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mex-1 and the general partitioning of cell fate in the early C-elegans embryo
Abstract
It is thought that at least some of the initial specification of the five somatic founder cells of the C. elegans embryo occurs cell-autonomously through the segregation of factors during cell divisions. It has been suggested that in embryos from mothers homozygous for mutations in the maternal-effect gene mex-1, four blastomeres of the 8-cell embryo adopt the fate of the MS blastomere. It was proposed that mex-1 functions to localise or regulate factors that determine the fate of this blastomere. Here, a detailed cell lineage analysis of 9 mex-1 mutants reveals that the fates of all somatic founder cells are affected by mutations in this gene. We propose that mex-1, like the par genes, is involved in establishing the initial polarity of the embryo.
... This general pattern of asymmetric localization of P granules happens prior to each of the subsequent cleavages, although additional mechanisms appear to contribute to P granule localization at later cleavages (Hird et al., 1996). Previous studies have shown that the maternal gene mex-1 is essential for the development of germ cells, and for the proper segregation of P granules during the first embryonic cleavages (Mello et al., 1992; Schnabel et al., 1996). P granules accumulate posteriorly in newly fertilized mex-1 mutant eggs, but they do not associate properly with the cortex and are thus spread throughout the posterior half of the egg. ...
... P granules accumulate posteriorly in newly fertilized mex-1 mutant eggs, but they do not associate properly with the cortex and are thus spread throughout the posterior half of the egg. In the next and subsequent cleavages, this incomplete localization leads to the mis-partitioning of P granules into somatic blastomeres, and a progressive loss of P granules in cells that would normally be germline blastomeres (Mello et al., 1992; Schnabel et al., 1996). mex-1 mutants also have complex defects in somatic differentiation (Mello et al., 1992; Schnabel et al., 1996 ). ...
... In the next and subsequent cleavages, this incomplete localization leads to the mis-partitioning of P granules into somatic blastomeres, and a progressive loss of P granules in cells that would normally be germline blastomeres (Mello et al., 1992; Schnabel et al., 1996). mex-1 mutants also have complex defects in somatic differentiation (Mello et al., 1992; Schnabel et al., 1996 ). The SKN- 1 transcription factor is required for the proper development of the ventral blastomeres MS and E, and is expressed at high levels in these blastomeres in wild-type embryos. ...
In the nematode Caenorhabditis elegans, germ cells arise from early embryonic cells called germline blastomeres. Cytoplasmic structures called P granules are present in the fertilized egg and are segregated into each of the germline blastomeres during the first few cleavages of the embryo. Mutations in the maternally expressed gene mex-1 disrupt the segregation of P granules, prevent the formation of germ cells, and cause inappropriate patterns of somatic cell differentiation. We have cloned the mex-1 gene and determined the distribution pattern of the mex-1 gene products. The MEX-1 protein contains two copies of an unusual 'finger' domain also found in the PIE-1 protein of C. elegans. PIE-1 has been shown to be expressed in germline blastomeres, and is a component of P granules. We show here that MEX-1 also is present in germline blastomeres and is a P granule component, although MEX-1 is a cytoplasmic protein while PIE-1 is present in both the nucleus and cytoplasm. We further show that MEX-1 is required to restrict PIE-1 expression and activity to the germline blastomeres during the early embryonic cleavages.
... As an additional assay of cell fate conversion, we tested the function of ectopic HLH-1 in embryos in which most cells had been specified as intestine. Loss of the maternal factor MEX-1 results in the four granddaughters of AB adopting an EMS-like fate (Mello et al., 1992;Schnabel et al., 1996). If these embryos are also depleted of POP-1/TCF, all daughters of the EMS and pseudo-EMS cells will develop like E, transforming the entire anterior of the embryo into intestine (Lin et al., 1995;Maduro et al., 2001). ...
... Wnt/MAP kinase signaling has also been implicated in the formation of body wall muscle cells from the MS lineage from previous studies of lit-1 (Kaletta et al., 1997). To explore this possibility further, we increased the number of MS-like blastomeres using mex-1 RNAi, which causes a transformation of AB granddaughter blastomeres to an MS-like lineage, in addition to other lineage defects (Mello et al., 1992;Schnabel et al., 1996). As a consequence of the reiteration of the MS lineage there is a large excess (four-to fivefold) of body wall and pharyngeal muscle that can be distinguished from each other using antibodies to MHC A and 3NB12, respectively. ...
... Most embryos had a small cluster of body wall muscle cells (~10) near the posterior of the embryo; these are probably descendants of D, which should be largely unaffected by lit-1 or wrm-1 RNAi as D has no detectable POP-1 (Lin et al., 1998) and only low levels of POP-1 are detected in D descendants (data not shown). mex-1 RNAi perturbs both the C and D lineages, in addition to affecting AB (Schnabel et al., 1996), explaining why we did not observed the normal number (20) of D descendants. Our results confirm that Wnt/MAPK signaling is required to knock down POP-1 levels in cells destined to be body wall muscle within the MS lineage. ...
In vertebrates, striated muscle development depends on both the expression of members of the myogenic regulatory factor family (MRFs) and on extrinsic cellular cues, including Wnt signaling. The 81 embryonically born body wall muscle cells in C. elegans are comparable to the striated muscle of vertebrates. These muscle cells all express the gene hlh-1, encoding HLH-1 (CeMyoD) which is the only MRF-related factor in the nematode. However, genetic studies have shown that body wall muscle development occurs in the absence of HLH-1 activity, making the role of this factor in nematode myogenesis unclear. By ectopically expressing hlh-1 in early blastomeres of the C. elegans embryo, we show that CeMyoD is a bona fide MRF that can convert almost all cells to a muscle-like fate, regardless of their lineage of origin. The window during which ectopic HLH-1 can function is surprisingly broad, spanning the first 3 hours of development when cell lineages are normally established and non-muscle cell fate markers begin to be expressed. We have begun to explore the maternal factors controlling zygotic hlh-1 expression. We find that the Caudal-related homeobox factor PAL-1 can activate hlh-1 in blastomeres that either lack POP-1/TCF or that have down-regulated POP-1/TCF in response to Wnt/MAP kinase signaling. The potent myogenic activity of HLH-1 highlights the remarkable developmental plasticity of early C. elegans blastomeres and reveals the evolutionary conservation of MyoD function.
... pe zinc-finger proteins, PIE-1, MEX-1, MEX-5/6 and POS-1, that possess the characteristic cysteine-X 8 − 10 -cysteine-X 5 -cysteine-X 3 -histidine sequence of the CCCH-type zinc-finger domain (Bai & Tolias 1996;Clarke & Berg 1998). Genetic studies have shown that they are important for differentiation of germ cells in C. elegans (Mello et al . 1992Schnabel et al . 1996;Guedes & Priess 1997;Tenenhaus et al . 1998;Tabara et al . 1999;Reese et al . 2000;Schubert et al . 2000). PIE-1 is a predominantly nuclear protein but also associates with P granules , and that appears to repress transcription in the germline blastomere (Seydoux et al . 1996;Batchelder et al . 1999;Seydoux & Strome 1999;Tenenhaus et al ...
... To examine possible interplay of OMA-1 localization with other factors that influence germline development, we analyzed the distribution of OMA-1 protein in mutants in which P granules are mislocalized: par-2, par-3, pie-1, mex-1 and mex-5 (Fig. 5). All are maternal-effect mutants; each exhibits unique cleavage and P granules partitioning defects as was previously reported (Fig. 5) (Kemphues et al. 1988;Mello et al. 1992;Etemad-Moghadam et al. 1995;Boyd et al. 1996;Schnabel et al. 1996;Guedes & Priess 1997;Schubert et al. 2000). Although OMA-1 protein clearly associated with P granules in par-2 or par-3 mutants, P granules localization of OMA-1 seems to be disturbed in some of the mutant embryos, such as mex-5 (Fig. 5). ...
... Eventually, PGL-1 became distributed abnormally in dividing embryos (76.8%, n = 148) ( Fig. 7A-b,c). It is known that the behavior of P granules is influenced by MEX-1 protein (Mello et al. 1992;Schnabel et al. 1996). We therefore tried to determine the distribution of MEX-1 protein. ...
In Caenorhabditis elegans, CCCH-type zinc-finger proteins have been shown to be involved in the differentiation of germ cells during embryonic development. Previously, we and others have identified novel redundant CCCH-type zinc-finger proteins, OMA-1 and OMA-2, that are involved in oocyte maturation. In this study, we report that the cytoplasmic expression level of OMA-1 protein was largely reduced after fertilization. In contrast to its cytoplasmic degradation, OMA-1 was found to accumulate exclusively on P granules in germline blastomeres during embryogenesis. A notable finding is that embryos with partially suppressed oma-1; oma-2 expression showed inappropriate germline specification, including abnormal distributions of PGL-1, MEX-1 and PIE-1 proteins. Thus, our results suggest that oma gene products are novel multifunctional proteins that participate in crucial processes for germline specification during embryonic development.
... In wildtype embryos, MS primarily produces mesodermal cells that build the posterior pharynx and contribute to body wall muscle. In mex-1 loss of function embryos, the four AB granddaughters adopt the MS fate as well (Figure 1.6) (Mello et al., 1992;Schnabel et al., 1996). The consequence of this blastomere fate misspecification is the Muscle EXcess Mex phenotype. ...
... The consequence of this blastomere fate misspecification is the Muscle EXcess Mex phenotype. As is the case with pie-1, reducing skn-1 function suppresses the ectopic MS differentiation in mex-1 embryos (Mello et al., 1992;Schnabel et al., 1996). ...
How do embryos develop with such poise from a single zygote to multiple cells with different identities, and yet survive? At the four-cell stage of the C. elegans embryo, only the blastomere EMS adopts the endo-mesoderm identity. This fate requires SKN-1, the master regulator of endoderm and mesoderm differentiation. However, in the absence of the RNA binding protein POS-1, EMS fails to fulfill its fate despite the presence of SKN-1. pos-1(-) embryos die gutless. Conversely, the RNA binding protein MEX-5 prevents ectoderm blastomeres from adopting the endo-mesoderm identity by repressing SKN-1. mex-5(-) embryos die with excess muscle at the expense of skin and neurons.
