495 research outputs found

    Lbx2 regulates formation of myofibrils

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    <p>Abstract</p> <p>Background</p> <p>Skeletal muscle differentiation requires assembly of contractile proteins into organized myofibrils. The <it>Drosophila ladybird homeobox </it>gene (<it>lad</it>) functions in founder cells of the segmental border muscle to promote myoblast fusion and muscle shaping. Tetrapods have two homologous genes (<it>Lbx</it>). Lbx1 functions in migration and/or proliferation of hypaxial myoblasts, whereas the function of Lbx2 is poorly understood.</p> <p>Results</p> <p>To elucidate the role of Lbx in vertebrate myogenesis, we examined Lbx function in zebrafish. Zebrafish <it>lbx2 </it>transcripts appear in newly formed paraxial mesoderm and become restricted to adaxial cells, precursors of slow muscle. Slow muscles lose <it>lbx2 </it>expression as they differentiate, while a subset of differentiating fast muscle cells transiently expresses <it>lbx2</it>. Fin and hyoid muscle express <it>lbx2 </it>later. In contrast, <it>lbx1b </it>expression first appears lateral to the somites at late segmentation stages and is later restricted to fin muscle. Morpholino knockdown of Lbx1b and Lbx2 suppresses hypaxial muscle development. Moreover, knockdown of Lbx2 results in malformation of muscle fibers and reduced fusion of fast precursors, although no obvious effects on induction or specification are observed. Expression of myofilament genes, including <it>actin </it>and <it>myosin</it>, requires the engrailed repressor domain of Lbx2.</p> <p>Conclusion</p> <p>Our results elucidate a new function of Lbx2 as a regulator of myofibril formation.</p

    PhOTO Zebrafish: A Transgenic Resource for In Vivo Lineage Tracing during Development and Regeneration

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    Background: Elucidating the complex cell dynamics (divisions, movement, morphological changes, etc.) underlying embryonic development and adult tissue regeneration requires an efficient means to track cells with high fidelity in space and time. To satisfy this criterion, we developed a transgenic zebrafish line, called PhOTO, that allows photoconvertible optical tracking of nuclear and membrane dynamics in vivo. Methodology: PhOTO zebrafish ubiquitously express targeted blue fluorescent protein (FP) Cerulean and photoconvertible FP Dendra2 fusions, allowing for instantaneous, precise targeting and tracking of any number of cells using Dendra2 photoconversion while simultaneously monitoring global cell behavior and morphology. Expression persists through adulthood, making the PhOTO zebrafish an excellent tool for studying tissue regeneration: after tail fin amputation and photoconversion of a ~100µm stripe along the cut area, marked differences seen in how cells contribute to the new tissue give detailed insight into the dynamic process of regeneration. Photoconverted cells that contributed to the regenerate were separated into three distinct populations corresponding to the extent of cell division 7 days after amputation, and a subset of cells that divided the least were organized into an evenly spaced, linear orientation along the length of the newly regenerating fin. Conclusions/Significance: PhOTO zebrafish have wide applicability for lineage tracing at the systems-level in the early embryo as well as in the adult, making them ideal candidate tools for future research in development, traumatic injury and regeneration, cancer progression, and stem cell behavior

    Flavones induce neutrophil apoptosis by down-regulation of Mcl-1 via a proteasomal-dependent pathway

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    Neutrophil apoptosis and subsequent nonphlogistic clearance by surrounding phagocytes are key to the successful resolution of neutrophilic inflammation, with dysregulated apoptosis reported in multiple human inflammatory diseases. Enhancing neutrophil apoptosis has proresolution and anti-inflammatory effects in preclinical models of inflammation. Here we investigate the ability of the flavones apigenin, luteolin, and wogonin to induce neutrophil apoptosis in vitro and resolve neutrophilic inflammation in vivo. Human neutrophil apoptosis was assessed morphologically and by flow cytometry following incubation with apigenin, luteolin, and wogonin. All three flavones induced time- and concentration-dependent neutrophil apoptosis (apigenin, EC(50)=12.2 μM; luteolin, EC(50)=14.6 μM; and wogonin, EC(50)=28.9 μM). Induction of apoptosis was caspase dependent, as it was blocked by the broad-spectrum caspase inhibitor Q-VD-OPh and was associated with both caspase-3 and caspase-9 activation. Flavone-induced apoptosis was preceded by down-regulation of the prosurvival protein Mcl-1, with proteasomal inhibition preventing flavone-induced Mcl-1 down-regulation and apoptosis. The flavones abrogated the survival effects of mediators that prolong neutrophil life span, including lipoteichoic acid, peptidoglycan, dexamethasone, and granulocyte-macrophage colony stimulating factor, by driving apoptosis. Furthermore, wogonin enhanced resolution of established neutrophilic inflammation in a zebrafish model of sterile tissue injury. Wogonin-induced resolution was dependent on apoptosis in vivo as it was blocked by caspase inhibition. Our data show that the flavones induce neutrophil apoptosis and have potential as neutrophil apoptosis-inducing anti-inflammatory, proresolution agents.—Lucas, C. D., Allen, K. C., Dorward, D. A., Hoodless, L. J., Melrose, L. A., Marwick, J. A., Tucker, C. S., Haslett, C., Duffin, R., Rossi, A. G. Flavones induce neutrophil apoptosis by down-regulation of Mcl-1 via a proteasomal-dependent pathway

