Publications (18)145.44 Total impact
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Article: Organization of lamprey variable lymphocyte receptor C locus and repertoire development.
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ABSTRACT: Jawless vertebrates are pivotal representatives for studies of the evolution of adaptive immunity due to their unique position in chordate phylogeny. Lamprey and hagfish, the extant jawless vertebrates, have an alternative lymphocyte-based adaptive immune system that is based on somatically diversifying leucine-rich repeat (LRR)-based antigen receptors, termed variable lymphocyte receptors (VLRs). Lamprey T-like and B-like lymphocyte lineages have been shown to express VLRA and VLRB types of anticipatory receptors, respectively. An additional VLR type, termed VLRC, has recently been identified in arctic lamprey (Lethenteron camtschaticum), and our analysis indicates that VLRC sequences are well conserved in sea lamprey (Petromyzon marinus), L. camtschaticum, and European brook lamprey (Lampetra planeri). Genome sequences of P. marinus were analyzed to determine the organization of the VLRC-encoding locus. In addition to the incomplete germ-line VLRC gene, we have identified 182 flanking donor genomic sequences that could be used to complete the assembly of mature VLRC genes. Donor LRR cassettes were classifiable into five basic structural groups, the composition of which determines their order of use during VLRC assembly by virtue of sequence similarities to the incomplete germ-line gene and to one another. Bidirectional VLRC assembly was predicted by comparisons of mature VLRC genes with the sequences of donor LRR cassettes and verified by analysis of partially assembled intermediates. Biased and repetitive use of certain donor LRR cassettes was demonstrable in mature VLRCs. Our analysis provides insight into the unique molecular strategies used for VLRC gene assembly and repertoire diversification.Proceedings of the National Academy of Sciences 03/2013; · 9.68 Impact Factor -
Article: Side chain modified peptide nucleic acids (PNA) for knock-down of six3 in medaka embryos.
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ABSTRACT: Synthetic antisense molecules have an enormous potential for therapeutic applications in humans. The major aim of such strategies is to specifically interfere with gene function, thus modulating cellular pathways according to the therapeutic demands. Among the molecules which can block mRNA function in a sequence specific manner are peptide nucleic acids (PNA). They are highly stable and efficiently and selectively interact with RNA. However, some properties of non-modified aminoethyl glycine PNAs (aegPNA) hamper their in vivo applications. We generated new backbone modifications of PNAs, which exhibit more hydrophilic properties. When we examined the activity and specificity of these novel phosphonic ester PNAs (pePNA) molecules in medaka (Oryzias latipes) embryos, high solubility and selective binding to mRNA was observed. In particular, mixing of the novel components with aegPNA components resulted in mixed PNAs with superior properties. Injection of mixed PNAs directed against the medaka six3 gene, which is important for eye and brain development, resulted in specific six3 phenotypes. PNAs are well established as powerful antisense molecules. Modification of the backbone with phosphonic ester side chains further improves their properties and allows the efficient knock down of a single gene in fish embryos.BMC Biotechnology 08/2012; 12:50. · 2.35 Impact Factor -
Article: A thymus candidate in lampreys.
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ABSTRACT: Immunologists and evolutionary biologists have been debating the nature of the immune system of jawless vertebrates--lampreys and hagfish--since the nineteenth century. In the past 50 years, these fish were shown to have antibody-like responses and the capacity to reject allografts but were found to lack the immunoglobulin-based adaptive immune system of jawed vertebrates. Recent work has shown that lampreys have lymphocytes that instead express somatically diversified antigen receptors that contain leucine-rich-repeats, termed variable lymphocyte receptors (VLRs), and that the type of VLR expressed is specific to the lymphocyte lineage: T-like lymphocytes express type A VLR (VLRA) genes, and B-like lymphocytes express VLRB genes. These clonally diverse anticipatory antigen receptors are assembled from incomplete genomic fragments by gene conversion, which is thought to be initiated by either of two genes encoding cytosine deaminase, cytosine deaminase 1 (CDA1) in T-like cells and CDA2 in B-like cells. It is unknown whether jawless fish, like jawed vertebrates, have dedicated primary lymphoid organs, such as the thymus, where the development and selection of lymphocytes takes place. Here we identify discrete thymus-like lympho-epithelial structures, termed thymoids, in the tips of the gill filaments and the neighbouring secondary lamellae (both within the gill basket) of lamprey larvae. Only in the thymoids was expression of the orthologue of the gene encoding forkhead box N1 (FOXN1), a marker of the thymopoietic microenvironment in jawed vertebrates, accompanied by expression of CDA1 and VLRA. This expression pattern was unaffected by immunization of lampreys or by stimulation with a T-cell mitogen. Non-functional VLRA gene assemblies were found frequently in the thymoids but not elsewhere, further implicating the thymoid as the site of development of T-like cells in lampreys. These findings suggest that the similarities underlying the dual nature of the adaptive immune systems in the two sister groups of vertebrates extend to primary lymphoid organs.Nature 02/2011; 470(7332):90-4. · 36.28 Impact Factor -
Article: Characterization of mononuclear phagocytic cells in medaka fish transgenic for a cxcr3a:gfp reporter.
