Article

Cyclin: A protein specified by maternal mRNA in sea urchin eggs that is destroyed at each cleavage division

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Abstract

Cleavage in embryos of the sea urchin Arbacia punctulata consists of eight very rapid divisions that require continual protein synthesis to sustain them. This synthesis is programmed by stored maternal mRNAs, which code for three or four particularly abundant proteins whose synthesis is barely if at all detectable in the unfertilized egg. One of these proteins is destroyed every time the cells divide. Eggs of the sea urchin Lytechinus pictus and oocytes of the surf clam Spisula solidissima also contain proteins that only start to be made after fertilization and are destroyed at certain points in the cell division cycle. We propose to call these proteins the cyclins.

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... Fertilization thus triggers entry into the S-phase and completion of the fi rst mitotic division. Thanks to the large number of cells that can be recovered from a single female and their embryonic mitotic division synchronicity, these gametes have been crucial for the development of biochemical approaches studying cell cycle progression and protein translation (Evans et al. 1983;Humphreys 1969 ). Unravelling the mechanisms controlling protein synthe sis has been a central area of research in the 20th century (Thieffry and Burian 1996 ). ...
... For this purpose, he studied the sea urchin Arbacia punctulata. Adding [ 35 S] methionine to an egg suspension and separating proteins by gel electropho resis, he produced an autoradiogram where one specifi c band, which was later identified as a Cyclin, showed an unexpected behaviour (Evans et al. 1983). Whereas most bands became stronger and stronger with time, this protein accumulated after fertilization but disappeared rapidly just before blasto mere cleavage. ...
Chapter
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Echinoderms, a sister group of chordates, is a group of exclusively marine animals. The Echinodermata is an ancient phylum dating to at least 450 million years old and including more than 10,000 extant species present throughout the world’s oceans. The modern echinoderms belong to five classes: Echinoidea (e.g., sea urchins and sand dollars), Asteroidea (e.g., starfishes), Ophiuroidea (e.g., brittle stars), Holothuroidea (e.g., sea cucumbers) and Crinoidea (e.g., sea lilies and feather stars). Sea urchin represents a well-established marine model in biological sciences. This chapter provides a general description of echinoderms and focuses on the significant advances in cell and developmental biology that the study of sea urchins has made possible. During the last decade, many genomic data concerning echinoderms and sea urchins in particular have become available. These new molecular tools have facilitated gene regulation analysis during development and have boosted the possibilities offered by sea urchins as experimental models.
... While Leland Hartwell and Paul Nurse made many discoveries using yeast, Tim Hunt examined protein synthesis using fertilized sea urchin eggs. Hunt and his colleagues analyzed the pattern of protein synthesis before and after fertilization using a two-dimensional gel separation technique and found that the pattern of expressed proteins was changed [33]. They first noticed that one of these proteins is destroyed every time the cells divide. ...
... Then, they performed protein analysis experiments using 35 S-methionine-added egg suspensions and discovered that some proteins start to be synthesized after fertilization and are destroyed at certain points in the cell division cycle. They examined this protein in detail and proposed to call these proteins "cyclins" [33]. Importantly, they also showed data indicating the existence of multiple cyclins and described two cyclins, cyclin A and cyclin B, discovered from the clam Spisula solidissima in this paper. ...
Article
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Cell division and cell cycle mechanism has been studied for 70 years. This research has revealed that the cell cycle is regulated by many factors, including cyclins and cyclin-dependent kinases (CDKs). Heat shock transcription factors (HSFs) have been noted as critical proteins for cell survival against various stresses; however, recent studies suggest that HSFs also have important roles in cell cycle regulation-independent cell-protective functions. During cell cycle progression, HSF1, and HSF2 bind to condensed chromatin to provide immediate precise gene expression after cell division. This review focuses on the function of these HSFs in cell cycle progression, cell cycle arrest, gene bookmarking, mitosis and meiosis.
... Genetic analysis of Drosophila demonstrated that three distinct cyclins (B, A, and B3) make overlapping contributions to mitosis [11,12]. The requirement of mitotic cyclin synthesis for the first mitotic division following fertilization was first inferred from the inhibition of total protein synthesis in clams and sea urchins [13]. Cyclin B mRNA translation depends on mTOR activity, inducing eIF4E release from its inhibitor 4E-BP following sea urchin egg fertilization [14]. ...
... This is coherent with previous results showing that eIF4B is required in mammals to efficiently translate several mRNAs involved in cell proliferation, such as those coding for Cdc25C, c-Myc, and the ornithine decarboxylase [20]. Moreover, many cell cycle regulators are also translationally activated upon egg fertilization in the sea urchin, including the three mitotic cyclins (A, B, and B3) and CDK1 [4,13,14]. Different mechanisms that underlie the translation control of mitotic player during the cell cycle and development transitions of model organisms have been reported [49]. For instance, eIF4A influences the polysomal association and protein level of the B-type cyclin (Cdc13) in S. pombe [50]. ...
Article
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During the first steps of sea urchin development, fertilization elicits a marked increase in protein synthesis essential for subsequent cell divisions. While the translation of mitotic cyclin mRNAs is crucial, we hypothesized that additional mRNAs must be translated to finely regulate the onset into mitosis. One of the maternal mRNAs recruited onto active polysomes at this stage codes for the initiation factor eIF4B. Here, we show that the sea urchin eIF4B orthologs present the four specific domains essential for eIF4B function and that Paracentrotus lividus eIF4B copurifies with eIF4E in a heterologous system. In addition, we investigated the role of eIF4B mRNA de novo translation during the two first embryonic divisions of two species, P. lividus and Sphaerechinus granularis. Our results show that injection of a morpholino directed against eIF4B mRNA results in a downregulation of translational activity and delays cell division in these two echinoids. Conversely, injection of an mRNA encoding for P. lividus eIF4B stimulates translation and significantly accelerates cleavage rates. Taken together, our findings suggest that eIF4B mRNA de novo translation participates in a conserved regulatory loop that contributes to orchestrating protein synthesis and modulates cell division rhythm during early sea urchin development.
... Cyclin Y (CCNY), a cyclin family protein known to regulate the cell cycle in dividing cells [20][21][22], can regulate synaptic functions in terminally differentiated neuronal cells [23][24][25][26]. Specifically, CCNY is particularly enriched in the postsynaptic compartment in rat brains and inhibits the trafficking of α-amino-3-hydroxy-5methyl-4-isoxazolepropionate (AMPA) receptor (AMPAR) to the synapse during glycine-induced LTP in cultured neurons [25]. ...
Article
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Spatial learning and memory flexibility are known to require long-term potentiation (LTP) and long-term depression (LTD), respectively, on a cellular basis. We previously showed that cyclin Y (CCNY), a synapse-remodeling cyclin, is a novel actin-binding protein and an inhibitory regulator of functional and structural LTP in vitro. In this study, we report that Ccny knockout (KO) mice exhibit enhanced LTP and weak LTD at Schaffer collateral-CA1 synapses in the hippocampus. In accordance with enhanced LTP, Ccny KO mice showed improved spatial learning and memory. However, although previous studies reported that normal LTD is necessary for memory flexibility, Ccny KO mice intriguingly showed improved memory flexibility, suggesting that weak LTD could exert memory flexibility when combined with enhanced LTP. At the molecular level, CCNY modulated spatial learning and memory flexibility by distinctively affecting the cofilin-actin signaling pathway in the hippocampus. Specifically, CCNY inhibited cofilin activation by original learning, but reversed such inhibition by reversal learning. Furthermore, viral-mediated overexpression of a phosphomimetic cofilin-S3E in hippocampal CA1 regions enhanced LTP, weakened LTD, and improved spatial learning and memory flexibility, thus mirroring the phenotype of Ccny KO mice. In contrast, the overexpression of a non-phosphorylatable cofilin-S3A in hippocampal CA1 regions of Ccny KO mice reversed the synaptic plasticity, spatial learning, and memory flexibility phenotypes observed in Ccny KO mice. Altogether, our findings demonstrate that LTP and LTD cooperatively regulate memory flexibility. Moreover, CCNY suppresses LTP while facilitating LTD in the hippocampus and negatively regulates spatial learning and memory flexibility through the control of cofilin-actin signaling, proposing CCNY as a learning regulator modulating both memorizing and forgetting processes.
... A final example, this time of destabilization, is the selective degradation of proteins during the cell cycle, another discovery rewarded by the Nobel Prize (Evans et al. 1983). Cyclins, so called because their abundance cycles through the different stages of the cell cycle, are regulatory proteins that bind to and activate cyclin-dependent kinases (CDKs), which phosphorylate downstream targets to drive cells through the cell cycle. ...
Article
While cellular proteins were initially thought to be stable, research over the last decades has firmly established that intracellular protein degradation is an active and highly regulated process: Lysosomal, proteasomal, and mitochondrial degradation systems were identified and found to be involved in a staggering number of biological functions. Here, we provide a global overview of the diverse roles of cellular protein degradation using seven categories: homeostasis, regulation, quality control, stoichiometry control, proteome remodeling, immune surveillance, and baseline turnover. Using selected examples, we outline how proteins are degraded and why this is functionally relevant. Expected final online publication date for the Annual Review of Cell and Developmental Biology Volume 38 is October 2022. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
... Basic research on the cell cycle of primitive eukaryotic model systems, sea urchins, enabled scientists to identify a diverse family of proteins named cyclins (Evans et al. 1983). Unlike cell cycle regulatory kinases, cyclins display Fig. 2 Schematic depiction of the structure and activation process of CDKs. ...
Article
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The discussion on cell proliferation cannot be continued without taking a look at the cell cycle regulatory machinery. Cyclin-dependent kinases (CDKs), cyclins, and CDK inhibitors (CKIs) are valuable members of this system and their equilibrium guarantees the proper progression of the cell cycle. As expected, any dysregulation in the expression or function of these components can provide a platform for excessive cell proliferation leading to tumorigenesis. The high frequency of CDK abnormalities in human cancers, together with their druggable structure has raised the possibility that perhaps designing a series of inhibitors targeting CDKs might be advantageous for restricting the survival of tumor cells; however, their application has faced a serious concern, since these groups of serine–threonine kinases possess non-canonical functions as well. In the present review, we aimed to take a look at the biology of CDKs and then magnify their contribution to tumorigenesis. Then, by arguing the bright and dark aspects of CDK inhibition in the treatment of human cancers, we intend to reach a consensus on the application of these inhibitors in clinical settings.
... To ensure a correct DNA-duplication and distribution of chromosomes to the daughter cells, the cell cycle is controlled by a complex interaction of different intra-and extracellular factors with proliferation-promoting or -inhibiting effects. The correct sequence of cell cycle phases is regulated by the cyclic activity of cyclin-dependent kinases (Cdk) and their binding to regulatory cyclins (Figure 1) (Evans et al., 1983;King et al., 1996;Miller and Cross, 2001). Before forming of the pre-replication complex (pre-RC) at the origin of replication during the initiation step of DNA replication, the cyclin activity has to be low in G1-phase. ...
Article
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To sustain genomic stability by correct DNA replication and mitosis, cell cycle progression is tightly controlled by the cyclic activity of cyclin-dependent kinases, their binding to cyclins in the respective phase and the regulation of cyclin levels by ubiquitin-dependent proteolysis. The spindle assembly checkpoint plays an important role at the metaphase-anaphase transition to ensure a correct separation of sister chromatids before cytokinesis and to initiate mitotic exit, as an incorrect chromosome distribution may lead to genetically unstable cells and tumorigenesis. The ubiquitin ligase anaphase-promoting complex or cyclosome (APC/C) is essential for these processes by mediating the proteasomal destruction of cyclins and other important cell cycle regulators. To this end, it interacts with the two regulatory subunits Cdh1 and Cdc20. Both play a role in tumorigenesis with Cdh1 being a tumor suppressor and Cdc20 an oncogene. In this review, we summarize the current knowledge about the APC/C-regulators Cdh1 and Cdc20 in tumorigenesis and potential targeted therapeutic approaches.
... De novo protein synthesis is dispensable for the S--phase but is required for the onset of M--phase and subsequent embryonic cell cycles [64,65]. The mitotic cyclins A and B were first discovered in sea urchin as key proteins, which are synthesized and degraded during the mitosis phase at each cell division [14]. ...
... Most notably, L. pictus has a relatively short generation time (4-6 months), that enables breeding in captivity (Hinegardner, 1969;Nesbit et al., 2019;Nesbit and Hamdoun, 2020), and a recently published genome (Warner et al., 2021), which together open the door to targeted stable mutagenesis. The contributions made using this species range from the seminal discovery of cyclins (Evans et al., 1983) to the first characterizations of echinoderm cisregulatory elements (Xiang et al., 1991), the cytoskeletal controls of cell division (Pal et al., 2022) and axis formation (Henson et al., 2021), and modeling embryonic adaptations against ocean pollution and acidification (Cserjesi et al., 1992;Wu et al., 2015;Smith et al., 2019). ...