Through forward and reverse genetics, I found that genes gld-3/Bicaudal C, cytoplasmic adenylase gld-2, cye-1/Cyclin E, glp-1/Notch and the novel gene neg-1 are suppressors that restore gut development despite the absence of pos-1. Both POS-1 and MEX-5 bind the 3’UTR of neg-1 mRNA and its poly(A) tail requires GLD-3/2 for elongation. Moreover, neg-1 requires MEX-5 for its expression in anterior ectoderm blastomeres and is repressed in EMS by POS-1. Most neg-1(-) embryos die with defects in anterior ectoderm development where the mesoderm transcription factor pha-4 becomes ectopically expressed. This lethality is reduced by the concomitant loss of med- 1, a key mesoderm-promoting transcription factor.
Thus the endo-mesoderm identity of EMS is determined by the presence of SKN- 1 and the POS-1 repression of neg-1, whose expression is promoted by MEX-5. Together they promote the anterior ectoderm identity by repressing mesoderm differentiation. Such checks and balances ensure the vital plurality of cellular identity without the lethal tyranny of a single fate.
... Other weaker regulatory inputs into gut specification have been found that result from more global effects on embryonic gene expression. These include the Sp1 orthologue SPTF-3, the histone deacetylase HDA-1, the p300-like factor CBP-1, the Pur-alpha orthologue PLP-1, the endopeptidase TASP-1, the Prion-like-(Q/N-rich)-domain-bearing protein PQN-82, and other factors that can modulate gene expression epigenetically [39][40][41][42][43]. Finally, other factors have been identified that are important for restricting the activity of SKN-1 to MS and E, such as the RNA-binding proteins MEX-1 and PIE-1 [44][45][46]. Table 1. Core genes in endoderm specification in Caenorhabditis. ...
Cells in a developing animal embryo become specified by the activation of cell-type-specific gene regulatory networks. The network that specifies the gut in the nematode Caenorhabditis elegans has been the subject of study for more than two decades. In this network, the maternal factors SKN-1/Nrf and POP-1/TCF activate a zygotic GATA factor cascade consisting of the regulators MED-1,2 → END-1,3 → ELT-2,7, leading to the specification of the gut in early embryos. Paradoxically, the MED, END, and ELT-7 regulators are present only in species closely related to C. elegans, raising the question of how the gut can be specified without them. Recent work found that ELT-3, a GATA factor without an endodermal role in C. elegans, acts in a simpler ELT-3 → ELT-2 network to specify gut in more distant species. The simpler ELT-3 → ELT-2 network may thus represent an ancestral pathway. In this review, we describe the elucidation of the gut specification network in C. elegans and related species and propose a model by which the more complex network might have formed. Because the evolution of this network occurred without a change in phenotype, it is an example of the phenomenon of Developmental System Drift.
... The function of PIE-1 is controlled by the P granule factor MEX-1 (two "finger" domain, similar to PIE-1), which restricts PIE-1 expression and activity to germline blastomeres during early embryogenesis (Guedes and Priess, 1997). MEX-1 also controls proper localization and activity of the maternally provided transcription factor SKN-1 and subsequently determines early cell fate (Schnabel et al., 1996). The P Granule factor MEX-3 (a presumptive RNA binding protein) apparently acts independently of the MEX-1 and SKN-1/PIE-1 regulated blastomere specification (Draper et al., 1996). ...
Alterations of general or specific mRNA levels are a universal manifestation of the ageing
process (Cookson, 2011). During their existence, mRNAs are constantly decorated by dynamically
changing factors, which form messenger ribonucleoprotein (mRNP) complexes and
determine the fate of an mRNA. Mechanisms that control mRNA turnover in the cytoplasm have
been described in great detail but whether they might be involved in the regulation of ageing
is unknown (Anderson and Kedersha, 2009; Decker and Parker, 2012). Bulk mRNA decay in
eukaryotes is initiated by irreversible shortening of the poly(A)-tail, subsequent decapping and
final 5’ to 3’ degradation (Houseley and Tollervey, 2009). We present compelling evidence that
EDC-3, a highly conserved modulator of decapping, is a novel determinant of ageing in C. elegans.
Decapping has been shown to regulate protein synthesis by competing with the mechanism
of translation initiation. Congruently, we find that EDC-3 regulates protein synthesis and
lifespan in interaction with the previously described translation initiation factor IFE-2, an isoform
of the human eIF4E, which has a conserved role in the control of ageing. We demonstrate that
EDC-3 and IFE-2 mediated regulation of C. elegans lifespan happens specifically in neuronal
tissue and governs neural integrity. Further, we show that loss of EDC-3 protects from oxidative
and heat induced stress and that lifespan extension depends on the activity of Nrf-like xenobiotic-
response factor SKN-1 and heat shock response factor HSF-1. Also, longevity upon loss of
EDC-3 triggers a ROS induced hormesis response that depends on SKN-1 activity.
Most mRNPs accumulate in distinct cellular foci termed processing bodies (P-bodies)
or stress granules, which store mRNAs stalled in modes of degradation or translation initiation
(Sheth and Parker, 2003, reviewed in Decker and Parker, 2012; Franks and Lykke-Andersen,
2008). Decapping factors, including EDC-3, are part of P-bodies, while IFE-2 localizes to stress
granules in C. elegans. We establish an increased formation of P-bodies and stress granules
and their co-localization upon specific stress insults and during age in the nematode, thereby
defining them as biomarkers of ageing. It is unknown, whether mRNP granule formation is
cause or consequence of mRNA decay and stress response (Eulalio et al., 2007). We demonstrate,
that loss of SKN-1 contributes to an increased formation of P-bodies upon oxidative
stress. Curiously, down-regulation of HSF-1 prevents P-body assembly specifically upon heat
stress and causes age-related granulation of IFE-2. These results implicate that mRNP aggregation
is a transcriptionally controlled process that contributes to maintenance of cellular stress
response and ageing.
Unexpectedly, we find that HSF-1 suppresses transcription, stability and nuclear granulation
of IFE-2 during ageing. These granules co-localize with components of P-bodies at the
nuclear envelope, which also have been shown to be involved in transcription regulation in the
nucleus (summarized in Reines, 2012). Excitingly, we observe decreased localization of IFE-
2 in the nucleus, upon depletion of EDC-3. Our findings suggest that HSF-1 modulates IFE-2
function and localization during ageing, and that IFE-2 also serves as nuclear mRNP export
factor in C. elegans. Thus, IFE-2 likely mediates the effects of the heat stress response on both
mRNA translation and degradation to influence ageing.
... Thus, maternal genes specify the fates of early blastomeres, and maternal-effect mutations have been the entry point for analysis of early pattern formation. The first asymmetric division and early A/P axis formation depend on six par genes (partitioning defective) (Kemphues et al., 1988;Morton et al., 1992;Watts et al., 1996), pkc-3 (protein kinase C) (Tabuse et al., 1998;Wu et al., 1998), mex-1 (muscled in excess), mex-3 (Schnabel et al., 1996), and mes-1 (maternal-effect sterile) (Strome et al., 1995;Tenenhaus et al., 1998). These initial asymmetries set up the unequal distribution of a number of developmental fate determinants such as SKN-1 (skin in excess), PIE-1 (pharynx and intestine in excess), and PAL-1 (posterior alae in males) to specific cells of the early embryo ( Figure 6). ...
Asymmetric cell divisions constitute a basic foundation of animal development, providing a mechanism for placing specific cell types at defined positions in a developing organism. In a 4-cell stage embryo in Caenorhabditis elegans the EMS cell divides asymmetrically to specify intestinal cells, which requires a polarizing signal from the neighboring P2 cell. Here we describe how the extracellular signal from P2 is transmitted from the membrane to the nucleus during asymmetric EMS cell division, and present the identification of additional components in the pathways that accomplish this signaling.
P2/EMS signaling involves multiple inputs, which impinge on the Wnt, MAPK-like, and Src pathways. Transcriptional outputs downstream of these pathways depend on a homolog of β-catenin, WRM-1. Here we analyze the regulation of WRM-1, and show that the MAPK-like pathway maintains WRM-1 at the membrane, while its release and nuclear translocation depend on Wnt/Src signaling and sequential phosphorylation events by the major cell-cycle regulator CDK-1 and by the membrane-bound GSK-3 during EMS cell division. Our results provide novel mechanistic insights into how the signaling events at the cortex are coupled to the asymmetric EMS cell division through WRM-1.
To identify additional regulators in the pathways governing gut specification, we performed suppressor genetic screens using temperature-sensitive alleles of the gutless mutant mom-2/Wnt, and extra-gut mutant cks-1. Five intragenic suppressors and three semi-dominant suppressors were isolated in mom-2 suppressor screens. One extragenic suppressor was mapped to the locus ifg-1, eukaryotic translation initiation factor eIF4G. From the suppressor screen using cks-1(ne549), an allele of the self-cleaving nucleopore protein npp-10 was identified as a suppressor of cks-1(ne549) and other extra-gut mutants.
Taken together, these results help us better understand how the fate of intestinal cells are specified and regulated in early C. elegans embryos and broaden our knowledge of cell polarity and fate specification.
... For example, P2 (and its descendants) is not properly specified in the pie-1 or pos-1 mutant. Furthermore, in the mex-1 mutant the AB lineage develops with irregular timing (Schnabel et al., 1996) and the ABar fate is changed (Mello et al., 1992;Tabara et al., 1999). ...