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ABSTRACT: Chemokines and chemokine receptors are key evolutionary innovations of vertebrates. They are involved in morphogenetic processes and play an important role in the immune system. Based on an analysis of the chemokine receptor gene family in teleost genomes, and the expression patterns of chemokine receptor genes during embryogenesis and the wounding response in young larvae of Oryzias latipes, we identified the chemokine receptor cxcr3a as a marker of innate immune cells. Cells expressing cxcr3a were characterized in fish transgenic for a cxcr3a:gfp reporter. In embryos and larvae, cxcr3a-expressing cells are motile in healthy and damaged tissues, and phagocytic; the majority of these cells has the morphology of tissue macrophages, whereas a small fraction has a dendritic phenotype. In adults, cxcr3a-positive cells continue to specifically express myeloid-associate markers and genes related to antigen uptake and presentation. By light microscopy and ultrastructural analysis, the majority of cxcr3a-expressing cells has a dendritic phenotype, whereas the remainder resembles macrophage-like cells. After challenge of adult fish with bacteria or CpG oligonucleotides, phagocytosing cxcr3a-positive cells in the blood up-regulated il12p40 genes, compatible with their function as part of the mononuclear phagocytic system. Our results identify a marker of teleost mononuclear phagocytic cells and suggest a surprising degree of morphological and functional similarity between the innate immune systems of lower and higher vertebrates.Proceedings of the National Academy of Sciences 10/2010; 107(42):18079-84. · 9.68 Impact Factor -
Article: Identification of starmaker-like in medaka as a putative target gene of Pax2 in the otic vesicle.
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ABSTRACT: Otoliths in bony fishes are involved in the function of the ear in the senses of balance and hearing. In a large-scale random in situ hybridization screen of genes expressed in the medaka developing ear, we identified starmaker-like (stm-l) gene, a novel homologue of zebrafish starmaker and human dentine sialo-phosphoprotein (dspp) gene. Despite the absence of sequence similarity between these genes, here we describe their similar genomic structure and expression patterns hinting for a conserved function. In medaka fry, stm-l is expressed in various organs such as otoliths, teeth, gills, and kidney. Additionally, our results provide evidence that stm-l is a putative downstream target gene of Pax2 transcription factor and Pax2 itself has a promoting function in otolith formation.Developmental Dynamics 09/2009; 238(11):2860-6. · 2.54 Impact Factor -
Article: Induction of otic structures by canonical Wnt signalling in medaka.
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ABSTRACT: The Wnt family of signalling proteins is known to participate in multiple developmental decisions during embryogenesis. We misexpressed Wnt1 in medaka embryos and observed anterior truncations, similar to those described for ectopic activation of canonical Wnt signalling in other species. Interestingly, when we induced a heat-shock Wnt1 transgenic line exactly at 30% epiboly, we observed multiple ectopic otic vesicles in the truncated embryos. The vesicles then fused, forming a single large ear structure. These "cyclopic ears" filled the complete anterior region of the embryos. The ectopic induction of otic development can be explained by the juxtaposition of hindbrain tissue with anterior ectoderm. Fibroblast growth factor (Fgf) ligands are thought to mediate the otic-inducing properties of the hindbrain. However, signals different from Fgf3 and Fgf8 are necessary to explain the formation of the ectopic ear structures, suggesting that Wnt signalling is involved in the otic induction process in medaka.Archiv für Entwickelungsmechanik der Organismen 09/2009; 219(8):391-8. · 1.77 Impact Factor -
Article: Evolution of genetic networks underlying the emergence of thymopoiesis in vertebrates.