Article
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Sea urchins are premier model organisms for the study of early development. However, the lengthy generation times of commonly used species have precluded application of stable genetic approaches. Here, we use the painted sea urchin Lytechinus pictus to address this limitation and to generate a homozygous mutant sea urchin line. L. pictus has one of the shortest generation times of any currently used sea urchin. We leveraged this advantage to generate a knockout mutant of the sea urchin homolog of the drug transporter ABCB1, a major player in xenobiotic disposition for all animals. Using CRISPR/Cas9, we generated large fragment deletions of ABCB1 and used these readily detected deletions to rapidly genotype and breed mutant animals to homozygosity in the F2 generation. The knockout larvae are produced according to expected Mendelian distribution, exhibit reduced xenobiotic efflux activity and can be grown to maturity. This study represents a major step towards more sophisticated genetic manipulation of the sea urchin and the establishment of reproducible sea urchin animal resources.
... Marine genomics, transcriptomics, and beyond in developmental, cell, and evolutionary biology Marine organisms, serving as superior materials in the life science fields, have provided us with the opportunity to obtain biological knowledge. For example, in developmental biology, the countless gametes of sea urchins and the synchronous development of their embryos helped scientists in finding the cell-cycle-specific protein cyclin (Evans et al., 1983). In neuroscience, experiments using sea slugs provided evidence of the sophisticated neural circuit for memory (Kandel, 1976(Kandel, , 2001. ...
... Cell cycle regulation depends on tight cyclin expression control to activate the cyclin-dependent protein kinase enzymes (CDK) that govern cell cycle progression (Evans et al, 1983). In G1 phase, growth factors induce cyclin D which activates CDK4 and CDK6, which in turn, inactivate the E2F inhibitor, the retinoblastoma protein (RB). ...
Article
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Cyclin E/CDK2 drives cell cycle progression from G1 to S phase. Despite the toxicity of cyclin E overproduction in mammalian cells, the cyclin E gene is overexpressed in some cancers. To further understand how cells can tolerate high cyclin E, we characterized non-transformed epithelial cells subjected to chronic cyclin E overproduction. Cells overproducing cyclin E, but not cyclins D or A, briefly experienced truncated G1 phases followed by a transient period of DNA replication origin underlicensing, replication stress, and impaired proliferation. Individual cells displayed substantial intercellular heterogeneity in cell cycle dynamics and CDK activity. Each phenotype improved rapidly despite high cyclin E–associated activity. Transcriptome analysis revealed adapted cells down-regulated a cohort of G1-regulated genes. Withdrawing cyclin E from adapted cells only partially reversed underlicensing indicating that adaptation is at least partly non-genetic. This study provides evidence that mammalian cyclin E/CDK inhibits origin licensing indirectly through premature S phase onset and provides mechanistic insight into the relationship between CDKs and licensing. It serves as an example of oncogene adaptation that may recapitulate molecular changes during tumorigenesis.
... In 1982, he was surprised to observe proteins that were newly synthesised before each division but, most importantly, that were immediately and totally degraded after each cleavage, and then resynthesised again, etc. Because of its cycling appearance, he called this protein 'cyclin' [33]. He knew he had discovered something important for the cell cycle field but he was at a bit of a loss as to how it was working (from Woods Hole Lab, Tim Hunt's personal letter to his friend Richard Jackson). ...
Article
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All living organisms on Earth are made up of cells, which are the functional unit of life. Eukaryotic organisms can consist of a single cell (unicellular) or a group of either identical or different cells (multicellular). Biologists have always been fascinated by how a single cell, such as an egg, can give rise to an entire organism, such as the human body, composed of billions of cells, including hundreds of different cell types. This is made possible by cell division, whereby a single cell divides to form two cells. During a symmetric cell division, a mother cell produces two daughter cells, while an asymmetric cell division results in a mother and a daughter cell that have different fates (different morphologies, cellular compositions, replicative potentials, and/or capacities to differentiate). In biology, the cell cycle refers to the sequence of events that a cell must go through in order to divide. These events, which always occur in the same order, define the different stages of the cell cycle: G1, S, G2, and M. What is fascinating about the cell cycle is its universality, and the main reason for this is that the genetic information of the cell is encoded by exactly the same molecular entity with exactly the same structure: the DNA double helix. Since both daughter cells always inherit their genetic information from their parent cell, the underlying fundamentals of the cell cycle—DNA replication and chromosome segregation—are shared by all organisms. This review goes back in time to provide a historical summary of the main discoveries that led to the current understanding of how cells divide and how cell division is regulated to remain highly reproducible.
... Eukaryotes (Hartwell et al., 1974;Evans et al., 1983), while death relies on a constant probability rate. As a consequence, cells can adjust their size and growth, whereas genotypes encode how heterogeneous these sizes are as well as decide how cells synchronise nutrient processing with growth. ...
Thesis
Since Life was born, its Evolution has created an exceptional diversity of entities spanning an extravagant range of sizes from tiny microscopic molecules to the giant organisms that embody Megafauna. This broad variability, which exists both between and within classes of biological entities (eg. proteins), has often been theoretically explained by assuming the existence of biological trade-offs – impossibility to optimise many traits at once - and/or specific niches (eg. two different nutrients in the environment). However, how these trade-offs build up at the cellular level has mostly remained elusive because models of specialisation overlook the very mechanistic underpinnings of cells, that is to say how they actually work. Here, we develop a model where the fitness of cells emerges from a sequence of enzyme-substrate reactions that each produce a specific metabolite like ATP, and first show that accounting for physical, ecological and cellular constraints sheds light on the reasons why enzyme properties resemble a zoo although they seemingly evolve under a similar directional selective pressure – and should thus, at first glance, all look the same. Based on these landscapes of metabolic fitness and adaptive dynamics, we then simulate cell competition to demonstrate how the simple and intrinsic physical constraint of membrane permeability can explain the emergence of cross-feeding in an environment where only one ecological niche seems to exist, thus violating Gause's principle of competitive exclusion. This form of specialization sees one of the types specializing at the exploitation of a waste product released by the other, and it has generally been explained through considerations on the cost of processing metabolites, but this does not allow one to explain why certain metabolites seem more often associated with cross-feeding (acetate, glycerol). Our model specifically makes it possible to predict which intermediate metabolites should give rise to cross-feeding interactions and we emphasize that the available data seems to match our predictions. Yet, in this model, the enzymatic properties cannot evolve and the optimization simply concerns their levels of expression. If enzyme kinetics could be easily improved, cross-feeding would probably not emerge, but this not what the data shows. Hence, we then develop a quantitative genetic model intended to clarify the mechanistic underpinnings of metabolic epistasis and its consequences on the fitness reached at the mutation-selection-drift equilibrium. Because of these consequences, optimising enzymes above a given level may be compromised. Finally, we discuss the open perspectives whose vocation would be to combine these approaches in order to bring the fields of systems biology closer to those of quantitative genetics, and, thereby, to feed the field of quantitative evolution.
... Eukaryotes (Hartwell et al., 1974;Evans et al., 1983), while death relies on a constant probability rate. As a consequence, cells can adjust their size and growth, whereas genotypes encode how heterogeneous these sizes are as well as decide how cells synchronise nutrient processing with growth. ...
Thesis
Full-text available
Since Life was born, its tireless evolution has created an exceptional diversity of entities spanning an extravagant range of sizes, from the microscopic molecules underlying heritability and the expression of phenotypes to multicellular organisms and their societies. This great variety of the living world, present both between classes of biological entities and within these classes (e.g. proteins), has often been explained theoretically by assuming the existence of trade-offs – impossibility to optimize multiple traits at once – and / or specific niches as produced by the co-occurrence of multiple nutrients. However, the way in which these internal compromises emerge at the cellular level has remained in general elusive, especially since models of evolution most often overlook the mechanistic foundations and the very functioning of cells. Across this thesis, I try to build mechanistic evolutionary models by studying one of the most fundamental property of living things: how to produce energy, and grow, faster than others? This property is based at the cellular level on the structure and expression of enzymes. Rather than the extreme optimization this role suggests, enzymes have extremely diverse characteristics – some are close to achievable physical limits while others are very far from them – that should be explained. Through a modeling approach of the kinetic processes involved, I have shown that these differences can be explained by different selective contexts, characterizing in particular the reactions in which these enzymes are involved. Furthermore, the expression of an enzyme is the result of a complex selective process involving the obvious interest of catalyzing a given reaction but also overall costs for the cell, both in terms of production of the enzyme and of crowding within the cytoplasm. These constraints can promote the evolution of a selective (partial) expression of a metabolic pathway, leading to the release into the medium of metabolites, which can be used as an energetic source. In turn,this can give rise to the evolution of organisms specialised at these metabolites through a process called cross-feeding. Taking into account these processes in an adaptive dynamic model while also integrating an ecological dimension allowed me to establish the restricted conditions in which the cross-feeding may evolve, shedding light on the preponderant implication of certain metabolites (acetate, glycerol). In a last part, outside the strictly mechanistic framework of the thesis, I develop a model of population genetics intended to clarify the mainsprings of metabolic (weakest link) epistasis and its deleterious consequences on fitness at the mutation-selection-drift equilibrium. Finally, I discuss the perspectives opened up by this whole work, the vocation of which would be to contribute to the development of more realistic genotype- phenotype-fitness maps and to document their quantitative influence on evolution, through the combination of population genetics and systems biology.
... Since the cell cyclins have been found in sea urchins by Hunt in the 1980s [9], tremendous advances have been made in the molecular mechanisms of the cell cycle. This provides a positive direction for the study of tumors and other physiological diseases caused by cell cycle regulation [10]. ...
Article
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Research Highlights: This study identified the cell cycle genes in birch that likely play important roles during the plant’s growth and development. This analysis provides a basis for understanding the regulatory mechanism of various cell cycles in Betula pendula Roth. Background and Objectives: The cell cycle factors not only influence cell cycles progression together, but also regulate accretion, division, and differentiation of cells, and then regulate growth and development of the plant. In this study, we identified the putative cell cycle genes in the B. pendula genome, based on the annotated cell cycle genes in Arabidopsis thaliana (L.) Heynh. It can be used as a basis for further functional research. Materials and Methods: RNA-seq technology was used to determine the transcription abundance of all cell cycle genes in xylem, roots, leaves, and floral tissues. Results: We identified 59 cell cycle gene models in the genome of B. pendula, with 17 highly expression genes among them. These genes were BpCDKA.1, BpCDKB1.1, BpCDKB2.1, BpCKS1.2, BpCYCB1.1, BpCYCB1.2, BpCYCB2.1, BpCYCD3.1, BpCYCD3.5, BpDEL1, BpDpa2, BpE2Fa, BpE2Fb, BpKRP1, BpKRP2, BpRb1, and BpWEE1. Conclusions: By combining phylogenetic analysis and tissue-specific expression data, we identified 17 core cell cycle genes in the Betulapendula genome.
... Since the cell cyclins have been found in sea urchins by Hunt in the 1980s [9], tremendous advances have been made in the molecular mechanisms of the cell cycle. This provides a positive direction for the study of tumors and other physiological diseases caused by cell cycle regulation [10]. ...
Preprint
Research Highlights: This study identified the cell cycle genes in birch that likely play important roles during plant growth and development. This analysis provides a basis for understanding the regulatory mechanism of various cell cycles in Betula pendula. Background and Objectives: The cell cycle factors not only influence cell cycle progression together, but also regulate accretion, division and differentiation of cells, and then regulate growth and development of plant. In this study, we identified the putative cell cycle genes in B. pendula genome, based on the annotated cell cycle genes in A. thaliana. It could serve as a foundation for further functional studies. Materials and Methods: The transcript abundance was determined for all the cell cycle genes in xylem, root, leaf and flower tissues using RNA-seq technology. Results: We identified 59 cell cycle gene models in the genome of B. pendula, 17 highly expression genes among them. These genes were BpCDKA.1, BpCDKB1.1, BpCDKB2.1, BpCKS1.2, BpCYCB1.1, BpCYCB1.2, BpCYCB2.1, BpCYCD3.1, BpCYCD3.5, BpDEL1, BpDpa2, BpE2Fa, BpE2Fb, BpKRP1, BpKRP2, BpRb1 and BpWEE1. Conclusions: We identified 17 core cell cycle genes in the genome of birch by combining phylogenetic analysis and tissue specific expression data.
... CDKs drive the events of the mammalian cell cycle and control the rhythm of mammalian cell cycle procession; besides, they also integrate extracellular and intracellular signals to ensure the fine coordination of cell cycle events (Morgan, 1997). CDKs function as cell cycle event drivers, which are completely dependent on the association with cyclins, being first found in sea urchin eggs by their cyclic oscillations during the cleavage division in the early 1980s (Evans et al., 1983). Oscillating synthesis of cyclins controls the stage-specific timing of CDK activity. ...
Article
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Precise regulation of cell cycle is essential for tissue homeostasis and development, while cell cycle dysregulation is associated with many human diseases including renal fibrosis, a common process of various chronic kidney diseases progressing to end-stage renal disease. Under normal physiological conditions, most of the renal cells are post-mitotic quiescent cells arrested in the G0 phase of cell cycle and renal cells turnover is very low. Injuries induced by toxins, hypoxia, and metabolic disorders can stimulate renal cells to enter the cell cycle, which is essential for kidney regeneration and renal function restoration. However, more severe or repeated injuries will lead to maladaptive repair, manifesting as cell cycle arrest or overproliferation of renal cells, both of which are closely related to renal fibrosis. Thus, cell cycle dysregulation of renal cells is a potential therapeutic target for the treatment of renal fibrosis. In this review, we focus on cell cycle regulation of renal cells in healthy and diseased kidney, discussing the role of cell cycle dysregulation of renal cells in renal fibrosis. Better understanding of the function of cell cycle dysregulation in renal fibrosis is essential for the development of therapeutics to halt renal fibrosis progression or promote regression.
