SEARCH · PubMed Health

Results for “zebrafish”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,009 records · Page 56Linked to original sources

PubMed2005

Molecular cloning of three zebrafish lin7 genes and their expression patterns in the retina.

The vertebrate retina develops from an undifferentiated sheet of neuroepithelial cells, whose differentiation requires the generation and maintenance of the correct cellular polarity. To examine the role of cell polarity in retinal development, we cloned three zebrafish lin7 genes (lin7a, lin7b, and lin7c), which each encodes a protein candidate that is required for generation/maintenance of neuroepithelial cell junctions. These three zebrafish Lin7 proteins share over 78% amino acid identity and contain both L27 and PDZ domains that are present in all Lin7 homologs. Immunoblots revealed that the Lin7b and Lin7c proteins were first expressed in the developing eye by 24hr postfertilization (hpf), while Lin7a was not detected in the eye until 72 hpf. At 33 hpf, the Lin7 proteins localized at, or slightly apical of, the actin-associated adherens junctions in the retinal neuroepithelium. This subcellular distribution required the expression of the Nok protein. In the absence of Nok, the Lin7 proteins failed to localize to either the ectopic adherens junctions or the cell membrane. At 4 days postfertilization, in situ hybridisation revealed that all three lin7 genes were expressed in both the ganglion cell layer and the bipolar cell region of the inner nuclear layer. The lin7a gene was also expressed in the amacrine and horizontal cell regions of the inner nuclear layer, while lin7c was also expressed in the outer nuclear layer. In the adult retina, where Lin7a is the predominant form expressed, the Lin7 proteins were localized to the outer and inner plexiform layers, the bipolar and horizontal cells of the inner nuclear layer, and the ganglion cells. These results suggest that the three zebrafish Lin7 proteins possess partially redundant, yet essential, roles in retinal development.

Animals↗

PubMed2006

Constitutive activation of zebrafish Stat5 expands hematopoietic cell populations in vivo.

OBJECTIVE: Constitutive activation of Stat5 has been observed in a variety of malignancies, particularly myeloid leukemias. To directly investigate the in vivo consequences of Stat5 perturbation, we expressed constitutively active forms in zebrafish. METHODS: We generated mutants of the zebrafish stat5.1 protein (N646H, H298R/N714F, and N714F) based on previously identified constitutively active mutants of murine Stat5a. The in vitro properties of these mutants were determined using phosphorylation-specific antibodies and luciferase reporter assays, and their in vivo effects were analyzed through microinjection of zebrafish embryos. RESULTS: Two of these stat5.1 mutants (N646H and H298R/N714F) showed increased tyrosine phosphorylation and transactivation activity compared to the wild-type protein. Expression of either mutant led to a range of hematological perturbations, which were more pronounced for the H298R/N714F mutant. Interestingly, expression of wild-type also produced generally similar phenotypes. Further analysis showed that expression of the H298R/N714F mutant led to increased numbers of early and late myeloid cells, erythrocytes, and B cells. Some nonhematopoietic developmental perturbations were also observed, but these were equally prominent with wild-type or mutant forms. CONCLUSION: These data implicate Stat5 activity as a direct critical regulator of hematological cell proliferation, suggesting a causal role for constitutively-active Stat5 in the etiology of hematological malignancies.

Amino Acid Substitution↗

PubMed2006

Snx5, as a Mind bomb-binding protein, is expressed in hematopoietic and endothelial precursor cells in zebrafish.

Notch signaling has an evolutionarily conserved function for cell fate determination and stem cell maintenance. Previously, we identified a novel component of the Notch signaling pathway in zebrafish, mind bomb, which encodes an E3 ubiquitin ligase essential for Notch signal activation. Further studies showed that Mind bomb(-/-) mouse embryos exhibited pan-Notch phenotypes in various tissues, suggesting that Mind bomb function is conserved in mammals. Therefore we sought to understand the various molecular partners of Mind bomb using yeast two-hybrid screening. In this search we identified Sorting nexin 5 (Snx5) as a novel interacting partner of Mind bomb. Furthermore we demonstrated that Snx5 colocalizes with Mind bomb in early endosomal compartments, suggesting that Snx5 is important for Mind bomb trafficking. In addition, we identified zebrafish orthologue of Snx5 and showed that snx5 is predominantly expressed in hematopoietic and endothelial precursor cells in zebrafish. We also found defects in hematopoiesis and blood vessel development in snx5 morpholino-injected embryos. Taken together, we show that Snx5, a novel interacting partner of Mind bomb, may have an essential role for cell fate determination in early development.