    Dcc Regulates Asymmetric Outgrowth of Forebrain Neurons in Zebrafish

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    The guidance receptor DCC (deleted in colorectal cancer) ortholog UNC-40 regulates neuronal asymmetry development in Caenorhabditis elegans, but it is not known whether DCC plays a role in the specification of neuronal polarity in vertebrates. To examine the roles of DCC in neuronal asymmetry regulation in vertebrates, we studied zebrafish anterior dorsal telencephalon (ADt) neuronal axons. We generated transgenic zebrafish animals expressing the photo-convertible fluorescent protein Kaede in ADt neurons and then photo-converted Kaede to label specifically the ADt neuron axons. We found that ADt axons normally project ventrally. Knock down of Dcc function by injecting antisense morpholino oligonucleotides caused the ADt neurons to project axons dorsally. To examine the axon projection pattern of individual ADt neurons, we labeled single ADt neurons using a forebrain-specific promoter to drive fluorescent protein expression. We found that individual ADt neurons projected axons dorsally or formed multiple processes after morpholino knock down of Dcc function. We further found that knock down of the Dcc ligand, Netrin1, also caused ADt neurons to project axons dorsally. Knockdown of Neogenin1, a guidance receptor closely related to Dcc, enhanced the formation of aberrant dorsal axons in embryos injected with Dcc morpholino. These experiments provide the first evidence that Dcc regulates polarized axon initiation and asymmetric outgrowth of forebrain neurons in vertebrates

    The Mych Gene Is Required for Neural Crest Survival during Zebrafish Development

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    Background: Amomg Myc family genes, c-Myc is known to have a role in neural crest specification in Xenopus and in craniofacial development in the mouse. There is no information on the function of other Myc genes in neural crest development, or about any developmental role: of zebrafish Myc genes. Principal Findings: We isolated the zebrafish mych (myc homologue) gene. Knockdown of mych leads to sever defects in craniofacial development and in certain other tissues including the eye. These phenotypes appear to be caused by cell death in the neural crest and in the eye field in the anterior brain. Significance: Mych is a novel factor required for neural crest cell survival in zebrafish

    ZFIN: enhancements and updates to the zebrafish model organism database

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    ZFIN, the Zebrafish Model Organism Database, http://zfin.org, serves as the central repository and web-based resource for zebrafish genetic, genomic, phenotypic and developmental data. ZFIN manually curates comprehensive data for zebrafish genes, phenotypes, genotypes, gene expression, antibodies, anatomical structures and publications. A wide-ranging collection of web-based search forms and tools facilitates access to integrated views of these data promoting analysis and scientific discovery. Data represented in ZFIN are derived from three primary sources: curation of zebrafish publications, individual research laboratories and collaborations with bioinformatics organizations. Data formats include text, images and graphical representations. ZFIN is a dynamic resource with data added daily as part of our ongoing curation process. Software updates are frequent. Here, we describe recent additions to ZFIN including (i) enhanced access to images, (ii) genomic features, (iii) genome browser, (iv) transcripts, (v) antibodies and (vi) a community wiki for protocols and antibodies

    The Zebrafish Information Network: the zebrafish model organism database provides expanded support for genotypes and phenotypes

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    The Zebrafish Information Network (ZFIN, http://zfin.org), the model organism database for zebrafish, provides the central location for curated zebrafish genetic, genomic and developmental data. Extensive data integration of mutant phenotypes, genes, expression patterns, sequences, genetic markers, morpholinos, map positions, publications and community resources facilitates the use of the zebrafish as a model for studying gene function, development, behavior and disease. Access to ZFIN data is provided via web-based query forms and through bulk data files. ZFIN is the definitive source for zebrafish gene and allele nomenclature, the zebrafish anatomical ontology (AO) and for zebrafish gene ontology (GO) annotations. ZFIN plays an active role in the development of cross-species ontologies such as the phenotypic quality ontology (PATO) and the gene ontology (GO). Recent enhancements to ZFIN include (i) a new home page and navigation bar, (ii) expanded support for genotypes and phenotypes, (iii) comprehensive phenotype annotations based on anatomical, phenotypic quality and gene ontologies, (iv) a BLAST server tightly integrated with the ZFIN database via ZFIN-specific datasets, (v) a global site search and (vi) help with hands-on resources

    A Forward Chemical Screen in Zebrafish Identifies a Retinoic Acid Derivative with Receptor Specificity

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    Background: Retinoids regulate key developmental pathways throughout life, and have potential uses for differentiation therapy. It should be possible to identify novel retinoids by coupling new chemical reactions with screens using the zebrafish embryonic model. Principal Findings: We synthesized novel retinoid analogues and derivatives by amide coupling, obtaining 80–92% yields. A small library of these compounds was screened for bioactivity in living zebrafish embryos. We found that several structurally related compounds significantly affect development. Distinct phenotypes are generated depending on time of exposure, and we characterize one compound (BT10) that produces specific cardiovascular defects when added 1 day post fertilization. When compared to retinoic acid (ATRA), BT10 shows similar but not identical changes in the expression pattern of embryonic genes that are known targets of the retinoid pathway. Reporter assays determined that BT10 interacts with all three RAR receptor sub-types, but has no activity for RXR receptors, at all concentrations tested. Conclusions: Our screen has identified a novel retinoid with specificity for retinoid receptors. This lead compound may be useful for manipulating components of retinoid signaling networks, and may be further derivatized for enhanced activity
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