... Hunt found that one protein is always destroyed each time the cells divide. This protein was named cyclin as the level of the protein vary periodically during the cell cycle [22]. In the following years, more cyclins were identified in various species by Hunt and other Cells 2021, 10, 3327 4 of 23 groups. ...
Article
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The cell cycle is the series of events that take place in a cell, which drives it to divide and produce two new daughter cells. The typical cell cycle in eukaryotes is composed of the following phases: G1, S, G2, and M phase. Cell cycle progression is mediated by cyclin-dependent kinases (Cdks) and their regulatory cyclin subunits. However, the driving force of cell cycle progression is growth factor-initiated signaling pathways that control the activity of various Cdk–cyclin complexes. While the mechanism underlying the role of growth factor signaling in G1 phase of cell cycle progression has been largely revealed due to early extensive research, little is known regarding the function and mechanism of growth factor signaling in regulating other phases of the cell cycle, including S, G2, and M phase. In this review, we briefly discuss the process of cell cycle progression through various phases, and we focus on the role of signaling pathways activated by growth factors and their receptor (mostly receptor tyrosine kinases) in regulating cell cycle progression through various phases.
... Although cell cycle in eukaryotes is far more complex than the bacterial one, its progression is also tightly regulated, involving multiple layers of regulation. Three sets of factors were identified as the key players in the coordination of the eukaryotic cell cycle: (a) a set of cell cycle-regulated proteins including cyclin-dependent kinases (Cdk)cyclin complexes and related kinases (Evans et al., 1983;Galderisi et al., 2003), (b) various metabolic enzymes and related metabolites, and (c) reactive-oxygen species (ROS) and cellular redox status (Diaz-Moralli et al., 2013). Cyclin-dependent kinases are the engine of sequential progression through the eukaryotic cell cycle. ...
Preprint
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Cell cycle regulation is of paramount importance for all forms of life. Here we report that a transcription factor and its viral homologs control cell division in Sulfolobales. The 58-aa-long RHH (ribbon-helix-helix) family protein (designated as aCcr1) is conserved in Crenarchaeota and diverse Sulfolobales viruses. aCcr1 is essential for cell viability of Saccharolobus islandicus REY15A (formally Sulfolobus islandicus REY15A) and exhibits a cyclic transcription pattern, with the highest level being reached during active cell division (D-phase), subsequent to the expression of CdvA, an archaea-specific cell division protein. Cells over-expressing cellular aCcr1 as well as the homologs encoded by large spindle-shaped viruses Acidianus two-tailed virus (ATV) and Sulfolobus monocaudavirus 3 (SMV3) show significant growth retardation and cell division failure, manifested as enlarged cells with multiple chromosomes. aCcr1 over-expression results in downregulation of over a dozen of genes including the cdvA gene. A conserved motif, aCcr1-box, located between the TATA-binding box and the translation initiation site in the promoters of 13 out of the 17 repressed genes, is critical for the aCcr1 binding. The aCcr1-box is present in the promoters of cdvA genes across Sulfolobales, suggesting that aCcr1-mediated cdvA repression is an evolutionarily conserved mechanism by which archaeal cells dictate cytokinesis progression, whereas their viruses take advantage of this mechanism to manipulate the host cell cycle.
... Interestingly, oscillations in the levels of key proteins have been observed in many biological phenomena (Novák and Tyson 2008), including the cell division cycle (Evans et al., 1983;Glotzer et al., 1991), circadian rhythm (Kume et al., 1999;Reppert and Weaver 2001), cellular stress response Geva-Zatorsky et al., 2006), and development (Hirata et al., 2002;Pourquié 2003). In cells, the oscillation of protein levels is achieved by coordinating of many factors, such as transcriptional activities, translation, post-translational control (e.g., phosphorylation), nuclear transport, and targeted protein degradation. ...
Article
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Generating robust, predictable perturbations in cellular protein levels will advance our understanding of protein function and enable the control of physiological outcomes in biotechnology applications. Timed periodic changes in protein levels play a critical role in the cell division cycle, cellular stress response, and development. Here we report the generation of robust protein level oscillations by controlling the protein degradation rate in the yeast Saccharomyces cerevisiae. Using a photo-sensitive degron and red fluorescent proteins as reporters, we show that under constitutive transcriptional induction, repeated triangular protein level oscillations as fast as 5-10 minute-scale can be generated by modulating the protein degradation rate. Consistent with oscillations generated though transcriptional control, we observed a continuous decrease in the magnitude of oscillations as the input modulation frequency increased, indicating low-pass filtering of input perturbation. By using two red fluorescent proteins with distinct maturation times, we show that the oscillations in protein level is largely unaffected by delays originating from functional protein formation. Our study demonstrates the potential for repeated control of protein levels by controlling the protein degradation rate without altering the transcription rate.
... This proliferative process includes a series of checkpoints that govern progression through the four primary cell cycle phases: G1 (gap 1), S (chromatin synthesis), G2 (gap 2), M (chromatin segregation and cell division). CDKs promote passage through these checkpoints and are activated by cyclin proteins whose abundance fluctuates in response to growth factor and mitogen signaling and throughout cell cycle phases [16]. The presence of cyclins (and other cell cycle factors) are also limited by ubiquitin-dependent proteolysis. ...
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Protein signaling networks are formed from diverse and inter-connected cell signaling pathways converging into webs of function and regulation. These signaling pathways both receive and conduct molecular messages, often by a series of post-translation modifications such as phosphorylation or through protein–protein interactions via intrinsic motifs. The mitogen activated protein kinases (MAPKs) are components of kinase cascades that transmit signals through phosphorylation. There are several MAPK subfamilies, and one subfamily is the stress-activated protein kinases, which in mammals is the p38 family. The p38 enzymes mediate a variety of cellular outcomes including DNA repair, cell survival/cell fate decisions, and cell cycle arrest. The cell cycle is itself a signaling system that precisely controls DNA replication, chromosome segregation, and cellular division. Another indispensable cell function influenced by the p38 stress response is programmed cell death (apoptosis). As the regulators of cell survival, the BCL2 family of proteins and their dynamics are exquisitely sensitive to cell stress. The BCL2 family forms a protein–protein interaction network divided into anti-apoptotic and pro-apoptotic members, and the balance of binding between these two sides determines cell survival. Here, we discuss the intersections among the p38 MAPK, cell cycle, and apoptosis signaling pathways.
... Echinoderm embryos, and the sea urchin in particular, have been models for cell and developmental biology for over a century (Briggs and Wessel, 2006), leading to fundamental discoveries that shaped our understanding of fertilization (Hertwig, 1875), cell differentiation (Driesch H., 1892;Hörstadius and Horstadius, 1973;Davidson, 2006;McClay, 2011), genetic inheritance (Boveri, 1902) and cell-cycle regulation (Evans et al., 1983), to name a few. Some of these discoveries were made possible by the optical clarity of sea urchin embryos and the ease with which they can be live-imaged: these characteristics allowed, for instance, the first observations of male and female pronuclear fusion during fertilization (Hertwig, 1875) and of microtubule spindles during cell division (Hertwig, 1875). ...
Preprint
Full-text available
Echinoderm embryos have been model systems for cell and developmental biology for over 150 years, in good part because of their optical clarity. Discoveries that shaped our understanding of fertilization, cell division and cell differentiation were only possible because of the transparency of sea urchin eggs and embryos, which allowed direct observations of intracellular structures. More recently, live imaging of sea urchin embryos, coupled with fluorescence microscopy, has proven pivotal to uncovering mechanisms of epithelial to mesenchymal transition, cell migration and gastrulation. However, live imaging has mainly been performed on sea urchin embryos, while echinoderms include numerous experimentally tractable species that present interesting variation in key aspects of morphogenesis, including differences in embryo compaction and mechanisms of blastula formation. The study of such variation would allow us not only to understand how tissues are formed in echinoderms, but also to identify which changes in cell shape, cell-matrix and cell-cell contact formation are more likely to result in evolution of new embryonic shapes. Here we argue that adapting live imaging techniques to more echinoderm species will be fundamental to exploit such an evolutionary approach to the study of morphogenesis, as it will allow measuring differences in dynamic cellular behaviors - such as changes in cell shape and cell adhesion - between species. We briefly review existing methods for live imaging of echinoderm embryos and describe in detail how we adapted those methods to allow long-term live imaging of several species, namely the sea urchin Lytechinus pictus and the sea stars Patiria miniata and Patiriella regularis . We outline procedures to successfully label, mount and image early embryos for 10-16 hours, from cleavage stages to early blastula. We show that data obtained with these methods allows 3D segmentation and tracking of individual cells over time, the first step to analyze how cell shape and cell contact differ among species. The methods presented here can be easily adopted by most cell and developmental biology laboratories and adapted to successfully image early embryos of additional species, therefore broadening our understanding of the evolution of morphogenesis.
... In the fourth research article, Limatola et al., studied the role of the vitelline layer (VL) of sea urchin (Lytechinus pictus) eggs in fertilization [4]. Sea urchins have been a model organism in cell cycle research and contributed to the discovery of cyclins [5]. In this study, the research team led by Italian scientist Santella attempted to partially disrupt the VL with a reducing agent, dithiothreitol (DTT), and then observed its effects on fertilization. ...
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To celebrate its 10th anniversary, the prestigious journal Cells launched a series of Special Issues in 2021 [...]
... Protein concentration in a cell is increased (up-regulated) or decreased (down-regulated) based on the encoded information and transcription rate. Cells control and monitor each stage of cell growth with the help of these proteins [20]. For example, the G2-checkpoint is regulated by the protein p53 (tumor protein), which either repairs the DNA damage before cell division or destroys the cell (apoptosis) if the damage is irreparable [21]. ...
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Iterative-Indirect-Immunofluorescence Imaging(4i) generates high-dimensional multichannel images representing spatial information of 40 proteins at single-cell resolution. To understand the underlying biological processes, dimensionality reduction methods with the conjunction of deep learning are widely used to compress such high-dimensional biological data into lower dimensional space by capturing the most salient features of the dataset. Applications of supervised deep learning methods are limited for such datasets because of biological noise and the scarcity of labels. In this study, we employed labeled free deep learning methods to extract the low-dimensional feature representations of high-dimensional multichannel fluorescence images to analyze the effect of 6 different cancer drugs on single cell nuclei. The quality of these representations is evaluated by measuring their capacity to capture the known variations on three different downstream tasks, namely cell cycle prediction, drug identification, and predict cellular transcription rate. We modified, compared, and evaluated SimCLR, BYOL, and SimSiam, state-of-the-art self-supervised contrastive methods to solve the problem of noisy labels. We also incorporated Vision-Transformer in SimSiam architecture (Tranformeese network) that improved the performance on the downstream tasks by $3$ to $5\%$ as compared to the Resnet based architecture. We concluded that Transformeese Network generates better representations than other methods. As the performance of these contrastive methods is strongly dependent on the augmentations, we performed extensive ablation studies to find the most suitable set of augmentations for our dataset. We also showed that through custom-engineered augmentations, the model could be forced to focus on either protein intensities or subcellular morphology. Through latent space learning, self-supervised methods cluster the perturbations in a subspace based on their impact on cells. Thereby, these methods can be exploited to analyze the large-scale drug assays to understand cellular trajectories and heterogeneities.
... Les CDK sont actives uniquement lorsqu'elles sont fixées aux cyclines ce qui permet la formation d'un complexe composé d'une sous-unité catalytique, la CDK, et une sous-unité régulatrice, la cycline. Contrairement au niveau d'expression des CDKs qui reste stable, celui des cyclines varie pendant le cycle cellulaire permettant ainsi la régulation de l'activité des CDKs durant les différentes phases (Evans et al., 1983). AMPK peut également phosphoryler directement p53 sur la Ser15 pour la stabiliser et induire l'arrêt du cycle cellulaire à la transition G1/S . ...