The restricted spatiotemporal translation of maternal mRNAs, which is crucial for correct cell fate specification in early C. elegans embryos, is regulated primarily through the 3?UTR. Although genetic screens have identified many maternally expressed cell fate-controlling RNA-binding proteins (RBPs), their in vivo targets and the mechanism(s) by which they regulate these targets are less clear. These RBPs are translated in oocytes and localize to one or a few blastomeres in a spatially and temporally dynamic fashion unique for each protein and each blastomere. Here, we characterize the translational regulation of maternally supplied mom-2 mRNA, which encodes a Wnt ligand essential for two separate cell-cell interactions in early embryos. A GFP reporter that includes only the mom-2 3?UTR is translationally repressed properly in oocytes and early embryos, and then correctly translated only in the known Wnt signaling cells. We show that the spatiotemporal translation pattern of this reporter is regulated combinatorially by a set of nine maternally supplied RBPs. These nine proteins all directly bind the mom-2 3?UTR in vitro and function as positive or negative regulators of mom-2 translation in vivo. The net translational readout for the mom-2 3?UTR reporter is determined by competitive binding between positive- and negative-acting RBPs for the 3?UTR, along with the distinct spatiotemporal localization patterns of these regulators. We propose that the 3?UTR of maternal mRNAs contains a combinatorial code that determines the topography of associated RBPs, integrating positive and negative translational inputs.
... At the other end of the embryo, the maternal factor MEX-1 restricts appearance of ectopic MS-like fates by preventing appearance of high levels of SKN-1 protein in the early AB lineage [10,12,38] . In embryos lacking mex-1 function, the AB granddaughters adopt MS-like fates, concomitant with ectopic expression of med-1,2 and tbx-35 [12,25,33,39]. As with pie-1, SKN-1 is required for the appearance of ectopic MS-derived tissues in mex-1 mutant embryos, consistent with the ectopic activation of the normal MS specification pathway in the AB lineage [12]. ...
The specification of the Caenorhabditis elegans endomesoderm has been the subject of study for more than 15 years. Specification of the 4-cell stage endomesoderm precursor, EMS, occurs as a result of the activation of a transcription factor cascade that starts with SKN-1, coupled with input from the Wnt/β-catenin asymmetry pathway through the nuclear effector POP-1. As development proceeds, transiently-expressed cell fate factors are succeeded by stable, tissue/organ-specific regulators. The pathway is complex and uses motifs found in all transcriptional networks. Here, the regulators that function in the C. elegans endomesoderm network are described. An examination of the motifs in the network suggests how they may have evolved from simpler gene interactions. Flexibility in the network is evident from the multitude of parallel functions that have been identified and from apparent changes in parts of the corresponding network in Caenorhabditis briggsae. Overall, the complexities of C. elegans endomesoderm specification build a picture of a network that is robust, complex, and still evolving.
... LIT-1 is a kinase that controls anterior/posterior daughter cell fates beginning at the 6-cell stage when the ventral-most embryonic cell called EMS divides along the anterior/posterior body axis [61,62,63]. MEX-1 is a zinc finger protein that restricts blastomere identity at the 8-cell stage but also has been shown to affect anterior-posterior polarity at the one-cell stage [64,65,66]. We found one new lit-1 mutant, or393 ts and one new mex-1 mutant, or286 ts, which failed to complement the previously identified alleles lit-1(or131 ts) and mex-1(zu120), respectively (data not shown). ...
To study essential maternal gene requirements in the early C. elegans embryo, we have screened for temperature-sensitive, embryonic lethal mutations in an effort to bypass essential zygotic requirements for such genes during larval and adult germline development. With conditional alleles, multiple essential requirements can be examined by shifting at different times from the permissive temperature of 15°C to the restrictive temperature of 26°C. Here we describe 24 conditional mutations that affect 13 different loci and report the identity of the gene mutations responsible for the conditional lethality in 22 of the mutants. All but four are mis-sense mutations, with two mutations affecting splice sites, another creating an in-frame deletion, and one creating a premature stop codon. Almost all of the mis-sense mutations affect residues conserved in orthologs, and thus may be useful for engineering conditional mutations in other organisms. We find that 62% of the mutants display additional phenotypes when shifted to the restrictive temperature as L1 larvae, in addition to causing embryonic lethality after L4 upshifts. Remarkably, we also found that 13 out of the 24 mutations appear to be fast-acting, making them particularly useful for careful dissection of multiple essential requirements. Our findings highlight the value of C. elegans for identifying useful temperature-sensitive mutations in essential genes, and provide new insights into the requirements for some of the affected loci.
... However, such changed fates do not always show a clear lineage transformation. In mex-1 mutant embryos, the AB granddaughters adopt MS-like fates, but the transformations in the lineage itself are not as clear (Mello et al., 1992; Schnabel et al., 1996). Presumably, incomplete transformations in mutant backgrounds result from competition among multiple factors in the same lineage, as well as a change in cell–cell interactions when a blastomere adopts a fate outside of its normal context. ...
Cell specification requires that particular subsets of cells adopt unique expression patterns that ultimately define the fates of their descendants. In C. elegans, cell fate specification involves the combinatorial action of multiple signals that produce activation of a small number of "blastomere specification" factors. These initiate expression of gene regulatory networks that drive development forward, leading to activation of "tissue specification" factors. In this review, the C. elegans embryo is considered as a model system for studies of cell specification. The techniques used to study cell fate in this species, and the themes that have emerged, are described.
... Mickey, R. Hill, unpublished observations). If POS-1 and MEX-1 have a similar biochemical function, then phenotypic differences between pos-1 and mex-1 mutants (Mello et al., 1992; Schnabel et al., 1996) could result from POS-1 and MEX-1 having different targets, or from differences in the time that the proteins act in the germline blastomeres. MEX-1 is expressed at high levels in 1-cell stage embryos (Guedes and Priess, 1997), while POS-1 is not expressed at high levels until the 2-cell stage. ...
Germ cells arise during early C. elegans embryogenesis from an invariant sequence of asymmetric divisions that separate germ cell precursors from somatic precursors. We show that maternal-effect lethal mutations in the gene pos-1 cause germ cell precursors to inappropriately adopt somatic cell fates. During early embryogenesis, pos-1 mRNA and POS-1 protein are present predominantly in the germ precursors. POS-1 is a novel protein with two copies of a CCCH finger motif previously described in the germline proteins PIE-1 and MEX-1 in C. elegans, and in the mammalian TIS11/Nup475/TTP protein. However, mutations in pos-1 cause several defects in the development of the germline blastomeres that are distinct from those caused by mutations in pie-1 or mex-1. The earliest defect detected in pos-1 mutants is the failure to express APX-1 protein from maternally provided apx-1 mRNA, suggesting that POS-1 may have an important role in regulating the expression of maternal mRNAs in germline blastomeres.
... gld-1 is required for proper regulation of cell proliferation in the gonad (Francis et al., 1995). The pie-1, mex-1 and pos-1 genes are required, at least in part, to prevent the germline precursors from producing only somatic tissues (Mello et al., 1992; Schnabel et al., 1996; Tabara et al., 1999). In contrast, the mex-3 gene is required in part to prevent certain somatic precursors from adopting germline fates (Draper et al., 1996). ...
P granules are cytoplasmic structures of unknown function that are associated with germ nuclei in the C. elegans gonad, and are localized exclusively to germ cells, or germ cell precursors, throughout the life cycle. All the known protein components of P granules contain putative RNA- binding motifs, suggesting that RNA is involved in either the structure or function of the granules. However, no specific mRNAs have been identified within P granules in the gonad. We show here that P granules normally contain a low level of RNA, and describe conditions that increase this level. We present evidence that several, diverse mRNAs, including pos-1, mex-1, par-3, skn-1, nos-2 and gld- 1 mRNA, are present at least transiently within P granules.
In contrast, actin and tubulin mRNA and rRNA are either not present in P granules, or are present at relatively low levels. We show that pgl-1 and the glh (Vasa-related) gene family, which encode protein components of P granules, do not appear essential for RNA to concentrate in P granules; these proteins may instead function in events that are a prerequisite for RNAs to be transported efficiently from the nuclear surface.
... 34 Recent work, however, suggests that the gene mex-1 may not function specifically to establish blastomere identity but rather functions in the specification of the initial polarity of the embryo like the par-genes. 35,36 For the pie-1 gene it was now shown that it functions in the general suppression of transcription in the germ line precursor P2. 37,38 Extensive lineage analysis using the 4D-microscope of embryos mutant in the gene glp-1 or of laser-ablated embryos showed that certain AB-derived blastomeres could acquire the fates (identities) normally only executed by other AB blastomeres. 18,39 During investigations of the role of the MS blastomere in the induction of pharyngeal muscles in the AB lineage 17,21 it was recognized that the MS blastomere induces blastomere identities along the left-right axis of the embryo rather than specific tissues. ...
C. elegans is renowned for its invariant embryogenesis and functions as a major paradigm for a mode of development coupled to an invariant lineage. Recent work, however, suggests that the embryogenesis of the nematode is much more flexible than anticipated. The invariant premorphogenetic stage is formed from variable earlier stages through a sorting of cells. Cells do not act as individuals but already early in embryogenesis a regionalization of the embryo occurs. Cells are diversified by a binary specification of 'abstract' blastomere (regional) identities. The determination of tissues may thus be a very late event. It appears that C. elegans, although assigning cell fates in an invariant lineage pattern, uses the same strategies and mechanisms for embryogenesis as organisms with variable lineages.
... POS-1 controls polarity and cell cycle timing during the second phase of germ cell asymmetric division, and unlike PIE-1, is not required to initiate transcriptional silencing. Other germ cell-specific proteins, such as MEX-3 and MEX-1, also regulate germ cell fate ( Mello et al., 1992;Draper et al., 1996;Schnabel et al., 1996;Guedes and Priess, 1997). ...