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ABSTRACT: About 500 million years ago, a new type of adaptive immune defense emerged in basal jawed vertebrates, accompanied by morphological innovations, including the thymus. Did these evolutionary novelties arise de novo or from elaboration of ancient genetic networks? We reconstructed the genetic changes underlying thymopoiesis by comparative genome and expression analyses in chordates and basal vertebrates. The derived models of genetic networks were experimentally verified in bony fishes. Ancestral networks defining circumscribed regions of the pharyngeal epithelium of jawless vertebrates expanded in cartilaginous fishes to incorporate novel genes, notably those encoding chemokines. Correspondingly, novel networks evolved in lymphocytes of jawed vertebrates to control the expression of additional chemokine receptors. These complementary changes enabled unprecedented Delta/Notch signaling between pharyngeal epithelium and lymphoid cells that was exploited for specification to the T cell lineage. Our results provide a framework elucidating the evolution of key features of the adaptive immune system in jawed vertebrates.Cell 07/2009; 138(1):186-97. · 32.40 Impact Factor -
Article: Genetic evidence for monozygotic twinning in medaka fish.
Molecular Reproduction and Development 03/2009; 76(5):430. · 2.53 Impact Factor -
Article: Rapid identification of PAX2/5/8 direct downstream targets in the otic vesicle by combinatorial use of bioinformatics tools.
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ABSTRACT: The pax2/5/8 genes belonging to the PAX family of transcription factors are key developmental regulators that are involved in the patterning of various embryonic tissues. More particularly, their function in inner ear specification has been widely described. However, little is known about the direct downstream targets and, so far, no global approaches have been performed to identify these target genes in this particular tissue. Here we present an original bioinformatics pipeline composed of comparative genomics, database querying and text mining tools, which is designed to rapidly and specifically discover PAX2/5/8 direct downstream targets involved in inner ear development. We provide evidence supported by experimental validation in medaka fish that brain 2 (POU domain, class 3, transcription factor 2), claudin-7, secretory pathway component sec31-like and meteorin-like precursor are novel direct downstream targets of PAX2/5/8. This study illustrates the power of extensive mining of public data repositories using bioinformatics methods to provide answers for a specific biological question. It furthermore demonstrates how the usage of such a combinatorial approach is advantageous for the biologist in terms of experimentation time and costs.Genome biology 11/2008; 9(10):R145. · 6.63 Impact Factor -
Article: Wnt/Axin1/beta-catenin signaling regulates asymmetric nodal activation, elaboration, and concordance of CNS asymmetries.
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ABSTRACT: Nodal activity in the left lateral plate mesoderm (LPM) is required to activate left-sided Nodal signaling in the epithalamic region of the zebrafish forebrain. Epithalamic Nodal signaling subsequently determines the laterality of neuroanatomical asymmetries. We show that overactivation of Wnt/Axin1/beta-catenin signaling during late gastrulation leads to bilateral epithalamic expression of Nodal pathway genes independently of LPM Nodal signaling. This is consistent with a model whereby epithalamic Nodal signaling is normally bilaterally repressed, with Nodal signaling from the LPM unilaterally alleviating repression. We suggest that Wnt signaling regulates the establishment of the bilateral repression. We identify a second role for the Wnt pathway in the left/right regulation of LPM Nodal pathway gene expression, and finally, we show that at later stages Axin1 is required for the elaboration of concordant neuroanatomical asymmetries.Neuron 09/2007; 55(3):393-405. · 14.74 Impact Factor -
Article: Distinct roles of Fgf8, Foxi1, Dlx3b and Pax8/2 during otic vesicle induction and maintenance in medaka.
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ABSTRACT: The development of the vertebrate inner ear is a complex process that has been investigated in several model organisms. In this work, we examined genetic interactions regulating early development of otic structures in medaka. We demonstrate that misexpression of Fgf8, Dlx3b and Foxi1 during early gastrulation is sufficient to produce ectopic otic vesicles. Combined misexpression strongly increases the appearance of this phenotype. By using a heat-inducible promoter we were furthermore able to separate the regulatory interactions among Fgf8, Foxi1, Dlx3b, Pax8 and Pax2 genes, which are active during different stages of early otic development. In the preplacodal stage we suggest a central position of Foxi1 within a regulatory network of early patterning genes including Dlx3b and Pax8. Different pathways are active after the placodal stage with Dlx3b playing a central role. There Dlx3b regulates members of the Pax-Six-Eya-Dach network and also strongly affects the early dorsoventral marker genes Otx1 and Gbx2.Developmental Biology 08/2007; 307(2):408-20. · 4.07 Impact Factor -
Article: The roles of Groucho/Tle in left-right asymmetry and Kupffer's vesicle organogenesis.