Thesis
L’érythropoïèse adulte est un processus complexe qui a lieu dans la moelle osseuse, il aboutit à la formation de globules rouges (GR) à partir des cellules souches hématopoïétiques. L’AMPK (AMP-activated protein kinase) est un complexe hétérotrimérique (αβγ) connu pour son rôle de régulateur du métabolisme énergétique cellulaire. L'implication de l’AMPK dans le maintien de la survie et de l’intégrité des globules rouges murins a été démontrée. En effet, les souris invalidées pour Ampk α1, β1 ou γ1 présentent une anémie hémolytique due à une anomalie de la déformabilité des globules rouges. Nous avons émis l’hypothèse que les altérations observées dans les érythrocytes déficients en AMPK pourraient se mettre en place au cours du processus de différenciation des érythroblastes. L’objectif de ce travail est donc d’étudier le rôle de l'AMPK au cours de l’érythropoïèse murine et humaine adulte. Chez la souris, nos résultats démontrent que l’absence de l’AMPK n’affecte ni la prolifération ni la survie ni la différenciation des érythroblastes Ampkα1-/-. De la même façon, l’activation de l’AMPK n’a pas d’effet sur les érythroblastes murins. Chez l’homme, nous avons montré par une approche shRNA que l’inhibition de l’expression de la sous-unité α1 de l’AMPK induit un ralentissement de la prolifération cellulaire et une anomalie de l’érythropoïèse révélée par une modification de l’expression des protéines membranaires à la surface des érythroblastes au cours de la différenciation. Nous avons également montré que l’AMPK est fortement activée dans les érythroblastes immatures (Pro-E –Baso-E) et que cette activation diminue dans les érythroblastes matures (Poly-E – Retic). Une activation de l’AMPK par des activateurs directs (GSK621 et 991) dans les érythroblastes immatures n’a pas d’effet. Par contre, le maintien de son activation par les activateurs directs dans les érythroblastes matures induit, un arrêt du cycle cellulaire en phase S, une induction de l’autophagie, une apoptose caspase dépendante conduisant à un arrêt de la différenciation au stade Baso-E. Ces résultats montrent l’importance de la diminution de l’activation de l’AMPK pour la survie et la différenciation des érythroblastes matures. L’AMPK est donc importante pour la différenciation des cellules érythroïdes humaines alors que chez la souris, elle est impliquée dans le fonctionnement du globule rouge. Notre travail illustre donc un nouveau point de divergence entre l’érythropoïèse murine et l’érythropoïèse humaine.
... Cyclin-dependent kinases are a protein family of serinethreonine kinases that were first discovered for their role in cell cycle regulation in budding yeast in the late 1970s. The breakthrough discovery of cdc2 [17][18][19][20][21] ushered in a new era of research, opening multiple avenues of inquiry directed at one central theme: the regulation of cell division. In the following decades, laboratories across the globe uncovered a wide array of molecular mechanisms that control the cell cycle and cell division phases in development, immunity, tissue and organ regeneration, and cancer. ...
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Glioma stem cells (GSCs) are thought to drive growth and therapy resistance in glioblastoma (GBM) by “hijacking” at least a subset of signaling pathways active in normal neural stem cells (NSCs). Though the origins of GSCs still remain elusive, uncovering the mechanisms of self-renewing division and cell differentiation in normal NSCs has shed light on their dysfunction in GSCs. However, the distinction between self-renewing division pathways utilized by NSC and GSC becomes critical when considering options for therapeutically targeting signaling pathways that are specifically active or altered in GSCs. It is well-established that cyclin-dependent kinases (CDKs) regulate the cell cycle, yet more recent studies have shown that CDKs also play important roles in the regulation of neuronal survival, metabolism, differentiation, and self-renewal. The intimate relationship between cell cycle regulation and the cellular programs that determine self-renewing division versus cell differentiation is only beginning to be understood, yet seems to suggest potential differential vulnerabilities in GSCs. In this timely review, we focus on the role of CDKs in regulating the self-renewal properties of normal NSCs and GSCs, highlighting novel opportunities to therapeutically target self-renewing signaling pathways specifically in GBM.
... Changes in cyclin concentration produce sequential activation or deactivation of the CDK catalytic partners, leading to the periodic development of cells through the cell cycle [5,6]. Following the discovery of the cyclin protein in sea urchin oocytes [7], more cyclins, CKIs, CDKs, and E2F transcription factors were identified in both plants and mammals [4,8,9]. Different cyclin-like genes were fully characterized, and a few cyclins (cyclins A, B, D, and E) seem to be key players in cell cycle regulation and reproduction [10]. ...
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Cyclin-like genes are primarily considered as cell cycle regulators and have shown to be crucial for insect growth, development, reproduction, and fertility. However, no research has been performed on the cyclin-like genes in the diamondback moth (Plutella xylostella). Here, we identified the 21 cyclin genes in the genome of P. xylostella and clustered them into four groups. Most cyclin genes showed a well-maintained gene structure and motif distribution within the same group. The putative promoter regions of cyclin genes contained several transcription binding factors related to reproduction, along with growth and development. Furthermore, 16 miRNAs were identified targeting the 13 cyclin genes. Transcriptome and quantitative real-time PCR (qRT-PCR)-based expression profiling of cyclin-like genes at different stages and tissues were evaluated, revealing that 16 out of 21 cyclin genes were highly expressed in reproductive tissues of adult females and males. The Cyclin B1 gene (PxCyc B1) was only expressed in the ovary of the adult female and selected for the subsequent analysis. RNAi-mediated suppression of PxCyc B1 interrupted the external genitalia and length of the ovariole of female adults. Furthermore, the egg-laying capacity and hatching rate were also significantly decreased by suppressing the PxCyc B1, indicating the importance of cyclin genes in the reproduction and fertility of P. xylostella. The current study explained the detailed genome-wide analysis of cyclin-like genes in P. xylostella, which provided a basis for subsequent research to assess the roles of cyclin genes in reproduction, and the cyclin gene may be considered an effective target site to control this pest.
... To further test whether dermal DCs and mature BMDCs are retained during the cell cycle, we examined the levels of cyclins over the course of differentiation. Cyclins control cell-cycle progression from G1, S, G2, into M phase by activating cyclin-dependent kinases (Cdks) at specific time points in the cell cycle (Evans et al., 1983). Immature BMDCs harbored considerable levels of the S phase cyclin A and the mitotic cyclin B1 (Fig. 3 c), which are required for G1-S phase transition, completion of S phase, and entry into mitosis (Nurse, 2000). ...
Article
Centrosomes play a crucial role during immune cell interactions and initiation of the immune response. In proliferating cells, centrosome numbers are tightly controlled and generally limited to one in G1 and two prior to mitosis. Defects in regulating centrosome numbers have been associated with cell transformation and tumorigenesis. Here, we report the emergence of extra centrosomes in leukocytes during immune activation. Upon antigen encounter, dendritic cells pass through incomplete mitosis and arrest in the subsequent G1 phase leading to tetraploid cells with accumulated centrosomes. In addition, cell stimulation increases expression of polo-like kinase 2, resulting in diploid cells with two centrosomes in G1-arrested cells. During cell migration, centrosomes tightly cluster and act as functional microtubule-organizing centers allowing for increased persistent locomotion along gradients of chemotactic cues. Moreover, dendritic cells with extra centrosomes display enhanced secretion of inflammatory cytokines and optimized T cell responses. Together, these results demonstrate a previously unappreciated role of extra centrosomes for regular cell and tissue homeostasis.
... Echinoderm embryos, and the sea urchin in particular, have been models for cell and developmental biology for over a century (Briggs and Wessel, 2006), leading to fundamental discoveries that shaped our understanding of fertilization (Hertwig, 1875), cell differentiation (Driesch, 1892;Hörstadius and Horstadius, 1973;Davidson, 2006;McClay, 2011), genetic inheritance (Boveri, 1902) and cell-cycle regulation (Evans et al., 1983), to name a few. Some of these discoveries were made possible by the optical clarity of sea urchin embryos and the ease with which they can be live-imaged: these characteristics allowed, for instance, the first observations of male and female pronuclear fusion during fertilization (Hertwig, 1875) and of microtubule spindles during cell division (Hertwig, 1875). ...
Article
Full-text available
Echinoderm embryos have been model systems for cell and developmental biology for over 150 years, in good part because of their optical clarity. Discoveries that shaped our understanding of fertilization, cell division and cell differentiation were only possible because of the transparency of sea urchin eggs and embryos, which allowed direct observations of intracellular structures. More recently, live imaging of sea urchin embryos, coupled with fluorescence microscopy, has proven pivotal to uncovering mechanisms of epithelial to mesenchymal transition, cell migration and gastrulation. However, live imaging has mainly been performed on sea urchin embryos, while echinoderms include numerous experimentally tractable species that present interesting variation in key aspects of morphogenesis, including differences in embryo compaction and mechanisms of blastula formation. The study of such variation would allow us not only to understand how tissues are formed in echinoderms, but also to identify which changes in cell shape, cell-matrix and cell-cell contact formation are more likely to result in evolution of new embryonic shapes. Here we argue that adapting live imaging techniques to more echinoderm species will be fundamental to exploit such an evolutionary approach to the study of morphogenesis, as it will allow measuring differences in dynamic cellular behaviors - such as changes in cell shape and cell adhesion - between species. We briefly review existing methods for live imaging of echinoderm embryos and describe in detail how we adapted those methods to allow long-term live imaging of several species, namely the sea urchin Lytechinus pictus and the sea stars Patiria miniata and Patiriella regularis . We outline procedures to successfully label, mount and image early embryos for 10–16 h, from cleavage stages to early blastula. We show that data obtained with these methods allows 3D segmentation and tracking of individual cells over time, the first step to analyze how cell shape and cell contact differ among species. The methods presented here can be easily adopted by most cell and developmental biology laboratories and adapted to successfully image early embryos of additional species, therefore broadening our understanding of the evolution of morphogenesis.
Article
In the last 20 years, a growing army of systems biologists has employed quantitative experimental methods and theoretical tools of data analysis and mathematical modeling to unravel the molecular details of biological control systems with novel studies of biochemical clocks, cellular decision-making, and signaling networks in time and space. Few people know that one of the roots of this new paradigm in cell biology can be traced to a serendipitous discovery by an obscure Russian biochemist, Boris Belousov, who was studying the oxidation of citric acid. The story is told here from an historical perspective, tracing its meandering path through glycolytic oscillations, cAMP signaling, and frog egg development. The connections among these diverse themes are drawn out by simple mathematical models (nonlinear differential equations) that share common structures and properties.
Chapter
In this case study we examine mathematical models of the molecular regulatory network controlling entry into and exit from mitosis in fertilized frog eggs and in frog-egg extracts. This subject is important because mitotic cell division is at the root of all aspects of organismal growth, development and repair. Furthermore, the molecular mechanisms of mitotic regulation display intriguing commonalities across all taxa of eukaryotes that have been studied in detail. Finally, the physiology and molecular biology of mitotic cycle regulation in frog eggs can be adequately addressed in much simpler terms than, say, the complex regulatory networks in mammalian somatic cells. Nonetheless, what we learn from our investigations of frog eggs reveals some basic control principles of the DNA replication-division cycle in cells of more direct relevance to human health and disease.
Article
Cell cycle regulation that plays a pivotal role during organism growth and development is primarily driven by cyclin-dependent kinases (CDKs) and Cyclins. Although CDK and Cyclin genes have been characterized in some animals, the studies of CDK and Cyclin families in molluscs, the ancient bilaterian groups with high morphological diversity, is still in its infancy. In this study, we identified and characterized 95 CDK genes and 114 Cyclin genes in seven representative species of molluscs, including Octopus bimaculoides, Pomacea canaliculata, Biomphalaria glabrata, Lottia gigantea, Mizuhopecten yessoensis, Crassostrea gigas and Aplysia californica. Genes in CDK and Cyclin families were grouped into eight and 15 subfamilies by phylogenetic analysis, respectively. It should be noted that duplication of CDK9 gene was detected in P. canaliculate, L. gigantea and M. yessoensis genomes, which has never been recorded in animals. It is speculated that duplication may be the main course of expansion of the CDK9 subfamily in the three molluscs, which also sheds new light on the function of CDK9. In addition, Cyclin B is the largest subfamily among the Cyclin family in the seven molluscs, with the average of three genes. Our findings are helpful in better understanding CDK and Cyclin function and evolution in molluscs.
Article
Trichomonas vaginalis is an early divergent protozoan parasite that causes trichomoniasis, the most common non-viral sexually transmitted infection. In metazoans, there is abundant and detailed research on the cell cycle and the components involved in the regulation mechanisms. Regulators such as the cyclin-dependent kinases (CDKs) and cyclins activate the highly regulated processes of cell division. While CDKs have important roles in the phosphorylation of specific substrates, cyclins are important activating-components of CDKs that allow orderly passage through the different stages of the cell cycle. Cell cycle cyclins are characterized by showing drastic changes in their concentration during the cell cycle progression. However, in protists such as T. vaginalis, some biological processes such as cell cycle regulation remain less well studied. In an attempt to gain insight into cell cycle regulation in T. vaginalis, as an initial approach we characterized four proteins with features of cyclins. The genes encoding these putative cyclins were cloned to produce the recombinant proteins TvCYC1, TvCYC2, TvCYC3, and TvCYC4. The functional activity of TvCYC2, TvCYC3, and TvCYC4 was assessed through their complementation of a yeast cln1,2,3Δ mutant strain; TvCYC1 was not able to complement this mutant. Furthermore, our results suggest that TvCYC1, TvCYC2, and TvCYC3, are able to interact with and activate the kinase activity of TvCRK1, a kinase previously characterized by our group. The present study represents the first characterization of cyclins potentially involved in cell cycle regulation in T. vaginalis.
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Genetic feedback loops can be used by cells as a means to regulate internal processes or keep track of time. It is often thought that, for a genetic circuit to display self-sustained oscillations, a degree of cooperativity is needed in the binding and unbinding of actor species. This cooperativity is usually modeled using a Hill function, regardless of the actual promoter architecture. Moreover, genetic circuits do not operate in isolation and often transcription factors are shared between different promoters. In this work we show how mathematical modelling of genetic feedback loops can be facilitated with a mechanistic fold-change function that takes into account the titration effect caused by competing binding sites for transcription factors. The model shows how the titration effect aids self-sustained oscillations in a minimal genetic feedback loop: a gene that produces its own repressor directly — without cooperative transcription factor binding. The use of delay- differential equations leads to a stability contour that predicts whether a genetic feedback loop will show self-sustained oscillations, even when taking the bursty nature of transcription into account.