Animals↗

PubMed2004

Novel cis-element in intron 1 represses somite expression of zebrafish myf-5.

Myf-5 is a basic helix-loop-helix (bHLH) transcription factor that controls muscle differentiation. During early embryogenesis, myf-5 expression is transient, somite- and stage-specific. However, the negative regulation of myf-5 is poorly understood. We constructed a plasmid [(-9977/-1)/E1/I1/E2/GFP] that contains the sequence -9977 to -1, exon 1 (E1), intron 1 (I1), and exon 2 (E2) of zebrafish (Danio rerio) myf-5 and a reporter GFP gene. This plasmid was microinjected into zebrafish zygotes. Surprisingly, the somite-specific expression rate of reporter GFP in the transgenic embryos was extremely low (2%, n=392), compared to that of (-9977/-1)/GFP (92%, n=210). Dramatic repression of myf-5 expression was also observed in embryos microinjected with plasmids in which the sequence -8600/-1, -2937/-1 or -290/-1 was linked to E1/I1/E2/GFP. Thus, intron 1 contains a silencer that specifically represses the activity of myf-5. Functional analysis of intron 1 showed a strong, negative, cis-regulatory element was located at +502/+835. Its function was orientation- and position-dependent. The repressive capability of this silencer was completely dependent on two core motifs, IE1 (+502/+527) and IE2 (+816/+835), and a 156-bp spanning sequence that lies between them. This is the first study to identify a novel, cis-acting silencer in intron 1 that is crucial to negatively regulating zebrafish myf-5 expression.

Animals↗

PubMed2005

Myostatin gene silenced by RNAi show a zebrafish giant phenotype.

Myostatin is a member of the transforming growth factor-beta (TGF-beta) family that functions as a negative regulator of skeletal muscle development and growth. Recently, it has been reported that the transgenic zebrafish expressing myostatin prodomain exhibited an increased number of fiber in skeletal muscle. Other novel results suggest that myostatin plays a mayor role during myogenesis, apart from inhibition of proliferation as well as differentiation. We have investigated the ability of double-stranded RNA (dsRNA) to inhibit myostatin function in the zebrafish. By microinjection dsRNA, corresponding to biologically active C-terminal domain from aminoacid 268 to end codon of tilapia myostatin protein, we produced an increased body mass in treated fish. The dsRNA injection in early development stage in zebrafish produced hyperplasia or hypertrophy. In addition, the interference of gene function showed a strong dependence on the amount of dsRNA.

Animals↗

PubMed2006

Functional genome analysis indicates loss of 17beta-hydroxysteroid dehydrogenase type 2 enzyme in the zebrafish.

Among the family of 17beta-hydroxysteroid dehydrogenases, the type 2 (17beta-HSD 2) is the main enzyme responsible for inactivation of estrogens and androgens, catalyzing the oxidation of the C17 hydroxyl group. 17beta-HSD 2 has been studied only in mammals, its occurrence and function in other vertebrates hardly known. We investigated the presence of homologs in non-mammalian species and found sequences of 17beta-HSD 2 and its closest homolog 11beta-HSD 2 in zebrafish (Danio rerio), Takifugu rubripes, Tetraodon nigroviridis, Xenopus tropicalis and chicken databases. Furthermore, we cloned zebrafish 17beta-HSD 2 from ovarian tissue and found high expression also in the testis of adult fish and throughout embryogenesis. The enzyme, though, is inactive likely due to a non-sense N-terminal region including a dysfunctional cofactor binding motif. Replacement of the affected part by the corresponding human 17beta-HSD 2 sequence fully restored enzymatic activity. Comparison of all retrieved 17beta-HSD 2 sequences indicates that this functional loss may have occurred only in zebrafish, where steroid inactivation at position C17 seems to pursue without the protein studied. The closely related 11beta-HSD 2 is unlikely to substitute for 17beta-HSD 2 since in our hands it did not catalyze the respective oxidation of testosterone or estradiol.

17-Hydroxysteroid Dehydrogenases↗

PubMed2006

Aromatase in zebrafish: a potential target for endocrine disrupting chemicals.