Sm and Sm-like proteins are core components of the splicesome but have other functions distinct from pre-mRNA processing. Here, we show that Sm proteins also regulate germ cell specification during early C. elegans embryogenesis. SmE and SmG were required to maintain transcriptional quiescence in embryonic germ cell precursors. In addition, depletion of SmE inhibited expression of the germ lineage-specific proteins PIE-1, GLD-1, and NOS-2, but did not affect maintenance of several maternal mRNAs. PIE-1 had previously been shown to activate transcriptional silencing and NOS-2 expression. We found that PIE-1 also promotes GLD-1 expression by a process that is independent of transcriptional silencing. Thus, Sm proteins could control transcriptional silencing and maternal protein expression by regulating PIE-1. However, loss of SmE function also caused defects in P granule localization and premature division in early germline blastomeres, processes that are independent of PIE-1 function. Therefore, the Sm proteins control multiple aspects of germ cell precursor development. Because depletion of several other core splicing factors did not affect these events, these Sm functions are likely distinct from pre-mRNA splicing. Sm family proteins assemble into ribonucleoprotein complexes (RNPs) that control RNA activities. We suggest that novel Sm RNPs directly or indirectly influence posttranscriptional control of maternal mRNAs to promote germ cell specification in the early C. elegans embryo.
... Ectopic expression of either unc-120 or hnd-1 in the skn- 1, pal-1 double-RNAi background resulted in widespread myogenesis as assayed by MHCA (Fig. 1B ). Similar effects were seen when RNAi was used to knockdown both mex-1 and pop-1, which results in the anterior three-quarters of embryonic blastomeres adopting a gut cell fate while the posterior quarter becomes muscle (Fig. 1C; Mello et al. 1992; Schnabel et al. 1996; Fukushige and Krause 2005; Maduro et al. 2005). Taken together, these embryonic conversion assay results demonstrated that both UNC-120 and HND-1 were myogenic on their own, albeit weakly so in comparison with HLH-1. ...
Myogenic regulatory factors (MRFs) are required for mammalian skeletal myogenesis. In contrast, bodywall muscle is readily detectable in Caenorhabditis elegans embryos lacking activity of the lone MRF ortholog HLH-1, indicating that additional myogenic factors must function in the nematode. We find that two additional C. elegans proteins, UNC-120/SRF and HND-1/HAND, can convert naïve blastomeres to muscle when overproduced ectopically in the embryo. In addition, we have used genetic null mutants to demonstrate that both of these factors act in concert with HLH-1 to regulate myogenesis. Loss of all three factors results in embryos that lack detectable bodywall muscle differentiation, identifying this trio as a set that is both necessary and sufficient for bodywall myogenesis in C. elegans. In mammals, SRF and HAND play prominent roles in regulating smooth and cardiac muscle development. That C. elegans bodywall muscle development is dependent on transcription factors that are associated with all three types of mammalian muscle supports a theory that all animal muscle types are derived from a common ancestral contractile cell type.
... PIE-1, MEX-1, and POS-1 each play a different but crucial role in C. elegans development. PIE-1 is required to maintain the transcriptional quiescence of the germline lineage , MEX-1 is required to restrict muscle fate to one blastomere at the eight-cell stage (Schnabel et al., 1996), and POS-1 is required to specify pharyngeal, intestinal, and germline fates (Tabara et al., 1999). Thus, tandem zinc finger proteins play a number of essential yet diverse roles in the development of the early embryo. ...
Most sexually reproducing metazoans are anisogamous, meaning that the two gametes that combine during fertilization differ greatly in size. By convention, the larger gametes are considered female and are called ova, while the smaller gametes are male and are called sperm. In most cases, both gametes contribute similarly to the chromosomal content of the new organism. In contrast, the maternal gamete contributes nearly all of the cytoplasm. This cytoplasmic contribution is crucial to patterning early development; it contains the maternal proteins and transcripts that guide the early steps of development prior to the activation of zygotic transcription. This review compares and contrasts early development in common laboratory model organisms in order to highlight the similarities and differences in the regulation of maternal factors. We will focus on the production and reversible silencing of maternal mRNAs during oogenesis, their asymmetric activation after fertilization, and their subsequent clearance at the midblastula transition. Where possible, insights from mechanistic studies are presented.
Asymmetric cell division (ACD) is a cellular process that forms two different cell types through a cell division and is thus critical for the development of all multicellular organisms. Not all but many of the ACD processes are mediated by proper orientation of the mitotic spindle, which segregates the fate determinants asymmetrically into daughter cells. In many cell types, the evolutionarily conserved protein complex of Gαi/AGS-family protein/NuMA-like protein appears to play critical roles in orienting the spindle and/or generating the polarized cortical forces to regulate ACD. Studies in various organisms reveal that this conserved protein complex is slightly modified in each phylum or even within species. In particular, AGS-family proteins appear to be modified with a variable number of motifs in their functional domains across taxa. This apparently creates different molecular interactions and mechanisms of ACD in each developmental program, ultimately contributing to developmental diversity across species. In this review, we discuss how a conserved ACD machinery has been modified in each phylum over the course of evolution with a major focus on the molecular evolution of AGS-family proteins and its impact on ACD regulation.
This chapter reviews some of the experimental embryology that has set the stage for a genetic and molecular understanding of the mechanisms that control pattern formation during embryogenesis in Caenorhabditis elegans. Some insight into how regulation of the actin cytoskeleton polarizes the 1-cell stage C. elegans zygote on fertilization has come from the identification of six maternally expressed par genes, par-1 through par-6. The three maternal genes skn-1, pal-1, and pie-1 encode putative transcriptional regulators that appear to act intrinsically to regulate initiation of the genetic programs that define the fates of the four P1-derived somatic founder cells MS, E, C, and D. This view is based in part on the observation that in skn-1;pal-1 double mutant embryos, P1 frequently fails to produce any differentiated somatic cell types and instead produces many small and apparently undifferentiated descendants. The genetics and sequenced genome of C. elegans have provided rapid access to the molecules and mechanisms that initiate pattern formation in this cellularized animal embryo.
TheCaenorhabditis elegans elt-2gene encodes a single-finger GATA factor, previously cloned by virtue of its binding to a tandem pair of GATA sites that control the gut-specificges-1esterase gene. In the present paper, we show thatelt-2expression is completely gut specific, beginning when the embryonic gut has only two cells (one cell cycle prior toges-1expression) and continuing in every cell of the gut throughout the life of the worm. Whenelt-2is expressed ectopically using a transgenic heat-shock construct, the endogenousges-1gene is now expressed in most if not all cells of the embryo; several other gut markers (including a transgenicelt-2-promoter::lacZreporter construct designed to test forelt-2autoregulation) are also expressed ectopically in the same experiment. These effects are specific in that two otherC. elegansGATA factors (elt-1andelt-3) do not cause ectopic gut gene expression. An imprecise transposon excision was identified that removes the entireelt-2coding region. Homozygouselt-2null mutants die at the L1 larval stage with an apparent malformation or degeneration of gut cells. Although the loss ofelt-2function has major consequences for later gut morphogenesis and function, mutant embryos still expressges-1.We suggest thatelt-2is part of a redundant network of genes that controls embryonic gut development; other factors may be able to compensate forelt-2loss in the earlier stages of gut development but not in later stages. We discuss whether elements of this regulatory network may be conserved in all metazoa.
This chapter focuses on the maternal control of the generation of unique cleavage patterns. In ascidian embryos, experimental transplantation of egg cytoplasm has revealed the presence of localized factors in the egg cytoplasm. In gastropods and nematodes, maternal-effect variants and mutants show genetic evidence for the maternal contribution in controlling the pattern of cleavage. In nematodes, the products of such genes, the PAR proteins, show clear localization in the egg cortex. These studies indicate the importance of egg organization and localized factors for generation of unique cleavage patterns. To create divergence of the cleavage pattern from the “default” equal and orthogonal pattern, various mechanisms for positioning the mitotic apparatus are used in different animal systems. In sea urchin, ascidian, and nematode embryos, movements of centrosomes during interphase play key roles in the orientation and asymmetric positioning of the mitotic apparatus. The movements of the centrosome involve the cortical site, the centrosome, and the microtubules between them. In the nematode embryo, and in the second and third cleavages of oligochaete eggs, the mitotic spindle migrates during the mitotic phase. Microfilaments are required for the migration in the latter case. In the first cleavage of oligochaete, a difference in the size of the asters causes unequal cleavage.
has been described completely on a cell-by-cell basis and found to be essentially invariant. With this knowledge in hands,
micromanipulated embryos and mutants have been analyzed for cell lineage defects and the distribution of specific gene products.
The results challenge the classical view of cell-autonomous development in nematodes and indicate that the early embryo of
C. elegans is a highly dynamic system. A network of inductive events between neighboring cells is being revealed, which is necessary
to assign different developmental programs to blastomeres. In those cases where molecules involved in these cell-cell interactions
have been identified, homologies to cell surface receptors, ligands and transcription factors found in other systems have
become obvious.
Recent findings suggest that C. elegans, albeit displaying an invariant cell lineage for embryonic development, uses the same basic strategy for embryogenesis as other organisms. The early embryo is regionalised by cell-cell interactions.
After the first division of the C. elegans embryo, the posterior blastomere can produce numerous muscles while the anterior blastomere cannot. We show here that maternal-effect lethal mutations in the gene mex-3 cause descendants of the anterior blastomere to produce muscles by a pattern of development similar to that of a descendant of the wild-type posterior blastomere. mex-3 encodes a probable RNA-binding protein that is distributed unequally in early embryos and that is a component of germline-specific granules called P granules. We propose that MEX-3 contributes to anterior-posterior asymmetry by regulating one or more mRNAs involved in specifying the fate of the posterior blastomere.
Blastomeres in C. elegans embryos execute lineage programs wherein the fate of a cell is correlated reproducibly with the division sequence by which that cell is born. We provide evidence that the pop-1 gene functions to link anterior-posterior cell divisions with cell fate decisions. Each anterior cell resulting from an anterior-posterior division appears to have a higher level of nuclear POP-1 protein than does its posterior sister. Genes in the C. elegans Wnt pathway are required for this inequality in POP-1 levels. We show that loss of pop-1(+) activity leads to several types of anterior cells adopting the fates of their posterior sisters. These results suggest a mechanism for the invariance of blastomere lineages.