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Cyclin-dependent kinases (CDKs) control the progression of the cell cycle. D-type cyclin (CYCD) is generally believed to form a complex with CDK and control the G1/S transition. In plants, CYCD and CDK gene families can be divided into 6 (D1–D7) and 7 (CDKA–CDKG) subclasses, respectively. Different subclasses in the CYCD and CDK families have different numbers, structures and functions. In some heterologous woody plants, the functions of these subclass family members remain unclear. In this study, 43 CYCD and 27 CDK gene family members were identified in the allodiploid Populus tomentosa Carr. Phylogenetic analysis suggested that these CYCDs and CDKs were divided into 6 and 7 subclasses, respectively, which were the same as other species. The analysis of protein properties, gene structure, motifs, domains, cis-acting elements and tissue-specific expression of all members of these CYCDs and CDKs showed that the differences between members of different subclasses varied widely, but members of the same subclass especially in the CDK gene family were very similar. These findings also demonstrated a strong correlation between CYCD and CDK gene family members in response to hormones and specific expression. The collinear analysis of P. tomentosa, Populus trichocarpa and Arabidopsis thaliana showed that the expansion patterns of CYCD and CDK gene families were predominantly whole genome duplications (WGD). The protein interaction prediction results of different subclasses of CYCD and CDKs showed that the interaction between different subclasses of CYCD and CDKs was significantly different. Our previous study found that transgenic PtoCYCD2;1 and PtoCYCD3;3 poplars exhibited opposite phenotypes. Y2H and BIFC results showed that the interaction between PtoCYCD2;1 and PtoCYCD3;3 was significantly different with CDKs. This finding might suggest that the functional differences of different CYCD subclasses in plant growth and development were closely related to the different interactions between CYCD and CDK. Our results provide a good idea and direction for the functional study of CYCD and CDK proteins in woody plants.
Chapter
Cell homeostasis is maintained by a balanced regulation of cell growth, cell proliferation, and cell death. Several cell cycle checkpoints tightly regulate the division of cells. Defects in these checkpoints may lead to the accumulation of genomic abnormalities resulting in a pathological condition, such as cancer. Cancer is characterized by the uncontrolled cell proliferation due to various defects at the genetic and molecular levels, including the aberrant activity of several proteins involved in the cell cycle checkpoints regulation. Therefore, the cell cycle regulatory proteins are considered to be the most attractive targets in the cancer therapy. After extensive research of decades on the physiological functions and regulation of cell cycle proteins, several cell cycle regulators are being targeted for the cancer therapy. Here in this chapter, we will discuss the cell cycle, cell cycle regulators, and the significance of targeting them for the cancer therapeutics.
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Biological time keeping, or the duration and tempo at which biological processes occur, is a phenomenon that drives dynamic molecular and morphological changes that manifest throughout many facets of life. In some cases, the molecular mechanisms regulating the timing of biological transitions are driven by genetic oscillations, or periodic increases and decreases in expression of genes described collectively as a “molecular clock.” In vertebrate animals, molecular clocks play a crucial role in fundamental patterning and cell differentiation processes throughout development. For example, during early vertebrate embryogenesis, the segmentation clock regulates the patterning of the embryonic mesoderm into segmented blocks of tissue called somites, which later give rise to axial skeletal muscle and vertebrae. Segmentation clock oscillations are characterized by rapid cycles of mRNA and protein expression. For segmentation clock oscillations to persist, the transcript and protein molecules of clock genes must be short‐lived. Faithful, rhythmic, genetic oscillations are sustained by precise regulation at many levels, including post‐transcriptional regulation, and such mechanisms are essential for proper vertebrate development. This article is categorized under: RNA Export and Localization > RNA Localization RNA Turnover and Surveillance > Regulation of RNA Stability Translation > Regulation Genetic oscillators play crucial roles in development. The hairy/Enhancer of split gene family (Hes/her) undergo cell‐autonomous genetic oscillations to regulate the timing of vertebrate segmentation and neural cell proliferation and differentiation. mRNA stability and translation are tightly regulated to maintain oscillatory expression.
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Otto Warburg published the first papers describing what became known as the Warburg effect in 1923. All that was known about glucose metabolism at that time was that it occurred in two stages: (i) fermentation or glycolysis, in which glucose was converted to lactate, which did not require oxygen, and (ii) oxidative metabolism, in which the carbon atoms derived from glycolysis were fully oxidized to carbon dioxide, which did require oxygen. Warburg discovered that most tumour tissues produced a large amount of lactate that was reduced but not eliminated in the presence of oxygen, while most normal tissues produced a much smaller amount of lactate that was eliminated by provision of oxygen. These findings were clearly well ahead of their time because it was another 80 years before they were to have any major impact, and even today the mechanisms underlying the Warburg effect are not completely understood.
Chapter
The cell cycle is the series of events that take place in a cell that drives it to divide and produce two new daughter cells. Through more than 100 years of efforts by scientists, we now have a much clearer picture of cell cycle progression and its regulation. The typical cell cycle in eukaryotes is composed of the G1, S, G2, and M phases. The M phase is further divided into prophase, prometaphase, metaphase, anaphase, telophase, and cytokinesis. Cell cycle progression is mediated by cyclin-dependent kinases (Cdks) and their regulatory cyclin subunits. However, the driving force of cell cycle progression is growth factor-initiated signaling pathways that controls the activity of various Cdk-cyclin complexes. Most cellular events, including DNA duplication, gene transcription, protein translation, and post-translational modification of proteins, occur in a cell-cycle-dependent manner. To understand these cellular events and their underlying molecular mechanisms, it is desirable to have a population of cells that are traversing the cell cycle synchronously. This can be achieved through a process called cell synchronization. Many methods have been developed to synchronize cells to the various phases of the cell cycle. These methods could be classified into two groups: synchronization methods using chemical inhibitors and synchronization methods without using chemical inhibitors. All these methods have their own merits and shortcomings.Key wordsCell cycleG1 phaseS phaseG2 phaseM phaseSynchronizationCdks
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Evolutionary developmental biology, the interdisciplinary effort of illuminating the conserved similarities and difference during animal development across all phylogenetic clades, has gained renewed interest in the past decades. As technology (immunohistochemistry, next generation sequencing, advanced imaging, computational resources) has advanced, so has our ability of resolving fundamental hypotheses and overcoming the genotype‐phenotype gap. This rapid progress, however, has also exposed gaps in the collective knowledge around the choice and representation of model organisms. It has become clear that evo‐devo requires a comparative, large‐scale approach including marine invertebrates to resolve some of the most urgent questions about the phylogenetic positioning and character traits of the last common ancestors. Many invertebrates at the base of the tree of life inhabit marine environments and have been used for some years due to their accessibility, husbandry, and morphology. Here, we briefly review the major concepts of evolutionary developmental biology and discuss the suitability of established model organisms to address current research questions, before focussing on the importance, application and state‐of‐the‐art of marine evo‐devo. We highlight novel technical advances that progress evo‐devo as a whole.
Chapter
A “cell‐cycle engine” that functions in all eukaryotic organisms was defined. This mechanism consists of cyclin‐regulated cell‐cycle kinases that provide the crucial checkpoints and feedback controls that regulate cell division. This chapter utilizes T cells to illustrate the transition steps that regulate cell‐cycle entry, progression, and exit. The kinases and phosphatases that carry out this phase of cyclin‐dependent kinase regulation provide additional links between environmental signals and activation of the cell‐cycle machinery. The S‐phase checkpoint relies on detection of stalled replication or DNA damage. The S‐phase checkpoint is fallible, so a partially redundant checkpoint exists during the G2 phase. Cancer is a condition where cells divide uncontrollably, making it easy to see how it can be linked to abnormal cell‐cycle function. The gross abnormalities are prognostic, but even subtle quantitative differences in cyclin expression, Rb and phosphorylation, can influence tumor progression and serve as useful indicators of prognosis for hematopoietic tumors.
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For over 20 years, the Schmid Training Course (STC) has offered unique opportunities for marine biology students from European universities to learn about marine model organisms. While the topics of the course have continuously changed over the years with the advent of new research techniques and discoveries, the pedagogical approach has remained largely the same – a combination of lectures, lab practicals, and field excursions. Several life science researchers, who have taught in the STC for many years, sought to bring the course's pedagogical approach into the 21st century, and with the support of Erasmus+ Programme of the European Community funding, the Digital Marine project was developed. Digital Marine began in 2018 as an international partnership between the six research centers from which the STC instructors hail, and its main objective was to introduce a flipped, blended approach to learning and teaching with respect to established and emerging marine biological model systems. The Digital Marine platform, which covers 12 marine model organisms, is now publicly available. A blended learning for the Schmid Training Course, a training on the use of marine organisms in experimental biology for Master students. This course has now a dedicated online platform called Digital Marine designed to allow autonomous learning. Face‐to‐face learning in marine station offers more time for lab experiences and exercises.
Thesis
La protéine F-box FBL17 est un régulateur clef de la progression du cycle cellulaire chez Arabidopsis thaliana. Elle est notamment impliquée dans la dégradation des protéines inhibitrices de CDK, ce qui conduit chez le mutant fbl17 à des phénotypes développementaux sévères et une réduction drastique de l'activité de division cellulaire dans les méristèmes. Si l’implication de FBL17 dans le cycle cellulaire a été bien caractérisée, son répertoire de substrats et ses autres fonctions biologiques restent encore méconnus. Durant cette thèse, l’interactome de FBL17 a été mis en évidence et son implication dans la réponse aux dommages à l’ADN (DDR) a été étudiée. Le mutant fbl17 présente une DDR constitutive associée à une augmentation du nombre de lésions de l’ADN, et l’induction de gènes de la DDR de façon SOG1-indépendante. Au niveau protéique, en condition de stress génotoxique, FBL17 est recrutée au niveau des sites de lésions de l’ADN où elle co-localise avec la protéine RBR1. Ces données supportent un rôle de FBL17 dans le maintien de l’intégrité génétique des cellules.
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Cyclin‑dependent kinase (CDK)4/6 inhibitors in combination with endocrine therapy are the current standard of care used in the first‑line treatment of hormone receptor‑positive/HER2‑negative metastatic breast cancer (BC). Although CDK4/6 inhibitors mainly target the cell cycle, emerging evidence has indicated further potential roles of CDKs other than regulating cell cycle progression. The G1 and G2/M transition regulators, including cyclins D and E, as well as their catalytic partners, CDK2, CDK4 and CDK6, have been reported to play crucial roles in pluripotency maintenance and cell fate decisions of human pluripotent stem cells by controlling transcription factors, signaling pathways and epigenetic regulators. Dinaciclib, a CDK1/2/5/9 inhibitor, is currently being evaluated in clinical trials against various cancer types, including BC. However, the underlying molecular mechanisms of CDK1/2/5/9 inhibitors in regulating BC stemness remain poorly understood. The present study aimed to examine the stemness‑inhibitory effects of dinaciclib in MCF‑7 (luminal) and HCC‑1806 (triple‑negative) BC cells. We found that this drug not only effectively reduced the self‑renewal abilities and other malignant properties, but also dose‑dependently decreased the protein expression levels of three BC stem cell markers, CD44, aldehyde dehydrogenase 1 family member A1 (ALDH1A1) and BMI1 proto‑oncogene, polycomb ring finger (Bmi1), as well as three embryonic stem cell markers, Oct4, Nanog and Sox2. Moreover, the dinaciclib‑induced decrease of Oct4 and Nanog protein expression was able to be restored by co‑treatment with MG‑132, a proteasome inhibitor. Forkhead box M1 (FoxM1), both a stemness‑stimulating transcription factor and a cell cycle regulator, along with the Hedgehog signaling pathway, were identified as the therapeutic targets of dinaciclib. Collectively, the present results demonstrated a novel role of dinaciclib in suppressing BC stemness and indicated its potential use for future cancer treatments.
Article
Sea urchins have a long history as model organisms in biology, but their use in genetics is limited due to their long breeding cycle. In sea urchin genetics, genome editing technology was first established in Hemicentrotus pulcherrimus, whose genome has already been published. However, because this species also has a long breeding cycle, new model sea urchins that are more suitable for genetics have been sought. Here, we report a draft genome of another western Pacific species, Temnopleurus reevesii, which we established as a new model sea urchin recently since this species has a comparable developmental process to other model sea urchins but a short breeding cycle of approximately half a year. The genome of T. reevesii was assembled into 28,742 scaffold sequences with an N50 length of 67.6 kb and an estimated genome size of 905.9 Mb. In the assembled genome, 27,064 genes were identified, 23,624 of which were expressed in at least one of the seven developmental stages. To provide genetic information, we constructed the genome database TrBase. We also constructed the Western Pacific Sea Urchin Genome Database (WestPac‐SUGDB) with the aim of establishing a portal site for genetic information on sea urchins in the West Pacific. This site contains genomic information on two species, T. reevesii and H. pulcherrimus, and is equipped with homology search programs for comparing the two datasets. Therefore, TrBase and WestPac‐SUGDB are expected to contribute not only to genetic research using sea urchins but also to comparative genomics and evolutionary research.