In zebrafish, two isoforms of the aromatase gene exist, namely cyp19a1 and cyp19a2, expressed predominantly in the gonads and brain, respectively. In this study, we focus on characterizing the specificity of antibodies against the aromatase isoforms, and on (xeno)estrogen-induced changes of individual cyp19a2 mRNA concentrations in the brains of adult male zebrafish. Among three polyclonal antibodies studied, the one against CYP19A2 was found to be specific in Western blots and immunohistochemistry. Real-time RT-PCR analyses revealed strong interindividual variation of cyp19a2 levels in the brains of adult male zebrafish. After a three-week-exposure to (xeno)estrogens, mean values of cyp19a2 mRNA levels tended to increase, with significant induction at 200 ng 17beta-estradiol/L, but interindividual variation of cyp19a2 expression was maintained.

Animals↗

PubMed2006

Differential expression of two tropoelastin genes in zebrafish.

Elastin is the extracellular matrix protein responsible for properties of extensibility and elastic recoil in large blood vessels, lung and skin of most vertebrates. Elastin is synthesized as a monomer, tropoelastin, but is rapidly transformed into its final polymeric form in the extracellular matrix. Until recently information on sequence and developmental expression of tropoelastins was limited to mammalian and avian species. We have recently identified and characterized two expressed tropoelastin genes in zebrafish. This was the first example of a species with multiple tropoelastin genes, raising the possibility of differential expression and function of these tropoelastins in elastic tissues of the zebrafish. Here we have investigated the temporal expression and tissue distribution of the two tropoelastin genes in developing and adult zebrafish. Expression was detected early in skeletal cartilage structures of the head, in the developing outflow tract of the heart, including the bulbus arteriosus and the ventral aorta, and in the wall of the swim bladder. While the temporal pattern of expression was similar for both genes, the upregulation of eln2 was much stronger than that of eln1. In general, both genes were expressed and their gene products deposited in most of the elastic tissues examined, with the notable exception of the bulbus arteriosus in which eln2 expression and its gene product was predominant. This finding may represent a sub-specialization of eln2 to provide the unique architecture of elastin and the specific mechanical properties required by this organ.

Amino Acid Sequence↗

PubMed2004

Ectopic expression and knockdown of a zebrafish sox21 reveal its role as a transcriptional repressor in early development.

Sox proteins are DNA-binding proteins belonging to the HMG box superfamily and they play key roles in animal embryonic development. Zebrafish Sox21a is part of group B Sox proteins and its chicken and mouse orthologs have been described as transcriptional repressor and activator, respectively, in two different target gene contexts. Zebrafish sox21a is present as a maternal transcript in the oocyte and is mainly expressed at the developing midbrain-hindbrain boundary from the onset of neurulation. In order to understand its role in vivo, we ectopically expressed sox21a by microinjection. Ectopic expression of full length sox21a leads to dorsalization of the embryos. A subset of the dorsalized embryos shows a partial axis splitting, and hence an ectopic neural tube, as an additional phenotype. At gastrulation, injected embryos show expansion of the expression domains of organizer-specific genes, such as chordin and goosecoid. Molecular markers used in somitogenesis highlight that sox21a-injected embryos have shortened AP axis, undulating axial structures, enlarged or even radialized paraxial territory. The developmental abnormalities caused by ectopic expression of sox21a are suggestive of defects in convergence-extension morphogenetic movements. Antisense morpholino oligonucleotides, designed to functionally knockdown sox21a, cause ventralization of the embryos. Moreover, gain-of-function experiments with chimeric constructs, where Sox21a DNA-binding domain is fused to a transcriptional activator (VP16) or repressor (EnR) domain, suggests that zebrafish Sox21a acts as a repressor in dorso-ventral patterning.

Animals↗

PubMed2004

Zebrafish gcmb is required for pharyngeal cartilage formation.

The glial cells missing (gcm) gene in Drosophila encodes a GCM-motif transcription factor that functions as a binary switch to select between glial and neuronal cell fates. To understand the function of gcm in vertebrates, we isolated the zebrafish gcmb and analyzed the function of this gene using antisense morpholino oligonucleotides against gcmb mRNA (gcmb-MO) and transgenic overexpression. Zebrafish gcmb is expressed in the pharyngeal arch epithelium and in cells of the macrophage lineage. gcmb-MO-injected larvae show significantly reduced branchial arch cartilages. fgf3-MO-injected larvae display a similar phenotype to that of gcmb-MO-injected larvae with respect to the lack of pharyngeal cartilage formation. In addition, gcmb expression in the pharyngeal arches is down-regulated in fgf3-MO-injected larvae. The gcmb transgenic larvae show a protrusion of the lower jaw and abnormal spatial arrangement of the pharyngeal cartilage elements. These results suggest that gcmb is required for normal pharyngeal cartilage formation in zebrafish and that its expression is dependent on fgf3 activity.