Genetic screens for recessive, maternal-effect, embryonic-lethal mutations have identified about 25 genes that control early steps of pattern formation in the nematode Caenorhabditis elegans. These maternal genes are discussed as belonging to one of three groups. The par group genes establish and maintain polarity in the one-cell zygote in response to sperm entry, defining an anterior/posterior body axis at least in part through interactions with the cyto-skeleton mediated by cortically localized proteins. Blastomere identity group genes act down-stream of the par group to specify the identities of individual embryonic cells, or blastomeres, using both cell autonomous and non-cell autonomous mechanisms. Requirements for the blastomere identity genes are consistent with previous studies suggesting that early asymmetric cleavages in the C. elegans embryo generate six "founder" cells that account for much of the C. elegans body plan. Intermediate group genes, most recently identified, may link the establishment of polarity in the zygote by par group genes to the localization of blastomere identity group gene functions. This review summarizes the known requirements for the members of each group, although it seems clear that additional regulatory genes controlling pattern formation in the early embryo have yet to be identified. An emerging challenge is to link the function of the genes in these three groups into interacting pathways that can account for the specification of the six founder cell identities in the early embryo, five of which produce somatic cell types and one of which produces the germline.
The Caenorhabditis elegans elt-2 gene encodes a single-finger GATA factor, previously cloned by virtue of its binding to a tandem pair of GATA sites that control the gut-specific ges-1 esterase gene. In the present paper, we show that elt-2 expression is completely gut specific, beginning when the embryonic gut has only two cells (one cell cycle prior to ges-1 expression) and continuing in every cell of the gut throughout the life of the worm. When elt-2 is expressed ectopically using a transgenic heat-shock construct, the endogenous ges-1 gene is now expressed in most if not all cells of the embryo; several other gut markers (including a transgenic elt-2-promoter::lacZ reporter construct designed to test for elt-2 autoregulation) are also expressed ectopically in the same experiment. These effects are specific in that two other C. elegans GATA factors (elt-1 and elt-3) do not cause ectopic gut gene expression. An imprecise transposon excision was identified that removes the entire elt-2 coding region. Homozygous elt-2 null mutants die at the L1 larval stage with an apparent malformation or degeneration of gut cells. Although the loss of elt-2 function has major consequences for later gut morphogenesis and function, mutant embryos still express ges-1. We suggest that elt-2 is part of a redundant network of genes that controls embryonic gut development; other factors may be able to compensate for elt-2 loss in the earlier stages of gut development but not in later stages. We discuss whether elements of this regulatory network may be conserved in all metazoa.
In the early Caenorhabditis elegans embryo five somatic founder cells are born during the first cleavages. The first of these founder cells, named AB, gives rise to 389 of the 558 nuclei present in the hatching larva. Very few genes directly involved in the specification of the AB lineage have been identified so far. Here we describe a screen of a large collection of maternal-effect embryonic lethal mutations for their effect on the early expression of a pes-1::lacZ fusion gene. This fusion gene is expressed in a characteristic pattern in 14 of the 32 AB descendants present shortly after the initiation of gastrulation. Of the 37 mutations in 36 genes suspected to be required specifically during development, 12 alter the expression of the pes-1::lacZ marker construct. The gene expression pattern alterations are of four types: reduction of expression, variable expression, ectopic expression in addition to the normal pattern, and reduction of the normal pattern together with ectopic expression. We estimate that approximately 100 maternal functions are required to establish the pes-1 expression pattern in the early embryo.
Studies of about 20 maternally expressed genes are providing an understanding of mechanisms of patterning and cell-fate determination in the early Caenorhabditis elegans embryo. The analyses have revealed that fates of the early blastomeres are specified by a combination of intrinsically asymmetric cell divisions and two types of cell-cell interactions: inductions and polarizing interactions. In this review we summarize the current level of understanding of the molecular mechanisms underlying these processes in the specification of cell fates in the pregastrulation embryo.
A wee1 homolog, wee-1.1, is expressed in both a temporally and spatially restricted pattern during early Caenorhabditis elegans embryogenesis, and is undetectable throughout the remainder of embryogenesis. The wee-1.1 message appears to be zygotically expressed in the somatic founder cell E of the 12-cell embryo. This expression disappears when the E blastomere divides for the first time. The wee-1.1 message then appears transiently in the nuclei of the eight great-granddaughter cells of the AB somatic founder cell, just before these cells divide in the 16-cell embryo. Following this division, the wee-1.1 mRNA is no longer detectable throughout the remainder of embryogenesis. The expression of wee-1.1 in the E blastomere and in the AB progeny appears to be restricted to nuclei in prophase and metaphase of the cell cycle. Analysis of the wee-1.1 mRNA expression pattern in maternal-effect lethal mutants suggests that this expression pattern is restricted to cells of the E and AB fates in the early embryo. This mRNA expression pattern is restricted to a 10-15-min span of embryonic development and may be regulating the timing of crucial cell divisions at this early stage of development.
In all animals information is passed from parent to offspring via the germline, which segregates from the soma early in development and undergoes a complex developmental program to give rise to the adult gametes. Many aspects of germline development have been conserved throughout the animal kingdom. Here we review the unique properties of germ cells, the initial determination of germ cell fates, the maintenance of germ cell identity, the migration of germ cells to the somatic gonadal primordia and the proliferation of germ cells during development invertebrates and invertebrates. Similarities in germline development in such diverse organisms as Drosophila melanogaster, Caenorhabditis elegans, Xenopus laevis and Mus musculus will be highlighted.
During Caenorhabditis elegans embryogenesis the primordial germ cell, P(4), is generated via a series of unequal divisions. These divisions produce germline blastomeres (P(1), P(2), P(3), P(4)) that differ from their somatic sisters in their size, fate and cytoplasmic content (e.g. germ granules). mes-1 mutant embryos display the striking phenotype of transformation of P(4) into a muscle precursor, like its somatic sister. A loss of polarity in P(2) and P(3) cell-specific events underlies the Mes-1 phenotype. In mes-1 embryos, P(2) and P(3) undergo symmetric divisions and partition germ granules to both daughters. This paper shows that mes-1 encodes a receptor tyrosine kinase-like protein, though it lacks several residues conserved in all kinases and therefore is predicted not to have kinase activity. Immunolocalization analysis shows that MES-1 is present in four- to 24-cell embryos, where it is localized in a crescent at the junction between the germline cell and its neighboring gut cell. This is the region of P(2) and P(3) to which the spindle and P granules must move to ensure normal division asymmetry and cytoplasmic partitioning. Indeed, during early stages of mitosis in P(2) and P(3), one centrosome is positioned adjacent to the MES-1 crescent. Staining of isolated blastomeres demonstrated that MES-1 was present in the membrane of the germline blastomeres, consistent with a cell-autonomous function. Analysis of MES-1 distribution in various cell-fate and patterning mutants suggests that its localization is not dependent on the correct fate of either the germline or the gut blastomere but is dependent upon correct spatial organization of the embryo. Our results suggest that MES-1 directly positions the developing mitotic spindle and its associated P granules within P(2) and P(3), or provides an orientation signal for P(2)- and P(3)-specific events.
Germ cells are essential for reproduction, yet the molecular mechanisms that underlie their unique development are only beginning to be understood. Here we review important events that lead to the establishment of the germline and the initiation of meiotic development in C. elegans. Formation of the germline begins in the pregastrulation embryo, where it depends on polarization along the anterior/posterior axis and on the asymmetric segregation of P granules and associated factors. During postembryonic development, the germline expands using the GLP-1/Notch signaling pathway to promote proliferation and regulate entry into meiosis. Throughout their development, germ cells also employ unique "silencing" mechanisms to regulate their genome and protect themselves against unwanted expression from repetitive sequences including transposable elements. Together these mechanisms preserve the health and reproductive potential of the germline.
Early C. elegans embryos exhibit protein asymmetries that allow rapid diversification of cells. Establishing these asymmetries requires the novel protein MEX-5. We show that mutations in the efl-1 and dpl-1 genes cause defects in protein localization resembling defects caused by mutations in mex-5. efl-1 and dpl-1 encode homologs of vertebrate E2F and DP proteins that regulate transcription as a heterodimer. efl-1 and dpl-1 mutants have elevated levels of activated Map kinase in oocytes. Their mutant phenotype and that of mex-5 mutants can be suppressed by reducing Ras/Map kinase signaling. We propose this signaling pathway has a role in embryonic asymmetry and that EFL-1/DPL-1 control the level of Map kinase activation.
The recent identification and characterization of the Caenorhabditis elegans gene spn-4 has shed new light on the mechanisms that link embryonic polarity to the specification of cell fates.
In C. elegans, mRNA production is initially repressed in the embryonic germline by a protein unique to C. elegans germ cells, PIE-1. PIE-1 is degraded upon the birth of the germ cell precursors, Z2 and Z3. We have identified a chromatin-based mechanism that succeeds PIE-1 repression in these cells. A subset of nucleosomal histone modifications, methylated lysine 4 on histone H3 (H3meK4) and acetylated lysine 8 on histone H4 (H4acetylK8), are globally lost and the DNA appears more condensed. This coincides with PIE-1 degradation and requires that germline identity is not disrupted. Drosophila pole cell chromatin also lacks H3meK4, indicating that a unique chromatin architecture is a conserved feature of embryonic germ cells. Regulation of the germline-specific chromatin architecture requires functional nanos activity in both organisms. These results indicate that genome-wide repression via a nanos-regulated, germ cell-specific chromatin organization is a conserved feature of germline maintenance during embryogenesis.
One of the best-studied model organisms in biology is Caenorhabditis elegans. Because of its simple architecture and other biological advantages, considerable data have been collected about the regulation of its development. In this review, currently available data concerning the early phase of embryonic development are presented in the form of genetic networks. We performed computer simulations of regulatory mechanisms in embryonic development, and the results are described and compared with experimental observations.