Article
Patterns are ubiquitous in living systems and underlie the dynamic organization of cells, tissues, and embryos. Mathematical frameworks have been devised to account for the self-organization of biological patterns, most famously the Turing framework. Patterns can be defined in space, for example, to form stripes; in time, such as during oscillations; or both, to form traveling waves. The formation of these patterns can have different origins: purely chemical, purely mechanical, or a combination of the two. Beyond the variety of molecular implementations of such patterns, we emphasize the unitary principles associated with them, across scales in space and time, within a general mechanochemical framework. We illustrate where such mechanisms of pattern formation arise in biological systems from cellular to tissue scales, with an emphasis on morphogenesis. Our goal is to convey a picture of pattern formation that draws attention to the principles rather than solely specific molecular mechanisms. Expected final online publication date for the Annual Review of Cell and Developmental Biology Volume 38 is October 2022. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
Article
Many psychiatric disorders accompany deficits in cognitive functions and synaptic plasticity, and abnormal lipid modifications of neuronal proteins are associated with their pathophysiology. Lipid modifications, including palmitoylation and myristoylation, play crucial roles in the subcellular localization and trafficking of proteins. Cyclin Y (CCNY), enriched in the postsynaptic compartment, acts as an inhibitory modulator of functional and structural long-term potentiation (LTP) in the hippocampal neurons. However, cellular and molecular mechanisms underlying CCNY-mediated inhibitory functions in the synapse remain largely unknown. Here, we report that myristoylation located CCNY to the trans-Golgi network (TGN), and subsequent palmitoylation directed the myristoylated CCNY from the TGN to the synaptic cell surface. This myristoylation-dependent palmitoylation of CCNY was required for the inhibitory role of CCNY in excitatory synaptic transmission, activity-induced dynamics of AMPA receptors and PSD-95, LTP, and spatial learning. Furthermore, spatial learning significantly reduced palmitoyl- and myristoyl-CCNY levels, indicating that spatial learning lowers the synaptic abundance of CCNY. Our findings provide mechanistic insight into how CCNY is clustered adjacent to postsynaptic sites where it could play its inhibitory roles in synaptic plasticity and spatial learning.
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Rosmarinus officinalis .L ‫المعمل‬ ‫في‬ ‫المرستيمية‬ ‫البصل‬ ‫خاليا‬ ‫باستخدام‬ Allium cepa .L ‫البيدوني‬ ‫خالد‬ ‫د.‬ / ‫م‬ ‫د.‬ ‫ب‬ ‫جبريل‬ ‫روكة‬ / ‫القطعاني‬ ‫دالل‬ ‫أ.‬ / ‫أ.‬ ‫حمدي‬ ‫الفرجاني‬ ‫المعروفة‬ ‫النباتات‬ ‫من‬ ‫الجبل‬ ‫إكليل‬ ‫نبات‬ ‫من‬ ‫أهم‬ ‫أنحاء‬ ‫جميع‬ ‫في‬ ‫الشعبي‬ ‫الطب‬ ‫في‬ ‫بكثرة‬ ‫المستخدمة‬ ‫الطبية‬ ‫النباتات‬ ‫العالم‬ ، ‫الطبي‬ ‫تأثيرها‬ ‫يعزى‬ ‫حيث‬ ‫اإلكليل‬ ‫نبات‬ ‫مستخلص‬ ‫اء‬ ‫الحتو‬ ‫على‬ (‫حامض‬ Caffeic acid ‫ومشتقاته‬) ‫تأثير‬ ‫اسة‬ ‫در‬ ‫تم‬. ‫نبات‬ ‫اق‬ ‫ألور‬ ‫المائي‬ ‫المستخلص‬ ‫االكليل‬ (‫اكيز‬ ‫تر‬ (0.01,0.1 and 1 mg/ml ‫مختلفة(‬ ‫زمنية‬ ‫ات‬ ‫وفتر‬ ,6,4,2 ‫و‬ 24) ‫على‬ ‫االنقسام‬ ‫دليل‬ ‫باستخدام‬ ‫االنقسام‬ ‫اثناء‬ ‫الصبغيات‬ ‫وسلوك‬ ‫البصل‬ ‫لنبات‬ ‫الميتوزي‬ ‫االنقسام‬ ‫ات.‬ ‫الطفر‬ ‫ونسبة‬ ‫النتائج‬ ‫خالل‬ ‫من‬ ‫عليها‬ ‫المتحصل‬ ‫الميتوزي‬ ‫االنقسام‬ ‫إيقاف‬ ‫على‬ ‫القدرة‬ ‫لها‬ ‫كبات‬ ‫مر‬ ‫تحتوي‬ ‫انها‬ ‫االكليل‬ ‫نبات‬ ‫اق‬ ‫ألور‬ ‫المائي‬ ‫المستخلص‬ ‫ان‬ ‫الصبغيات‬ ‫سلوك‬ ‫على‬ ‫التأثير‬ ‫و‬. ‫منتظم،‬ ‫غير‬ ‫تمهيدي‬ ‫طور‬ ‫مثل‬ ‫ات‬ ‫الطفر‬ ‫من‬ ‫مختلفة‬ ‫مظاهر‬ ‫في‬ ‫اتضح‬ ‫المائي‬ ‫المستخلص‬ ‫تأثير‬ ‫الوسطي‬ ‫الطور‬ ‫الكروموسومات،‬ ‫التصاق‬ C ‫الكرمو‬ ‫و‬ ‫الجسور،‬ ، ‫المتأخر‬ ‫سوم‬ ‫الميوزي‬ ‫االنقسام‬ ‫خالل‬. ‫هذه‬ ‫بشكل‬ ‫تعتمد‬ ‫النتاج‬ ‫المعاملة.‬ ‫وزمن‬ ‫المستخدم‬ ‫كيز‬ ‫التر‬ ‫على‬ ‫كبير‬ ‫المقدمة‬ ‫ما‬ ‫عن‬ ‫البحث‬ ‫ته‬ ‫بفطر‬ ‫اإلنسان‬ ‫استطاع‬ ‫فقد‬ ، ‫ووجوده‬ ‫خلقه‬ ‫مع‬ ‫بدأ‬ ‫قد‬ ‫السامة‬ ‫و‬ ‫ية‬ ‫العطر‬ ‫و‬ ‫الطبية‬ ‫بالنباتات‬ ‫اإلنسان‬ ‫اهتمام‬ ‫النباتات‬ ‫باستخدام‬ ‫اضه‬ ‫أمر‬ ‫و‬ ‫آالمه‬ ‫يخفف‬ ‫اد‬ ‫بالمو‬ ‫تعرف‬ ‫فسيولوجي‬ ‫تأثير‬ ‫و‬ ‫فعالية‬ ‫ذات‬ ‫اد‬ ‫مو‬ ‫علي‬ ‫تحتوي‬ ‫التي‬ ‫و‬ ، ‫به‬ ‫المحيطة‬ ‫لتحضير‬ ‫الصيدلة‬ ‫في‬ ‫المستخدمة‬ ‫كبات‬ ‫المر‬ ‫أو‬ ‫اد‬ ‫المو‬ ‫علي‬ ‫منها‬ ‫نحصل‬ ‫التي‬ ‫النباتات‬ ‫أيضا‬ ‫الطبية‬ ‫النباتات‬ ‫تشمل‬ ‫كما‬ ، ‫الفعالة‬ ‫النبا‬ ‫و‬ ‫العقاقير‬ ‫من‬ ‫المئات‬ ‫هناك‬ ‫إن‬ ‫إال‬ ، ‫المختلفة‬ ‫بأشكالها‬ ‫األدوية‬ ‫الكثير‬ ‫و‬ ‫المختلفة‬ ‫اض‬ ‫األمر‬ ‫لعالج‬ ‫تستخدم‬ ‫التي‬ ‫الطبية‬ ‫تات‬ ‫كما‬ ، ‫تعاطيها‬ ‫ووقت‬ ‫الجرعة‬ ‫مقدار‬ ‫بها‬ ‫محدد‬ ‫طبية‬ ‫وصفة‬ ‫بدون‬ ‫استعمالها‬ ‫عدم‬ ‫الضروري‬ ‫و‬ ‫اجب‬ ‫الو‬ ‫من‬ ‫و‬ ‫السمية‬ ‫شديد‬ ‫منها‬ ‫استخدامها‬ ‫في‬ ‫الحيطة‬ ‫و‬ ‫الحذر‬ ‫اتخاذ‬ ‫عدم‬ ‫إن‬ ‫كبيرة‬ ‫بمخاطر‬ ‫مصحوبا‬ ‫عادة‬ ‫يكون‬ Effraim et al., 2001) .) ‫اآل‬ ‫في‬ ‫استخدام‬ ‫يادة‬ ‫ز‬ ‫لوحظ‬ ‫األخيرة‬ ‫ونة‬ ‫تحوير‬ ‫في‬ ‫اضح‬ ‫و‬ ‫دور‬ ‫من‬ ‫لها‬ ‫لما‬ ‫الشعبي‬ ‫الطب‬ ‫في‬ ‫الطبية‬ ‫النباتات‬ ‫مستخلصات‬ ‫الخاليا‬ ‫نشاط‬ ‫على‬ ‫الطبية‬ ‫للمستخلصات‬ ‫اثية‬ ‫الور‬ ‫المادة‬ ‫علي‬ ‫التأثير‬ ‫فة‬ ‫لمعر‬ ‫اسات‬ ‫الدر‬ ‫من‬ ‫العديد‬ ‫وقامت‬ ، ‫ات‬ ‫المطفر‬ ‫نشاط
Chapter
The cyclins and cyclin-dependent kinases (CDKs) are two large protein families that function in a wide variety of cellular contexts. Most notably, the cyclin-CDKs govern and orchestrate progression through the eukaryotic cell cycle. In a sequential fashion, different cyclin-CDK complexes execute the signaling, transcriptional, and enzymatic events that effect transitions through and between cell cycle phases. As such, cyclin-CDKs are intricately regulated through complex formation, phosphorylation, stability, subcellular localization, and association with CDK inhibitors. Given their connection to cellular proliferation, cyclin-CDKs are poised at the nexus of oncogenes and tumor suppressors and have a unique role in the pathogenesis of cancer and developmental disorders.
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Cell signalling engenders cells with the capability to receive and process information from the intracellular and extracellular environments, trigger and execute biological responses, and communicate with each other. Ultimately, cell signalling is responsible for maintaining homeostasis at the cellular, tissue and systemic level. For this reason, cell signalling is a topic of intense research efforts aimed to elucidate how cells coordinate transitions between states in developing and adult organisms in physiological and pathological conditions. Here, we review current knowledge of how cell signalling operates at multiple spatial and temporal scales, focusing on how single-cell analytical techniques reveal mechanisms underpinning cell-to-cell variability, signalling plasticity, and collective cellular responses.
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Micromolar amounts of the divalent ionophore A23187 can activate echinoderm eggs. The activations by ionophore A23187 were examined in terms of membrane elevation, the program of membrane conductance changes, the respiratory burst, and the increases in protein and DNA synthesis which normally accompany activation by sperm. In all these respects activation by the ionophore was fairly normal although subsequent cleavage and embryonic development was limited. Ionophore A23187 activations of the cortex of Lytechinus pictus and Strongylocentrotus purpuratus eggs were compared in various ionic media and were found to be completely independent of the ionic composition of the external solution. Respiration and protein synthesis of L. pictus eggs in singly substituted ionic media also indicated that these activations were independent of external sodium, calcium, or magnesium. These results suggest that the ionophore acts by releasing intracellular Ca⁺⁺. Consistent with this interpretation is the finding that eggs preloaded with ⁴⁵Ca show a 20-fold increase in ⁴⁵Ca-efflux when activated by ionophore A23187 or fertilization. Measurements of the “free” and “bound” calcium and magnesium in homogenates of the unfertilized eggs show that most of the Mg⁺⁺ is already available in the soluble form, whereas Ca⁺⁺ is sequestered but available for release. We propose that both normal fertilization and ionophore activation affect the metabolism of the egg by releasing Ca⁺⁺ sequestered in intracellular stores.
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Hybrid myeloma cell lines secreting monoclonal antibodies to tubulin have been prepared using rat myelomas and spleen cells from rats immunized with yeast tubulin. A comparison between the results obtained with the rat myeloma Y3-Ag 1.2.3., which secretes a light chain, and a new line, YB2/O, which does not, shows that they are both excellent parental lines and that the second produces hybrids with no myeloma chain components. The antitubulin antibodies in the serum of rats bearing two of the hybrid myeloma tumors gave titers of up to 1:10(6) from which large amounts of monoclonal antibodies could be easily purified. They recognized tubulin from yeast as well as from birds and mammals. The two antibodies gave clear immunofluorescent staining of yeast mitotic spindles as well as the interphase microtubule network of tissue culture cells. Some difference in the pattern of immunofluorescence staining of yeast cells and nuclei was observed between the two antibodies. The purified antibodies could be conjugated to colloidal gold particles and used for direct labeling of yeast microtubules for electron microscopy.