Amino Acid Sequence↗

PubMed2004

Touchtone promotes survival of embryonic melanophores in zebrafish.

An outstanding problem in the study of vertebrate development is the identification of the genes that direct neural crest precursor cells to adopt and maintain specific differentiated cell fates. In an effort to identify such genes, we have carried out a mutagenesis screen in zebrafish and isolated mutants that lack neural crest-derived melanophores. In this manuscript we describe the phenotype of one such mutant, touchtone(b722) (tct), and the map position of the gene it defines. Analysis of expression of dopachrome tautomerase (dct) and microphthalmia (mitfa) suggests that melanophore precursors are specified normally in homozygous tct mutants. However, differentiated melanophores are pale, small, and about half of them have disappeared by 48 h of development, apparently by cell death. We show that melanophores require Tct function cell autonomously. Signals from the receptor tyrosine kinase receptor C-kit are essential for survival of melanophores in zebrafish and mammals. However, differences in the phenotypes of tct and c-kit homozygous mutants, and an absence of interaction between c-kit and tct heterozygotes, suggest that Tct functions independently of the C-kit pathway. Other neural crest derivatives, including other pigment cell types, appear normal in tct mutants. Interestingly, tct mutant embryos undergo a temporary period of near complete paralyzis during the second day of development, although markers of axons of motor and sensory neurons look normal in this period. A fraction of tct(b722) mutants survive to adulthood, but mutant adults are small, indicating a role for Tct in post-larval growth. The tct gene maps to a small interval near a telomere of chromosome 18. Thus, we have identified a zebrafish gene that when mutated produces semi-viable offspring and that may serve as a model of human diseases that have both pigmentation and neurological symptoms.

Animals↗

PubMed2004

Melanophore sublineage-specific requirement for zebrafish touchtone during neural crest development.

The specification, differentiation and maintenance of diverse cell types are of central importance to the development of multicellular organisms. The neural crest of vertebrate animals gives rise to many derivatives, including pigment cells, peripheral neurons, glia and elements of the craniofacial skeleton. The development of neural crest-derived pigment cells has been studied extensively to elucidate mechanisms involved in cell fate specification, differentiation, migration and survival. This analysis has been advanced considerably by the availability of large numbers of mouse and, more recently, zebrafish mutants with defects in pigment cell development. We have identified the zebrafish mutant touchtone (tct), which is characterized by the selective absence of most neural crest-derived melanophores. We find that although wild-type numbers of melanophore precursors are generated in the first day of development and migrate normally in tct mutants, most differentiated melanophores subsequently fail to appear. We demonstrate that the failure in melanophore differentiation in tct mutant embryos is due at least in part to the death of melanoblasts and that tct function is required cell autonomously by melanoblasts. The tct locus is located on chromosome 18 in a genomic region apparently devoid of genes known to be involved in melanophore development. Thus, zebrafish tct may represent a novel as well as selective regulator of melanoblast development within the neural crest lineage. Further, our results suggest that, like other neural crest-derived sublineages, melanogenic precursors constitute a heterogeneous population with respect to genetic requirements for development.

Alleles↗

PubMed2006

Knockdown of zebrafish crim1 results in a bent tail phenotype with defects in somite and vascular development.

The Crim1 gene encodes a transmembrane protein containing six cysteine-rich repeats similar to those found in the BMP antagonist, chordin (chd). To investigate its physiological role, zebrafish crim1 was cloned and shown to be both maternally and zygotically expressed during zebrafish development in sites including the vasculature, intermediate cell mass, notochord, and otic vesicle. Bent or hooked tails with U-shaped somites were observed in 85% of morphants from 12 hpf. This was accompanied by a loss of muscle pioneer cells. While morpholino knockdown of crim1 showed some evidence of ventralisation, including expansion of the intermediate cell mass (ICM), reduction in head size bent tails and disruption to the somites and notochord, this did not mimic the classically ventralised phenotype, as assessed by the pattern of expression of the dorsal markers chordin, otx2 and the ventral markers eve1, pax2.1, tal1 and gata1 between 75% epiboly and six-somites. From 24 hpf, morphants displayed an expansion of the ventral mesoderm-derived ICM, as evidenced by expansion of tal1, lmo2 and crim1 itself. Analysis of the crim1 morphant phenotype in Tg(fli:EGFP) fish showed a clear reduction in the endothelial cells forming the intersegmental vessels and a loss of the dorsal longitudinal anastomotic vessel (DLAV). Hence, the primary role of zebrafish crim1 is likely to be the regulation of somitic and vascular development.