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ABSTRACT: The heart is the first organ to form and function in the vertebrate embryo. Furthermore, differences between the left and right sides of the embryo become first detectable during cardiac development. We observed strong cardiac laterality phenotypes in medaka embryos by manipulating Groucho protein activity. The phenotypes produced by misexpressing Tle4 and the dominant-negative Aes reveal a general effect of these corepressor proteins on left-right (LR) development. With the help of an inducible expression system, we were able to define temporally different phases for these effects. In an early phase during gastrulation, Groucho proteins regulate Brachyury expression in the dorsal forerunner cells, which later gives rise to the Kupffer's vesicle (KV). The interference of endogenous Groucho proteins by misexpression of Aes leads to KVs of reduced size, whereas overexpression of Tle4 results in enlarged KVs. The expression level of the cilia marker Lrd was also affected both positively and negatively from these treatments. In the late phase during somitogenesis, Groucho proteins regulate the asymmetric activities of Nodal and Lefty genes. Altering canonical Wnt signaling produced similar results in late embryos, however, this did not affect KV morphogenesis or Lrd expression in early embryos. Therefore, changes in Kupffer's vesicle morphogenesis and the laterality of visceral organs following alterations in Groucho corepressor levels demonstrate two distinct phases in which Groucho proteins help establish LR asymmetry in medaka fish.Developmental Biology 04/2007; 303(1):347-61. · 4.07 Impact Factor -
Article: Gbx2 and Otx2 interact with the WD40 domain of Groucho/Tle corepressors.
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ABSTRACT: One of the earliest organizational decisions in the development of the vertebrate brain is the division of the neural plate into Otx2-positive anterior and Gbx2-positive posterior territories. At the junction of these two expression domains, a local signaling center is formed, known as the midbrain-hindbrain boundary (MHB). This tissue coordinates or "organizes" the development of neighboring brain structures, such as the midbrain and cerebellum. Correct positioning of the MHB is thought to depend on mutual repression involving these two homeobox genes. Using a cell culture colocalization assay and coimmunoprecipitation experiments, we show that engrailed homology region 1 (eh1)-like motifs of both transcription factors physically interact with the WD40 domain of Groucho/Tle corepressor proteins. In addition, heat shock-induced expression of wild-type and mutant Otx2 and Gbx2 in medaka embryos demonstrates that Groucho is required for the repression of Otx2 by Gbx2. On the other hand, the repressive functions of Otx2 on Gbx2 do not appear to be dependent on corepressor interaction. Interestingly, the association of Groucho with Otx2 is also required for the repression of Fgf8 in the MHB. Therefore Groucho/Tle family members appear to regulate key aspects in the MHB development of the vertebrate brain.Molecular and Cellular Biology 02/2007; 27(1):340-51. · 5.53 Impact Factor -
Article: Expression of marker genes during early ear development in medaka.
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ABSTRACT: Induction of the otic placode involves a number of regulatory interactions. Early studies revealed that the induction of this program is initiated by instructive signals from the mesendoderm as well as from the adjacent hindbrain. Further investigations on the molecular level identified in zebrafish Fgf3, Fgf8, Foxi1, Pax8, Dlx3b and Dlx4b genes as key players during the induction phase. Thereafter an increasing number of genes participates in the regulatory interactions finally resulting in a highly structured sensory organ. Based on data from zebrafish we selected medaka genes with presumptive functions during early ear development for an expression analysis. In addition we isolated Foxi1 and Dlx3b gene fragments from embryonic cDNA. Altogether we screened the spatio-temporal distribution of more than 20 representative marker genes for otic development in medaka embryos, with special emphasis on the early phases. Whereas the spatial distribution of these genes is largely conserved between medaka and zebrafish, our comparative analysis revealed several differences, in particular for the timing of expression.Gene Expression Patterns 02/2007; 7(3):355-62. · 2.02 Impact Factor -
Article: Dynamic expression pattern of Nodal-related genes during left-right development in medaka.