As part of a general study of genes specifying a pattern of muscle attachments, we identified and genetically characterised mutants in the mup-1 gene. The body wall muscles of early stage mup-1 embryos have a wild-type myofilament pattern but may extend ectopic processes. Later in embryogenesis, some body wall muscles detach from the hypodermis. Genetic analysis suggests that mup-1 has both a maternal and a zygotic component and is not required for postembryonic muscle growth and attachment. mup-1 mutants are suppressed by mutations in several genes that encode extracellular matrix components. We propose that mup-1 may encode a cell surface/extracellular matrix molecule required both for the positioning of body wall muscle attachments in early embryogenesis and the subsequent maintenance of these attachments to the hypodermis until after cuticle synthesis.
Bilateral pairs of blastomeres derived from the founder cell AB, the anterior blastomere of the 2-cell stage, in the Caenorhabditis elegans embryo are initially equivalent in their developmental potential. Recently, we showed that an induction at the 12-cell stage by a blastomere called MS is necessary to establish the differences between left and right pairs of blastomeres in the anterior part of the embryo. Further analysis of the process of creating left-right asymmetry reveals that the induction at the 12-cell stage is only the first of a series of inductions establishing the left-right asymmetry of the embryo. We describe here two further inductions that create additional asymmetries in the posterior part of the embryo. One induction occurs at the 24-cell stage among AB descendants themselves. This induction is restricted to the left side of the embryo as a consequence of the fate changes induced by MS at the 12-cell stage. The second induction requires again blastomeres of the MS lineage and also occurs around the 24-cell stage. Together these inductions establish the fate differences observed in the development of left-right pairs of blastomeres in the embryo.
During the first four cleavage rounds of the Caenorhabdi- tis elegans embryo, five somatic founder cells AB, MS, E, C and D are born, which later form the tissues of the embryo. The classical criterion for a cell-autonomous specification of a tissue is the capability of primordial cells to produce this tissue in isolation from the remainder of the embryo. By this criterion, the somatic founder cells MS, C and D develop cell-autonomously. Laser ablation experiments, however, reveal that within the embryonic context these blastomeres form a network of duelling cellular interactions.
During normal development, the blastomere D inhibits muscle specification in the MS and the C lineage inhibits muscle specification in the D lineage. These inhibitory interactions are counteracted by two activating inductions. As described before the inhibition of body wall muscle in MS is counteracted by an activating signal from the ABa lineage. Body wall muscle in the D lineage is induced by MS descendants, which suppress an inhibitory activity of the C lineage. The interaction between the D and the MS lineage occurs through the C lineage. An interesting feature of these cell-cell interactions is that they do not serve to dis- criminate between equivalent cells but are permissive or nonpermissive inductions.
No evidence was found that the C-derived body wall muscle also depends on an induction, which suggests that possibly three different pathways coexist in the early embryo to specify body wall muscle, two of which are, in different ways, influenced by cell-cell interactions and a third that is autonomous.
This work supplies evidence that cells may acquire transient states during embryogenesis that influence the specification of other cells in the embryo. These states, however, may not be reflected in the developmental potentials of the cells themselves. They can only be scored indi- rectly by their action on the specification of other cells in the embryo.
Blastomeres that behave cell-autonomously in isolation are nevertheless subjected to cell-cell interactions in the embryonic context. Why this should be is an intriguing question. The classical notion has been that blastomeres are specified autonomously in nematodes. In recent years, it was established that at least five inductions are required to determine the AB descendants of C. elegans, whereas the P1 descendants have been typically viewed to develop more autonomously. It appears now that inductions also play a major role during the determination of P1-derived blastomeres.
Most somatic cells in the nematode Caenorhabditis elegans arise from AB, the anterior blastomere of the 2-cell embryo. While the daughters of AB, ABa and ABp, are equivalent in potential at birth, they adopt different fates as a result of their unique positions. One such difference is that the distribution of epidermal precursors arising from ABp is reversed along the anterior-posterior axis relative to those arising from ABa. We have found that a strong mutation in the glp-1 gene eliminates this ABa/ABp difference. Furthermore, extensive cell lineage analyses showed that ABp adopts an ABa-like fate in this mutant. This suggests that glp-1 acts in a cellular interaction that makes ABp distinct from ABa. One ABp-specific cell type was previously shown to be induced by an interaction with a neighboring cell, P2. By removing P2 from early embryos, we have found that the widespread differences between ABa and ABp arise from induction of the entire ABp fate by P2. Lineage analyses of genetically and physically manipulated embryos further suggest that the identifies of the AB great-granddaughters (AB8 cells) are controlled by three regulatory inputs that act in various combinations. These inputs are: (1) induction of the ABp-specific fate by P2, (2) a previously described induction of particular AB8 cells by a cell called MS, and (3) a process that controls whether an AB8 cell is an epidermal precursor in the absence of either induction. When an AB8 cell is caused to receive a new combination of these regulatory inputs, its lineage pattern is transformed to resemble the lineage of the wild-type AB8 cell normally receiving that combination of inputs. These lineage patterns are faithfully reproduced irrespective of position in the embryo, suggesting that each combination of regulatory inputs directs a unique lineage program that is intrinsic to each AB8 cell.
The first cleavage of C. elegans is asymmetric, generating daughter cells with different sizes, cytoplasmic components, and fates. Mutations in the par-1 gene disrupt this asymmetry. We report here that par-1 encodes a putative Ser/Thr kinase with similarity to kinases from yeasts and mammals. Two strong alleles have mutations in the kinase domain, suggesting that kinase activity is essential for par-1 function. PAR-1 protein is localized to the posterior periphery of the zygote and is distributed in a polar fashion preceding the asymmetric divisions of the germline lineage. Because PAR-1 distribution in the germline correlates with the distribution of germline-specific P granules, it is possible that PAR-1 functions in germline development as well as in establishing embryonic polarity.
In a C. elegans embryo the third cleavages of descendants of the anterior blastomere AB of the 2-cell stage create pairs of blastomeres that develop differently. By laser ablation experiments we show that the fates of all the posterior daughters of this division depend on an induction occurring three cleavages before these blastomeres are born. The time of induction precludes a direct effect on cell fate. Alternatively, we suggest that the induction creates a heritable cell polarity which is propagated through several divisions. We suggest a model to demonstrate how a signal could be propagated through several rounds of cell division. An important implication of our observations is that this early induction acts to specify blastomere identity, not tissue type. A detailed lineage analysis revealed that altering the inductive signal alters complex lineage patterns as a whole. The induction described here, together with two inductions described previously can be used to illustrate how the anterior portion of the C. elegans embryo can be successively subdivided into blastomeres with unique developmental potential.
In a 4-cell Caenorhabditis elegans embryo, two sister blastomeres called ABa and ABp are born with equivalent developmental potential, but eventually produce distinct patterns of cell fate. The different fates of ABa and ABp are specified at least in part by inductive interactions with neighboring blastomeres. Previous studies indicate that, at the 4-cell stage, a signal from the posterior-most blastomere, P2, is required for ABp to produce at least one of its unique cell types. This P2/ABp interaction depends on glp-1, a putative receptor for intercellular interactions. To investigate this early induction further, we isolated mutants in which ABp developed abnormally. We describe the effects of recessive mutations in apx-1, a maternal gene that appears to be required for P2 to signal ABp. In embryos from mothers homozygous for mutations in apx-1 (apx-1 embryos), ABp fails to produce its characteristic cell types. Instead, ABp from apx-1 embryos develops more like its sister ABa: it produces ABa-like pharyngeal cells and it recapitulates ABa-like cell lineages. Because mutations in apx-1 affect the development of only the ABp blastomere, we suggest that the wild-type gene encodes a component of the P2/ABp signalling pathway. To explain the observation that ABp in apx-1 embryos adopts an ABa-like fate, we propose a model in which the P2 signal is required to break the initial equivalence of ABa and ABp. We performed two independent tests of this model. First, we examined ABp development in pie-1 mutant embryos, in which P2 adopts the identity of another blastomere. We find that, in pie-1 embryos, APp fails to produce its characteristic cell types and instead adopts a fate similar to that of ABa. We conclude that the changed identity of P2 in pie-1 embryos prevents the P2/ABp interaction. As a second test, we examined ABp development in wild-type embryos after physically removing P2. These operated embryos produce extra pharyngeal cells, consistent with out proposal that a signal from P2 breaks the initially equivalent developmental state of ABa and ABp. We discuss the possibility that apx-1 acts as a ligand in this glp-1-dependent signalling pathway.
Two successive inductions specify blastomere identities, that is complex cell lineages and not specific tissues, in a major part of the early C. elegans embryo. The first induction acts along the anterior-posterior axis of the embryo and the second along the left-right axis. During the first induction a specific lineage program is induced in the posterior of the two AB blastomeres present in the four cell embryo. During the second induction, almost all of the left-right differences of the embryo are specified by interactions between a single signalling blastomere, MS, and the AB blastomeres that surround it. In both cases the inductions break the equivalence of pairs of blastomeres. The inductions correlate with the cell-cell contacts to the inducing blastomeres. The stereotype cleavage patterns of the early embryo results in invariant cell-cell contacts that guarantee the specificity of the inductions. Both inductions are affected in embryos mutant for glp-1 suggesting that in both cases glp-1 is involved in the reception of the signal.
EMS, a blastomere of the Caenorhabditis elegans embryo, produces body wall muscle cell-autonomously in isolation. Within the embryonic context, however, the specification of body wall muscle derived from EMS depends on inductive interactions between its daughter MS and ABa descendants that are required to overcome inhibitory interactions with other cells. The inductive events between the MS and ABa descendants are reciprocal, specifying subsequent fates in both lineages. Both induction events are blocked by mutations in the gene glp-1, known to encode a Notch-like transmembrane receptor protein.