Book
The reproduction of biological systems may be referred to the reproduction of cells. Subcellular entities such as viruses cannot maintain themselves indefinitely without parallel reproduction of the cells in which they live. The asexual reproduction of organisms such as flatworms is limited by the production of new cells, and all cases of sexual reproduction imply the generation of new cells after the mixing of genetic material from the parent cells. An evolutionary consequence of cellularity is implicit in the designation of multicellular organisms as higher organisms. Single cells can develop a high level of variety and complexity of structure and function. In the development of a multicellular organism, cell division, beginning with the egg, is the essential step toward differentiation. If mitosis is a device for the distribution of sister genes to sister cells, the chromosomes may be viewed as a system in which the numerous genes are packaged into a small number of units. Mitosis is accompanied by some changes in the physical state or texture of the cytoplasm, which have been assessed as viscosity changes.
Article
This chapter discusses the establishment of protein synthesis and the timing of its initiation, an assessment of the relevance of protein synthesis in unfertilized eggs, and the evidence for the necessity of post-fertilization synthesis. The kinetics of labeling is described and some peculiarities of the pattern are discussed. The extent to which this pattern is or is not linked to cytological events is examined. The chapter reviews the source of information for polypeptide assembly comes and some points have been added concerning the extent to which the stored program alone can make anything resembling a normal embryo at any stage. The use of the term “program” is then justified, as it implies that the stored information is sufficient to code for many proteins. The relation of protein synthesis to differentiation can be studied in many ways. The chapter discusses two of the obvious ones, which are (1) following the amounts of one or more proteins, and the control of those amounts, in tissue cells of late embryonic stages or in differentiating cell populations of the adult organism; and (2) following the course of protein synthesis after fertilization of the egg.
Article
Animalization was induced with evans blue and with Zn++ in embryos of Arbacia punctulata and of Lytechinus variegatus, respectively. Li+ induced vegetalization in A. punctulata embryos. While animalization did not affect the rate of cleavage, vegetalized embryos exhibited a reduction in cell number at post-morula stages. Mid-gastrulae and corresponding experimental embryos each were labeled for 4 hr with uridine-[5-3H] and with L-[3H-methyl]-methionine. The rate of uptake of each exogenous RNA precursor was similar in control and in experimental embryos. Purified RNA preparations were fractionated by electrophoresis on 2.4% acrylamide+0.5 % agarose gels. Comparison of rates of incorporation of each RNA precursor into 26s and 18s RNAs indicated that on a per cell basis evans blue- and Zn++-animalized embryos showed a reduction (0.53–0.56) and Li+-vegetalized embryos an enhancement (1.41—1.53) in the rate of accumulation of newly made 26s and 18s RNAs compared to controls (1.00). These results suggest that chemically-induced animalized and vegetalized embryos provide useful tools for studying possible differential gene expression in different embryonic germ layers of the developing sea urchin embryo.
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Fully grown oocytes of the frog (Rana pipiens) undergo cytoplasmic and nuclear maturation when treated with progesterone after the follicular envelopes have been removed. The mechanism of this maturation was investigated by injection of cytoplasm from progesterone-treated oocytes at various stages of maturation into fully grown but immature oocytes. The injected cytoplasm becomes effective in inducing maturation by 12 hours after progesterone administration, reaches a maximum effectiveness around 20 hours, and then declines after the donor oocytes complete maturation. However, even cytoplasm from early embryos retains some capacity to induce oocyte maturation. The frequency with which maturation is induced is proportional to the volume of the injected cytoplasm. Progesterone itself is not directly responsible for the maturation-producing effect of injected cytoplasm since injected progesterone does not promote maturation. However, externally applied progesterone does induce the completion of the first meiotic division, presumably by releasing a cytoplasmic “maturation promoting factor.” The production of this cytoplasmic factor was not affected by removal of the nucleus.
Article
Changes in the pattern of polypeptide synthesis during the in vitro maturation of the porcine oocyte have been studied by high-resolution, two-dimensional, polyacrylamide gel electrophoresis. The findings reveal not only that pig oocytes mature in vitro, but also that they are engaged in the synthesis of a complex pattern of polypeptides throughout maturation. The results also demonstrate the presence of molecular markers, the appearance, disappearance, and significant change in intensity of which are temporally related with the sequential stages of nuclear meiotic maturation. Collectively, the evidence indicates the presence of a developmental program during oocyte maturation.
Article
When meiotic maturation of primary oocytes of the starfish Asterias forbesi is induced by 1-methyladenine, rapid and striking changes in the pattern of protein synthesis detectable by electrophoresis occur after germinal vesicle breakdown. These include a decline in relative labeling with [35S]methionine of several polypeptides synthesized in the oocyte, and increased labeling and new appearance of several polypeptides. Fertilization does not result in other detectable changes. The population of total mRNA translatable in a rabbit reticulocyte lysate cell-free system does not change, but the distribution of mRNAs between polysomes and the postribosomal supernatant reflects the changes observed in vivo. Thus these changes are regulated at the translational level. A review of the literature indicates that translationally mediated changes in patterns of protein synthesis during maturation of oocytes may be a widespread phenomenon.
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The absolute rate of protein synthesis in developing embryos of Strongylocentrotus purpuratus has been measured by lysine incorporation. Protein synthesis rises to about 240 pg hr−1 embryo−1 from the two- to eight-cell stage, and then gradually increases to a maximum of over 500 pg hr−1 embryo−1 in the blastula. The changes in protein synthesis are accompanied by similar increase in the polyribosomes in the embryo, so that 60–65% of the ribosomes are in polyribosomes by the blastula stage. The data are used to calculate an average peptide elongation rate of 1.8 amino acids ribosome−1 sec−1.
Article
Incubation of unfertilized eggs in ammonia has been reported to initiate such late responses to fertilization as K+-conductance, DNA synthesis, chromosome condensation and cytoplasmic mRNA polyadenylation. It does not initiate such early responses as Na+-influx and the cortical reactions. We have further analyzed this metabolic derepression and find that ammonia activation does not result in the early respiratory burst and also does not initiate the late activation of Na+-dependent amino acid transport. Protein synthesis is increased, similar to that following normal fertilization. This indicates that augmentation of protein synthesis is causally linked neither to the earlier Na+-influx, cortical reactions, and respiratory burst nor to the later activation of amino acid transport. The temporal correlation between activation of transport and increased protein synthesis is therefore coincidental. The association between increased protein synthesis and establishment of K+-conductance was analyzed by abolishing K+-conductance through acidification of the sea water. This did not affect protein synthesis, indicating that K+-conductance and protein synthesis are also not causally linked.There is also no obligate link between protein synthesis and chromosome condensation. Incubation in low concentrations of ammonia results in increased protein synthesis but not chromosome condensation. Higher ammonia concentrations cause chromosome condensation but with no further increase in rate of protein synthesis. This suggests a concentration-dependent hierarchy of activation.These results are consistent with the concept that the late fertilization changes are not causally linked and proceed independently of each other. As we have not been able to disassociate the early changes, they may be obligately linked and dependent on each other.
Article
Research on the early development of the sea urchin offers new insights into the process of embryogenesis. Maternal messenger RNA stored in the unfertilized egg supports most of the protein synthesis in the early embryo, but the structure of maternal transcripts suggests that additional functions are also possible. The overall developmental patterns of transcription and protein synthesis are known, and current measurements describe the expression of specific genes, including the histone genes, the ribosomal genes, and the actin genes. Possible mechanisms of developmental commitment are explored for regions of the early embryo that give rise to specified cell lineages, such as the micromere-mesenchyme cell lineage.
Article
Microtubule protein in RNA is present in considerable quantities in the unfertilized egg. After fertilization, the synthesis of microtubule proteins begins and proceeds at an increasing rate during cleavage.
Article
High resolution two-dimensional electrophoresis has been used to examine the pattern of protein synthesis during meiotic maturation of mouse oocytes in vitro. Fluorograms of [35S]methionine-labeled oocyte proteins have revealed that meiotic progression from dictyate to metaphase II (meiotic maturation) is accompanied by marked changes in the pattern of proteins synthesized by oocytes. Virtually all of the changes observed take place subsequent to the breakdown of the oocyte's germinal vesicle, but are not dependent upon the occurrence of other morphological events, such as spindle formation or polar body emission. These changes in protein synthesis do not take place in oocytes that fail to undergo breakdown of germinal vesicles spontaneously or in oocytes arrested at the germinal vesicle stage by dibutyryl 3':5'-cyclic AMP. These data suggest that mixing of the oocyte's nucleoplasm and cytoplasm may trigger many of the changes in protein synthesis that accompany meiotic maturation of mouse oocytes in vitro.
Article
Measurements of the rates of incorporation of [35S]methionine into protein and the specific activities of endogenous free methionine pools have been used to calculate the absolute rates of protein synthesis in mouse oocytes during spontaneous meiotic maturation in vitro. Fluorodinitro[3H]benzene was used to determine the specific activity of the oocyte's free methionine pool. It was found that the absolute rate of protein synthesis decreased from 43 to 31 pg/hr per oocyte during meiotic progression from dictyate to metaphase II (meiotic maturation), while the size of the intracellular free methionine pool decreased from 61 to 35 fmol per oocyte during the same period. Comparable measurements made on ovulated mouse oocytes that had undergone meiotic maturation in vivo strongly suggest that the decrease in the absolute rate of protein synthesis observed during meiotic maturation in vitro is physiologically significant. An alternative method that depends upon differential expansion of the oocyte's endogenous methionine pool was also used to determine absolute rates of protein synthesis. The results of these experiments are in excellent agreement with those obtained by using fluorodinitro[3H]benzene, indicating that the oocyte's free methionine pool is not compartmentalized.
Article
The activity of a cytoplasmic factor (MPF), capable of inducing nuclear membrane breakdown (germinal vesicle breakdown) when injected into amphibian oocytes, has been studied during the course of early cleavage in amphibian embryos. Mature egg cytoplasm was found to contain high levels of this activity, but this was quickly lost after fertilization or artificial activation. MPF activity later reappeared in the egg cytoplasm and started to cycle with time. The peak of embryonic MPF activity during each cycle coincided with the time the embryonic nuclei were entering the G2-M transition, i.e., mitosis. However, in colchicine-arrested embryos, this activity remained at an elevated level and no longer oscillated. The timing of the appearance and disappearance of this activity appeared to be under the control of the cytoplasm because such behavior was still observed in enucleated eggs. Continued protein synthesis in the embryo was required for the reappearance, but not for the disappearance, of this activity. MPF, previously thought to be restricted to oocyte maturation, may play a more general role in controlling nuclear membrane breakdown during mitosis as well as meiosis.
Article
The earliest stages of mouse embryogenesis, from fertilisation to the two-cell stage, are characterised by an extremely low level of RNA synthesis. Indeed, during this period, RNA polymerase II activity and incorporation of labelled precurosrs into heterogeneous RNA are not detectable, and there is no increase in the poly(A) content of the embryo, but rather a slight decrease. The rate of protein synthesis remains low and relatively constant throughout the one- and two-cell stages. However, qualitative analysis of the protein synthetic profile on SDS gels has revealed changes which appear around the late one-cell to early two-cell stage. This early change in the pattern of polypeptide synthesis represents the first major qualitative molecular change found so far in development. We present evidence which suggests that the increased synthesis at the early two-cell stage of a small number of polypeptides of molecular weight 35,000 is not dependent on transcription, but rather represents control at a post-transcriptional level using mRNAs synthesised before fertilisation.
Article
Unfertilized sea urchin eggs enter a mitotic chromosome cycle after treatment with sea water containing ammonia. Centrioles cannot be found but microtubules are formed in the later stages of the cycle. The microtubules are displayed in an astral arrangement centered on clusters of osmiophilic bodies. In early stages, distinct kinetochores on the condensed chromosomes show no attachments to microtubules. Later, a few microtubules may be attached to the kinetochores. The chromosomes and microtubules are contained in a "clear zone", a large compact accumulation of membranes which displaces yolk particles and mitochondria, but not ribosomes, from that region of the cell. No bipolar spindle is formed.
Article
Microtubule protein pools have been demonstrated to exist in unfertilized eggs and the early embryonic stages of several organisms. The microtubule pool of the sea urchin embryo is constant in size (about 0.4% of the total embryo protein) throughout early development. Protein withdrawn from this pool for organelle assembly is replaced by new synthesis. Eggs and embryos of Drosophila similarly contain a pool of microtubule proteins (larger than or equal to 0.4% of the total embryo protein, congruent to 3% of the soluble protein), which is constant in size throughout early development. The Drosophila egg microtubule proteins are easily purified by self-assembly in vitro of microtubules, and are similar to microtubule proteins from other organisms in molecular weight and other properties. Synthesis of microtubule proteins in sea urchin embryos is supported by oogenetic mRNA. This appears also to be the case in molluscan (Ilyanassa) embryos. It is not known whether Drosophila embryos synthesize microtubule proteins during the early stages of development.