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ABSTRACT: Nodal-related genes have been implicated in mesendoderm induction, establishment of embryonic axes, neural patterning and left-right development among vertebrates. Here we report the isolation of three Nodal-related genes in medaka (Oryzias latipes). Based on sequence analysis and in accordance to zebrafish orthologues we named the isolated genes Ndr1, Ndr2 and Spaw. Gene expression analysis throughout medaka development confirmed this assignment. Ndr1 and Ndr2 are detectable during gastrulation whereas Ndr2 and Spaw are expressed asymmetrically during somitogenesis. In accordance with its zebrafish orthologue, Spaw is expressed as the first asymmetric marker in the left lateral plate mesoderm (LPM) and Ndr2 displays asymmetric expression domains in the brain and the LPM. In general, the spatial distribution of Nodal transcripts resembles those reported for zebrafish, in case of Ndr2, however, we report a novel left-right asymmetry in the posterior paraxial mesoderm flanking the Kupffer's vesicle.Gene Expression Patterns 02/2007; 7(1-2):93-101. · 2.02 Impact Factor -
Article: Duplicated members of the Groucho/Tle gene family in fish.
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ABSTRACT: The highly conserved Groucho/Tle gene family has widespread functions during embryonic development and in adults. For mammalians, four full-length Tle paralogues are known, whereas the whole spectrum of this gene family in fish species has not been analysed yet. Most detailed data exist for medaka, where 3 Tle genes have been described, Tle1, Tle3, and Tle4. We now isolated 3 additional Tle genes from the medaka genome. Sequence analysis identifies these genes as Tle2a, Tle2b, and Tle3b. Database searches of genomic sequences revealed an identical set of Tle paralogues being present in distantly related fish species, indicating duplicated Tle2 and Tle3 genes for the complete teleost lineage. Like the previously analysed medaka Tle genes, the three new genes show a broad expression pattern during embryogenesis. Nevertheless, a detailed comparison of all six Tle genes reveals critical differences in certain aspects of their expression pattern. In particular, we concentrated on the activity of Tle genes during ear development and found Tle2a and Tle2b expressed in this sensory organ.Developmental Dynamics 10/2005; 234(1):143-50. · 2.54 Impact Factor -
Article: Groucho corepressor proteins regulate otic vesicle outgrowth.
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ABSTRACT: The Groucho/Tle family of corepressor proteins is known to regulate multiple developmental pathways. Applying the dominant-negative effect of the short member Aes, we demonstrate here a critical role of this gene family also for ear development. Misexpression of Aes in medaka embryos resulted in reduced size or loss of otic vesicles, whereas overexpression of the full-length Groucho protein Tle4 gave the opposite phenotype. These results are in close agreement with phenotypes observed for eye formation, suggesting a similar role for Groucho/Tle proteins in the developmental pathways of both sensory organs. Furthermore, by using the heat-inducible HSE promoter, we observed reversible branching of the embryonic axis upon Aes misexpression, indicating a transient duplication of the organizer. Groucho proteins, therefore, are critical for organizer maintenance.Developmental Dynamics 08/2005; 233(3):760-71. · 2.54 Impact Factor -
Article: An artificial promoter construct for heat-inducible misexpression during fish embryogenesis.
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ABSTRACT: Beside spatial distribution, timing of gene expression is a key parameter controlling gene function during embryonic development. Gain-of-function experiments can therefore have quite opposing results, depending on the time of gene activation. Induction techniques are necessary to control timing in these experiments from outside of the organism. Natural heat shock promoters constitute a simple inducible misexpression system, the main disadvantage is a high background level of expression. We present here a new heat stress-inducible bidirectional promoter consisting of multimerized heat shock elements (HSE). The simplified architecture of this promoter largely improves the properties needed for an efficient induction system: dramatically reduced background activity, improved inducibility, and loss of all tissue specific components. Based on this new artificial promoter, we present a transient induction system for fish embryos. Application of this new induction system for Fgf8 misexpression during embryonic development reveals different windows of competence during eye development. A dramatic early phenotype resulting in loss of the eyes is observed for conventional mRNA injection. Later activation, by using our inducible promoter, uncovers different eye phenotypes like cyclopic eyes. Even after 14 days, an efficient heat stress response could be evoked in the injected embryos. The HSE promoter therefore represents a new artificial heat shock promoter with superior properties, making possible transient experiments with inducible misexpression at various stages of development.Developmental Biology 08/2004; 271(2):416-30. · 4.07 Impact Factor
Top co-authors
Top Journals
Institutions
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2004–2012
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University of Veterinary Medicine in Vienna
- • Department für Biomedizinische Wissenschaften
- • Institut für Tierzucht und Genetik
Vienna, Vienna, Austria
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2010–2011
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Max-Planck-Institut für Immunbiologie und Epigenetik
- Department of Developmental Immunology
Freiburg, Baden-Wuerttemberg, Germany
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2009
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Max-Planck-Institut für Entwicklungsbiologie
Tübingen, Baden-Wuerttemberg, Germany
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