We have examined the cortex of Caenorhabditis elegans eggs during pseudocleavage (PC), a period of the first cell cycle which is important for the generation of asymmetry at first cleavage (Strome, S. 1989. Int. Rev. Cytol. 114: 81-123). We have found that directed, actin dependent, cytoplasmic, and cortical flow occurs during this period coincident with a rearrangement of the cortical actin cytoskeleton (Strome, S. 1986. J. Cell Biol. 103: 2241-2252). The flow velocity (4-7 microns/min) is similar to previously determined particle movements driven by cortical actin flows in motile cells. We show that directed flows occur in one of the daughters of the first division that itself divides asymmetrically, but not in its sister that divides symmetrically. The cortical and cytoplasmic events of PC can be mimicked in other cells during cytokinesis by displacing the mitotic apparatus with the microtubule polymerization inhibitor nocodazole. In all cases, the polarity of the resulting cortical and cytoplasmic flows correlates with the position of the attenuated mitotic spindle formed. These cortical flows are also accompanied by a change in the distribution of the cortical actin network. The polarity of this redistribution is similarly correlated with the location of the attenuated spindle. These observations suggest a mechanism for generating polarized flows of cytoplasmic and cortical material during embryonic cleavages. We present a model for the events of PC and suggest how the poles of the mitotic spindle mediate the formation of the contractile ring during cytokinesis in C. elegans.
This paper provides a brief summary of the Caenorhabditis elegans cell lineage, the evidence for both intrinsic and extrinsic cell specification, and experiments that suggest mechanisms for cell differentiation and patterning.
Two dominant suppressors of crossing over have been identified following X-ray treatment of the small nematode C. elegans. They suppress crossing over in linkage group II (LGII) about 100-fold and 50-fold and are both tightly linked to LGII markers. One, called C1, segregates independently of all other linkage groups and is homozygous fertile. The other is a translocation involving LGII and X. The translocation also suppresses crossing over along the right half of X and is homozygous lethal. C1 has been used as a balancer of LGII recessive lethal and sterile mutations induced by EMS. The frequencies of occurrence of lethals and steriles were approximately equal. Fourteen mutations were assigned to complementation groups and mapped. They tended to map in the same region where LGII visibles are clustered.
By the 4-cell stage of C. elegans embryogenesis, a ventral blastomere, called EMS, is already committed to producing pharyngeal and intestinal cell types. Recessive, maternal-effect mutations in the gene skn-1 prevent EMS from producing both pharyngeal and intestinal cells. In skn-1 mutant embryos, EMS instead produces hypodermal cells and body wall muscle cells, much like its sister blastomere. Genetic analysis suggests that the skn-1 gene product is also required post-embryonically for development of the intestine. We have cloned and sequenced the skn-1 gene and describe sequence similarities to the basic regions of bZIP transcription factors. We propose that the maternally expressed skn-1 gene product acts to specify the fate of the EMS blastomere.
Specification of some cell fates in the early Caenorhabditis elegans embryo is mediated by cytoplasmic localization under control of the maternal genome. Using nine newly isolated mutations, and two existing mutations, we have analyzed the role of the maternally expressed gene par-4 in cytoplasmic localization. We recovered seven new par-4 alleles in screens for maternal effect lethal mutations that result in failure to differentiate intestinal cells. Two additional par-4 mutations were identified in noncomplementation screens using strains with a high frequency of transposon mobility. All 11 mutations cause defects early in development of embryos produced by homozygous mutant mothers. Analysis with a deficiency in the region indicates that it33 is a strong loss-of-function mutation. par-4(it33) terminal stage embryos contain many cells, but show no morphogenesis, and are lacking intestinal cells. Temperature shifts with the it57ts allele suggest that the critical period for both intestinal differentiation and embryo viability begins during oogenesis, about 1.5 hr before fertilization, and ends before the four-cell stage. We propose that the primary function of the par-4 gene is to act as part of a maternally encoded system for cytoplasmic localization in the first cell cycle, with par-4 playing a particularly important role in the determination of intestine. Analysis of a par-4; par-2 double mutant suggests that par-4 and par-2 gene products interact in this system.
Two types of developmental events can cause an embryonic cell to adopt a fate different from that of its neighbours: during a cell division particular contents may be segregated to only one daughter cell and cells may experience different external cues, commonly in the form of inductive cell interactions. Work on development in the nematode Caenorhabditis elegans suggests that most cell fates are specified without a need for cell interactions. In particular, the gut cell lineage of C. elegans has been used as a primary example of specification by differential segregation of determinants. Here I re-examine the role of induction in gut specification by isolating early blastomeres. In C. elegans, the gut derives from all the progeny of a single blastomere (E) of the eight-cell stage. When a gut precursor cell (EMS) is isolated during the first half of the four-cell stage, gut does not differentiate. Gut differentiation is rescued by recombining EMS with its posterior neighbour (P2), but not by recombining EMS with one or both of the other two cells of the four-cell stage. These results demonstrate that P2 induces EMS to form gut in C. elegans.
During C. elegans embryogenesis an 8-cell stage blastomere, called MS, undergoes a reproducible cleavage pattern, producing pharyngeal cells, body wall muscles, and cell deaths. We show here that maternal-effect mutations in the pie-1 and mex-1 genes cause additional 8-cell stage blastomeres to adopt a fate very similar to that of the wild-type MS blastomere. In pie-1 mutants one additional posterior blastomere adopts an MS-like fate, and in mex-1 mutants four additional anterior blastomeres adopt an MS-like fate. We propose that maternally provided pie-1(+) and mex-1(+) gene products may function in the early embryo to localize or regulate factors that determine the fate of the MS blastomere.
Classical work implied that early nematode embryogenesis is completely mosaic. This view was lately challenged by the demonstration that in C. elegans an early interaction has to occur to induce the production of muscle from a blastomere. Here, early embryonic blastomeres were inactivated by laser microsurgery. The cell lineages of irradiated embryos were compared to those of intact embryos. It is shown that one blastomere, MS, is required for the specification of mesodermal pharyngeal fates and another blastomere, P2, for the specification of hypodermal fates from the descendants of the AB blastomere, whereas the proper specification of the nervous system requires the presence of both. The irradiation of a third blastomere shows that interactions also occur within the ectoderm. I propose that the body plan of the C. elegans embryo may be established by two primary signals followed by secondary interactions. The suggested mechanisms are reminiscent of those involved in amphibian development.
The early somatic blastomeres founding the tissues in the C. elegans embryo are derived in a stem-cell-like lineage from the P cells. We have isolated maternal effect lethal mutations defining the gene cib-1 in which the P cells, P1-P3, skip a cell cycle and acquire the fates of only their somatic daughters. Therefore, the cib-1 gene is required for the specification of the stem-cell-like fate of these cells. The analysis of the development of these mutants suggests that the clock controlling the cell cycles in the early embryo is directly coupled to the fate of a cell and that there must be another developmental clock that activates the determinative inventory for the early decision-making.
We have analyzed a set of linkage group (LG) II maternal-effect lethal mutations in Caenorhabditis elegans isolated by a new screening procedure. Screens of 12,455 F1 progeny from mutagenized adults resulted in the recovery of 54 maternal-effect lethal mutations identifying 29 genes. Of the 54 mutations, 39 are strict maternal-effect mutations defining 17 genes. These 17 genes fall into two classes distinguished by frequency of mutation to strict maternal-effect lethality. The smaller class, comprised of four genes, mutated to strict maternal-effect lethality at a frequency close to 5 X 10(-4), a rate typical of essential genes in C. elegans. Two of these genes are expressed during oogenesis and required exclusively for embryogenesis (pure maternal genes), one appears to be required specifically for meiosis, and the fourth has a more complex pattern of expression. The other 13 genes were represented by only one or two strict maternal alleles each. Two of these are identical genes previously identified by nonmaternal embryonic lethal mutations. We interpret our results to mean that although many C. elegans genes can mutate to strict maternal-effect lethality, most genes mutate to that phenotype rarely. Pure maternal genes, however, are among a smaller class of genes that mutate to maternal-effect lethality at typical rates. If our interpretation is correct, we are near saturation for pure maternal genes in the region of LG II balanced by mnC1. We conclude that the number of pure maternal genes in C. elegans is small, being probably not much higher than 12.
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We have isolated and analyzed eight strict maternal effect mutations identifying four genes, par-1, par-2, par-3, and par-4, required for cytoplasmic localization in early embryos of the nematode C. elegans. Mutations in these genes lead to defects in cleavage patterns, timing of cleavages, and localization of germ line-specific P granules. Four mutations in par-1 and par-4 are fully expressed maternal effect lethal mutations; all embryos from mothers homozygous for these mutations arrest as amorphous masses of differentiated cells but are specifically lacking intestinal cells. Four mutations in par-2, par-3, and par-4 are incompletely expressed maternal effect lethal mutations and are also grandchildless; some embryos from homozygous mothers survive and grow to become infertile adults due to absence of functional germ cells. We propose that all of these defects result from the failure of a maternally encoded system for intracellular localization in early embryos.
During early embryogenesis of Caenorhabditis elegans the serial stem cell-like cleavages of the germ line cells P0-P3 generate a number of somatic founder cells with different developmental potentials. Observations on partial embryos show that in the first two of these unequal divisions in the germ line the somatic daughter cell comes to lie anterior to the new germ line cell. In the following two, however, the somatic daughter cell comes to lie posterior to the new germ line cell, suggesting a reversal of polarity in the germ line. By the use of a laser microbeam, egg fragments can be extruded from young embryos; the fragments often cleave like partial twins. Depending on whether the fragment is derived from the posterior region of the uncleaved zygote P0 or its daughter P1, the mirror image duplications that are generated are joined at their larger soma-like cells or at their smaller germ line-like cells, respectively. This result is best explained as a reversal of polarity taking place in the germ line cell P2. This notion is strengthened by the finding that partial embryos derived from the posterior region of the P2 cell in late interphase do not undergo stem cell-like (i.e., unequal) cleavages in contrast to those derived from P0 or P1. Finally, an apparent early cell-cell interaction is described which is inconsistent with the classical notion of "mosaic" nematode development: removal of the germline cell P2 results in an altered developmental pattern of its somatic sister cell EMS. A working model is presented linking reversal of polarity and cell-cell interaction and offers an explanation for the unique behavior of the EMS cell in normal development.