Article
Cell-free extracts of Arbacia eggs (Arbacia punctulata) apparently do not contain an enzymatic system for the reduction of ribonucleotides to deoxyribonucleotides. However, during an interval of 5 hr after fertilization at 23 degrees , an enzymatic system is produced that is capable of catalyzing the reduction of CDP to dCDP in the presence of Mg(2+), ethylenediaminetetraacetate, ATP, and a reducing agent, dithiothreitol. The activity is first seen about 1 hr after fertilization, and reaches a peak at about 5 hr. The appearance of the ribonucleotide reductase is prevented by the addition of emetine or puromycin, inhibitors of protein synthesis, to the cells before fertilization. Inclusion of actinomycin D in the cell suspension at a concentration sufficient to inhibit synthesis of messenger RNA does not appreciably affect the production of the enzyme activity. Preexisting, maternal RNA is thus used for synthesis of reductase. Ribonucleotide reductase may, therefore, represent the first example of an enzyme system absent in unfertilized eggs that is produced in response to fertilization.
Article
Analysis of (35)S-methionine-labeled extracts of adenovirus 2-infected KB cells revealed 22 virus-induced polypeptide components. Most proteins of the virion were easily detected in extracts of whole cells labeled for short periods between 15 and 30 h after infection; however, several virion components were conspicuously absent. Radioactivity appeared in two of these virion components during a chase in nonradioactive medium, and this appearance was paralleled by a decrease in the radioactivity associated with two nonvirion adenovirus-induced proteins, results which imply precursor-product relationships for these components. Comparison of one of the chasable adenovirus-induced components (designated P-VII; mass of 20,000 daltons) and the major core protein (VII; mass of 18,500 daltons) of the virion showed that they have four common methionine-containing tryptic peptides; P-VII has an additional methionine residue which is not found in the major core protein. We propose that at least two of the adenovirus 2 virion components are derived by the cleavage of higher molecular weight precursor polypeptides.
Article
Annulate lamellae are peculiar cytoplasmic organelles composed of stacked sheets of endoplasmic reticulum-like membranes interrupted by annuli or pores. They are most commonly found in oocytes as well as embryonic and neoplastic cells, but they can be observed, albeit to a lesser degree, in virtually any eukaryotic cell type.
Article
Emetine is a potent inhibitor of protein synthesis in sea urchin embryos. At a concentration of the drug that rapidly inhibits protein synthesis in blastulae by 95%, uridine incorporation into RNA continues for more than 1 hr and presumptive histone messenger RNA is synthesized and transported into the cytoplasm where it is apparently associated with polyribosomes. Possible explanations of this result and its implications for the "informasome" theory of messenger transport in embryonic cells are discussed.
Article
Unfertilized eggs of Paracentrotus lividus activated with thymol and KCl can develop at least until the pluteus stage. Cytological studies showed that development of artificially activated eggs follows one of four different pathways: monaster formation, cytaster formation, haploid mitosis, or intranuclear chromosome replication. Cytospectrophotometric determination of DNA values from blastula nuclei indicated that parthenogenetic blastulae may contain either 1n, 2n or 4n nuclei or they may be a mosaic regarding the ploidy of their nuclei. DNA synthesis in artificially activated eggs begins about 2.5 times later as compared to fertilized controls; its rate is about half that of fertilized eggs. The rate of RNA synthesis in activated eggs is similar, though somewhat lower, than in fertilized eggs. Parthenogenetically activated and fertilized eggs show a very similar uptake of thymidine and uridine.
Article
A variety of steroid hormones are taken up from the incubation medium by isolated Rana pipiens oocytes, and of those tried, eight were capable of inducing maturation in vitro. Uptake of progesterone from the medium was linear with time and proportional to the concentration of hormone in the medium. Once bound, about 50% or more of the hormone remained permanently associated with the oocyte. However, the amount of uptake varied greatly among the hormones tested, as did effectiveness in inducing maturation, and there was no clear correlation between amount of uptake and induction effectiveness.Primary interaction of the hormone with the oocyte must occur at the oocyte surface, as injection of the hormone into the oocyte does not effect the maturation response. However, injection of cytoplasm from maturing eggs will effect maturation in oocytes not treated with hormone. We suggest that the steroid acts at or near the surface to cause the production of a second effector which, when accumulated to a sufficient level in the oocyte, induces the observable events associated with maturation.
Article
The kinetics of incorporation of tritiated leucine were used to study protein synthesis in activated eggs and early embryos of Rana pipiens. The results support the conclusion that yolk proteins are degraded to provide steady-state amino acid pools intermediate in the synthesis of cytoplasmic proteins de novo. The kinetic data can be used to calculate the rate of protein synthesis.
Article
The contribution of newly synthesized cell protein to the structural proteins of the mitotic apparatus during the first cell division of sea urchin eggs was investigated. Addition of puromycin 30 minutes after fertilization reduced amino acid incorporation during the remainder of the cell cycle by more than 75%, yet the eggs undergo mitosis at 100 minutes post-fertilization. If puromycin is present from the time of fertilization, DNA replication for the ensuing mitosis occurs, but mitosis itself is prevented. Amino acid incorporation into purified, solubilized mitotic apparatus is equal to, or only slightly greater than, incorporation into bulk cell protein. About 15% of the mitotic apparatus is insoluble in salt solutions and is apparently made at much faster rates than the bulk cell protein. It is concluded that synthesis of solubilized structural mitotic apparatus protein does not represent a heavy investment of the protein synthesis of the cell prior to the first cell division. Synthesis of catalytic components necessary for mitosis may, however, be important.
Article
The Xenopus embryo undergoes 12 rapid synchronous cleavages followed by a period of slower asynchronous divisions more typical of somatic cells. This change in cell cleavage has been termed the midblastula transition (MBT). We show that at the MBT the blastomeres become motile and transcriptionally active for the first time. We have investigated the timing of the MBT and found that it does not depend on cell division, on time since fertilization or on a counting mechanism involving the sequential modification of DNA. Rather, the timing of the MBT depends on reaching a critical ratio of nucleus to cytoplasm. We view the MBT as a consequence of the titration of some substance, originally present in the egg, by the exponentially increasing nuclear material. When this substance is exhausted a new cell program is engaged, leading to the acquisition of several new cell properties.
Article
Conditions that induce the formation of asters in unfertilized sea-urchin eggs have been investigated. Monasters were formed by treatment of eggs with acidic or basic sea-water, or procaine- or thymol-containing sea-water. A second treatment step, incubation with D2O-containing, ethanol-containing or hypertonic sea-water induced multiple cytasters. The number and size of cytasters varied according to the concentration of agents and duration of the first and second treatments, and also upon the species of eggs and the season in which the eggs were obtained. Generally, a longer second treatment or a higher concentration of the second medium resulted in a higher number of cytasters per egg. Asters were isolated and then examined by light and electron microscopy. Isolated monasters apparently lacked centrioles, whereas cytasters obtained from eggs undergoing the two-step treatment contained one or more centrioles. Up to eight centrioles were seen in a single aster; the centrioles appeared to have been produced during the second incubation. Centrospheres prepared from isolated asters retained the capacity to nucleate the formation of microtubules in vitro as assayed by light and electron microscopy. Many microtubules radiated from the centre of isolated asters, whether they contained centrioles or not. This observation is consistent with many other reports that microtubule-organizing centres need not contain centrioles.
Article
Hybrid myeloma cell lines secreting monoclonal antibodies to tubulin have been prepared using rat myelomas and spleen cells from rats immunized with yeast tubulin. A comparison between the results obtained with the rat myeloma Y3-Ag 1.2.3., which secretes a light chain, and a new line, YB2/O, which does not, shows that they are both excellent parental lines and that the second produces hybrids with no myeloma chain components. The antitubulin antibodies in the serum of rats bearing two of the hybrid myeloma tumors gave titers of up to 1:10(6) from which large amounts of monoclonal antibodies could be easily purified. They recognized tubulin from yeast as well as from birds and mammals. The two antibodies gave clear immunofluorescent staining of yeast mitotic spindles as well as the interphase microtubule network of tissue culture cells. Some difference in the pattern of immunofluorescence staining of yeast cells and nuclei was observed between the two antibodies. The purified antibodies could be conjugated to colloidal gold particles and used for direct labeling of yeast microtubules for electron microscopy.
Article
Striking changes in the pattern of protein synthesis occur shortly after the fertilization of Spisula solidissima oocytes. These changes include a strong reduction in the synthesis of prominent oocyte-specific proteins and a large increase in the synthesis of at least three proteins whose labeling dominates the pattern of protein synthesis in early embryos. Several independent lines of evidence suggest that these changes are modulated at the translational level:
Article
RNA transcripts complementary to a genomic histone repeat are found in high concentration in sea urchin egg pronuclei. In situ hybridizations with the recombinant plasmid pCO2 indicate that the nuclear concentration is at least 25 to 50 fold higher than that in the cytoplasm. If nuclear transcripts are predominantly histone mRNAs, they comprise about 12% of the histone mRNA in eggs, or about 0.36 pg. After fertilization these molecules persist through pronuclear fusion but disappear from nuclei by mid 2-cell stage. A similar high nuclear concentration is not observed for polyadenylated mRNAs. The high steady-state concentration of nuclear histone repeat transcripts suggests that they have an unusually long lifetime in pronuclei of unfertilized sea urchin eggs.
Article
During the early stages of embryogenesis, the sea urchin embryo uses maternally synthesized mRNA stored in the egg as inactive messenger ribonucleoproteins (mRNPs)1. Release of this mRNA for translation allows the embryo to develop even in the absence of new mRNA synthesis. Comparison using two-dimensional gel electrophoresis shows that the same spectrum of prevalent proteins are synthesized by eggs and zygotes 30-60 min after fertilization2. Thus, unlike the case of the clam Spisula in which there are several prominent changes in translation associated with fertilization3, sea urchins have been thought to show little or no regulation of translation of specific mRNA sequences. Histone mRNAs are major components of the mRNA pool, comprising as much as 4-8% of the total mRNA of eggs4, yet their products, histones, would not normally be detectable on two-dimensional gels. We report here that mRNA complementary to a histone H3 cloned probe remains in the inactive mRNA pool for 90 min after fertilization before it begins to be translated. This is long after the rapid increase in overall protein synthesis, using stored mRNAs, has begun. As we can demonstrate no significant synthesis of histone H3 mRNA during this period, stored histone H3 mRNA must be subject to sequence-specific translational regulation.
Article
The general metabolic activation of the sea urchin egg at fertilization is dependent on a release of intracellular stores of calcium and the subsequent transient elevation of intracellular Ca2+ (refs 1--3). However, this elevation does not by itself lead to increased macromolecular synthesis and development but initiates steps which result in a long-term elevation of intracellular pH (refs 4--6). Among the developmental processes dependent on the elevation of intracellular pH is the large acceleration in the rate of protein synthesis at fertilization. Weak penetrating bases such as ammonia can be used to mimic the processes resulting in an increase in intracellular pH and so show the corresponding increases in protein synthesis rate. Conversely, it is possible to demonstrate a gradual but complete shut down of protein synthesis if the intracellular pH is reduced to the unfertilized level with penetrating weak acids. However, the rate of protein synthesis in ammonia-activated eggs lags behind that of fertilized controls even though ammonia activation can result in an intracellular pH increase greater than occurs in the fertilized egg. This result has led to the suggestion that factors other than intracellular pH may be regulating protein synthesis following fertilization. To investigate the possibility that the Ca2+ transient may have such a role, we measured the rate of amino acid incorporation in eggs that were activated in various ionic conditions which enabled the effects of Ca2+ and pH changes to be studied separately. Our results, reported here, show that if intracellular pH is elevated, increases in intracellular Ca2+ play an additional part in the activation of protein synthesis at fertilization.
Article
Eggs and embryos of the sea urchin Lytechinus pictus were labeled with [35S]methionine. Aqueous extracts of protein were prepared and analyzed by a high resolution two-dimensional polyacrylamide gel electrophoresis system described recently by O'Farrell utilizing isoelectric focusing and sodium dodecyl sulfate electrophoresis. Out of about 400 distinctly resolved newly synthesized proteins, all but a few detectable in the zygote were being synthesized in the egg. Thus, the activation of translation of stored maternal messenger RNA following fertilization is due to a quantitative rather than qualitative change in the population of messenger RNA available for translation. The patterns of protein synthesis change only slightly during cleavage, but major differences appear by the beginning of gastrulation.
Article
Mitotic apparatus isolated at metaphase from sea urchin eggs, and placed in sea water containing leucine-carbon-14 after fertilization, has been shown by radioautographs to have taken up carbon-14 in such a way that it is not removable by repeated washing, by "chasing" with leucine-carbon-12, or by treatment with hot trichloroacetic acid.
Article
Résumé La puromycine inhibe fortement l'incorporation de la14C-L-valine dans les protéines, aussi bien dans les œufs fécondés d'oursins intacts que dans les préparations subcellulaires faites à partir de ceux-ci. Cependant ces préparations se sont montrées nettement plus sensibles à l'action de la puromycine que les œufs entiers. Parallèlement le blocage des divisions cellulaires a été proportionnel à cette action inhibitrice sur le métabolisme des protéines. Le développement s'est arrêté au stade «clear streak».
The Amerrcan Arbacia and other sea urchrns
  • E Harvey
Harvey, E. 8. (1956). The Amerrcan Arbacia and other sea urchrns. (Princeton, New Jersey Pnnceton Unrversrty Press).