In normal development both the anterior and posterior blastomeres in a 2-cell C. elegans embryo produce some descendants that become muscles. We show that cellular interactions appear to be necessary in order for the anterior blastomere to produce these muscles. The anterior blastomere does not produce any muscle descendants after either the posterior blastomere or one of the daughters of the posterior blastomere is removed from the egg. Moreover, we demonstrate that a daughter of the anterior blastomere that normally does not produce muscles appears capable of generating muscles when interchanged with its sister, a cell that normally does produce muscles. Embryos develop normally after these blastomeres are interchanged, suggesting that cellular interactions play a major role in determining the fates of some cells in early embryogenesis.
We have analyzed the differentiation potential of cells in early embryos of Caenorhabditis elegans by assessing the production of markers for intestinal, muscle, and hypodermal cell differentiation in cleavage-arrested blastomeres. Our results show that differentiation potential does not always segregate during cleavage in a linear fashion, i.e., a blastomere can express a differentiation potential that is absent in its parent blastomere and vice versa. Furthermore, the expression of a particular differentiation program by certain cleavage-arrested blastomeres is an exclusive event in that each cell will express only one program of differentiation, even though it may have the potential to express several.
We describe an esterase activity that, by the criterion of histochemical staining, is completely localized to the intestine of the nematode Caenorhabditis elegans. Esterase activity appears in the embryonic gut when the embryo contains 4-8 intestinal precursor cells and 100-150 total cells. Esterase activity is abolished by treating early embryos with alpha-amanitin, indicating that expression depends upon transcription by RNA polymerase II within the developing embryo. In partial embryos produced by lysing one blastomere of a two-cell embryo, esterase expression appears only in descendants of the blastomere that normally produces the gut; esterase expression appears independent of the other non-gut blastomere. In early cleavage-stage embryos in which cytokinesis has been blocked by cytochalasin D, esterase expression appears at the normal time and only in cells in the gut lineage; thus neither normal cell division nor normal embryogenesis is required for lineage-specific expression. However, esterase does not appear in cytochalasin D blocked one-cell embryos. These observations confirm the traditional view that C. elegans development is "mosaic," with each cell following a defined independent program of gene expression.
Ninety-five mutants of the nematode Caenorhabditis elegans altered in the cell lineages of the vulva have been isolated on the basis of their displaying one of two phenotypes, Vulvaless or Multivulva. In Vulvaless mutants, which define 12 genes, no vulva is present. In Multivulva mutants, which define ten genes, one or more supernumerary vulva-like protrusions are located along the ventral side of the animal. A single recessive mutation is responsible for the phenotypes of most, but not all, of these strains. Fifteen of these 22 genes are represented by multiple alleles. We have shown by a variety of genetic criteria that mutations that result in a Vulvaless or Multivulva phenotype in six of the 22 genes most likely eliminate gene function. In addition, Vulvaless or Multivulva mutations in seven of the other genes most likely result in a partial reduction of gene function; the absence of the activity of any of these genes probably results in lethality or sterility. Our results suggest that we may have identified most, or all, genes of these two classes.
Methods are described for the isolation, complementation and mapping of mutants of Caenorhabditis elegans, a small free-living nematode worm. About 300 EMS-induced mutants affecting behavior and morphology have been characterized and about one hundred genes have been defined. Mutations in 77 of these alter the movement of the animal. Estimates of the induced mutation frequency of both the visible mutants and X chromosome lethals suggests that, just as in Drosophila, the genetic units in C. elegans are large.
Six schemes were used to identify 80 independent recessive lethal deficiencies of linkage group (LG) II following X-ray treatment of the nematode Caenorhabditis elegans. Complementation tests between the deficiencies and ethyl methanesulfonate-induced recessive visible, lethal and sterile mutations and between different deficiencies were used to characterize the extents of the deficiencies. Deficiency endpoints thus helped to order 36 sites within a region representing about half of the loci on LG II and extending over about 5 map units. New mutations occurring in this region can be assigned to particular segments of the map by complementation tests against a small number of deficiencies; this facilitates the assignment of single-site mutations to particular genes, as we illustrate. Five sperm-defective and five oocyte-defective LG II sterile mutants were identified and mapped. Certain deficiency-by-deficiency complementation tests allowed us to suggest that the phenotypes of null mutations at two loci represented by visible alleles are wild type and that null mutations at a third locus confer a visible phenotype. A segment of LG II that is about 12 map units long and largely devoid of identified loci seems to be greatly favored for crossing over.
Germ-line granules in C. elegans embryos (P granules) can be visualized by immunofluorescence microscopy using a monoclonal antibody. In mutant zygotes with abnormal spindle orientations and in wild-type zygotes treated with the microtubule inhibitors nocodazole, colcemid, vinblastine, and griseofulvin, both P-granule segregation to the posterior pole and the concomitant pseudocleavage occur apparently normally, but the normally concurrent migration of the pronuclei is inhibited. Conversely, treatment of wild-type embryos with the microfilament inhibitors cytochalasins D and B inhibits P-granule segregation and pseudocleavage, as well as other manifestations of polarity, without preventing pronuclear migration. The results suggest that P-granule segregation does not require either the spindle or cytoplasmic microtubules, but that this process as well as generation of other asymmetries does require cytoskeletal functions that depend on microfilaments.
The embryonic cell lineage of Caenorhabditis elegans has been traced from zygote to newly hatched larva, with the result that the entire cell lineage of this organism is now known. During embryogenesis 671 cells are generated; in the hermaphrodite 113 of these (in the male 111) undergo programmed death and the remainder either differentiate terminally or become postembryonic blast cells. The embryonic lineage is highly invariant, as are the fates of the cells to which it gives rise. In spite of the fixed relationship between cell ancestry and cell fate, the correlation between them lacks much obvious pattern. Thus, although most neurons arise from the embryonic ectoderm, some are produced by the mesoderm and a few are sisters to muscles; again, lineal boundaries do not necessarily coincide with functional boundaries. Nevertheless, cell ablation experiments (as well as previous cell isolation experiments) demonstrate substantial cell autonomy in at least some sections of embryogenesis. We conclude that the cell lineage itself, complex as it is, plays an important role in determining cell fate. We discuss the origin of the repeat units (partial segments) in the body wall, the generation of the various orders of symmetry, the analysis of the lineage in terms of sublineages, and evolutionary implications.
We have followed the appearance of differentiation markers in cleavage-inhibited and uninhibited early blastomeres of C. elegans and have compared the cleavage patterns of blastomeres in partial and complete embryos. The results indicate that at least some primary differentiation of embryonic cells is determined by internal factors that segregate in early cleavages, whereas patterns of cleavage are dictated by both internally segregating determinants and external cues.
Establishment of the gut founder cell (E) in C. elegans involves an interaction between the P2 and the EMS cell at the four cell stage. Here I show that the fate of only one daughter of EMS, the E cell, is affected by this induction. In the absence of the P2-EMS interaction, both E and its sister cell, MS, produce pharyngeal muscle cells and body wall muscle cells, much as MS normally does.
By cell manipulations and inhibitor studies, I show first that EMS loses the competence to respond before it divides even once, but P2 presents an inducing signal for at least three cell cycles. Second, induction on one side of the EMS cell usually blocks the other side from responding to a second P2-derived signal. Third, microfilaments and microtubules may be required near the time of the interaction for subsequent gut differentiation. Lastly, cell manipulations in pie-1 mutant embryos, in which the P2 cell is transformed to an EMS-like fate and produces a gut cell lineage, revealed that gut fate is segregated to one of P2’s daughters cell-autonomously. The results contrast with previous results from similar experiments on the response to other inductions, and suggest that this induction may generate cell diversity by a different mechanism.
The par-2 gene of Caenorhabditis elegans functions in early embryogenesis to ensure an asymmetric first cleavage and the segregation of cytoplasmic factors. Both processes appear to be required to generate daughter blastomeres with distinct developmental potential. We isolated an allele of par-2 by using a screen for maternal-effect lethal mutations in a strain known for its high frequency of transposition events. A transposable element was found to be linked to this allele. Sequences flanking the site of transposon insertion were cloned and found to rescue the par-2 mutant phenotype. DNA in the par-2 region hybridized to a 2.3-kb germ-line-enriched mRNA. The cDNA corresponding to this germ-line-enriched message was cloned, sequenced, and used to identify the molecular lesions associated with three par-2 alleles. Sequence analysis of the par-2 cDNA revealed that the predicted protein contained two distinct motifs found in other known proteins: an ATP-binding site and a cysteine-rich motif which identifies the par-2 gene product as a member of a growing class of putative zinc-binding proteins.
The sister blastomeres ABp and ABa are equipotent at the beginning of the 4-cell stage in C. elegans embryos, but soon become committed to different fates. We show that the glp-1 gene, a homolog of the Notch gene of Drosophila, functions in two distinct cell-cell interactions that specify the ABp and ABa fates. These interactions both require maternal expression of glp-1. We show that a second maternal gene, apx-1, functions with glp-1 only in the specification of the ABp fate and that apx-1 can encode a protein homologous to the Delta protein of Drosophila. Our results suggest how interactions mediated by glp-1 and apx-1 contribute to the establishment of the dorsal-ventral axis in the early C. elegans embryo.
The autonomous or cell-intrinsic developmental properties of early embryonic blastomeres in nematodes are thought to result from the action of maternally provided determinants. After the first cleavage of the C. elegans embryo, only the posterior blastomere, P1, has a cell-intrinsic ability to produce pharyngeal cells. The product of the maternal gene skn-1 is required for P1 to produce pharyngeal cells. We show here that the Skn-1 protein is nuclear localized and that P1 appears to accumulate markedly higher levels of Skn-1 protein than its sister, the AB blastomere. We have examined the distribution of Skn-1 protein in embryos from mothers with maternal-effect mutations in the genes mex-1, par-1, and pie-1. These results suggest that mex-1(+) and par-1(+) activities are required for the unequal distribution of the Skn-1 protein and that pie-1(+) activity may function to regulate the activity of Skn-1 protein in the descendants of the posterior blastomere P1.
- Goh
- Goldstein
- Herman
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- Mango
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- Hutter
- Moskowitz
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