Animals↗

PubMed2006

Zebrafish Numb homologue: phylogenetic evolution and involvement in regulation of left-right asymmetry.

Numb and its homologue, Numb-like (Numbl), play important roles in mammalian development, but their role in embryonic development of lower vertebrates remains unknown. We cloned a zebrafish numb homologue (znumb) by searching database. znumb shows approximately 60% identity with mammalian Numb orthologs. Interstingly, znumb lacks two specific sequence motifs unique to Numbl orthologs. However, chromosomal localization of znumb gene revealed colinearity with genes located around mouse and human Numbl genes. Furthermore, multi-species comparisons of conserved phosphotyrosine-binding (PTB) domain sequences in Numb and Numbl proteins suggest that znumb is more closely related to Numbl than Numb. znumb mRNA was expressed in a wide variety of zebrafish adult tissues. Overexpression of znumb in embryos resulted in an absence, or reversal, of the normal leftward shift of the developing heart tube. Furthermore, no or bi-lateral transcripts of lefty2 were observed in znumb-expressing embryos, suggesting that the Notch signaling was essential for left-right field formation and maintenance in zebrafish, and that znumb perturbed this process through down-regulation of endogenous Notch signaling.

Animals↗

PubMed2006

An essential role for zebrafish Fgfrl1 during gill cartilage development.

The vertebrate craniofacial skeleton develops via a complex process involving signaling cascades in all three germ layers. Fibroblast growth factor (FGF) signaling is essential for several steps in pharyngeal arch development. In zebrafish, Fgf3 and Fgf8 in the mesoderm and hindbrain have an early role to pattern the pouch endoderm, influencing craniofacial integrity. Endodermal FGF signaling is required for the differentiation and survival of postmigratory neural crest cells that form the pharyngeal skeleton. We identify a novel role for zebrafish Fgf receptor-like 1a (Fgfrl1a) that is indispensable during gill cartilage development. We show that depletion of Fgfrl1a is sufficient to abolish cartilage derivatives of the ceratobranchials. Using an Fgfrl1a-deficient model, we analyzed expression of genes critical for chondrogenesis in the different compartments of the developing pharyngeal arch. Fgfrl1a-depleted animals demonstrate typical neural crest specification and migration to populate the arch primordia as well as normal pouch segmentation. However, in the absence of Fgfrl1a, larvae fail to express the transcription factor glial cells missing 2 (gcm2), a gene necessary for cartilage and gill filament formation, in the ectodermal lining of the branchial arches. In addition, two transcription factors essential for chondrogenesis, sox9a and runx2b, fail to express within the mesenchymal condensations of the branchial arches. A duplicate zebrafish gene, fgfrl1b, has now been identified. We show that Fgfrl1b is also required for proper formation of all ventral cartilage elements and acts cooperatively with Fgfrl1a during gill cartilage formation.

Amino Acid Sequence↗

PubMed2004

Identification and developmental expression pattern of van gogh-like 1, a second zebrafish strabismus homologue.

Cell movement plays a central role in both normal embryogenesis and the development of diseases such as cancer. Therefore, identification and analysis of proteins controlling cell movement is of special importance. The zebrafish trilobite locus encodes a Van Gogh/Strabismus homologue, which regulates diverse cell migratory behaviors during embryogenesis. Trilobite is most similar to human Van Gogh-like 2 (VANGL2)/Strabismus 1 and mouse Loop-tail associated protein/Lpp1. Both human and mouse genomes encode a second Strabismus homologue referred to as VANGL1/Strabismus 2 and Lpp2, respectively. This prompted us to ask whether another van gogh/strabismus gene, one more closely related to human VANGL1, exists in the zebrafish genome. This paper describes the identification of zebrafish vangl1 and provides the first spatiotemporal expression and functional analysis of a vertebrate vangl1 homologue. Our data indicate that vangl1 and trilobite/vangl2 are expressed in largely non-overlapping domains during embryogenesis. Injection of synthetic vangl1 RNA partially suppressed the gastrulation defect in trilobite mutant embryos, suggesting that Vangl1 and Trilobite/Vangl2 have similar biochemical activities.

Amino Acid Sequence↗

PubMed2004

Expression and mapping of duplicate neuropilin-1 and neuropilin-2 genes in developing zebrafish.

Previously, we described the isolation and characterization of the first zebrafish neuropilin gene, which we now call nrp1a, and found its protein to be a mediator of vascular endothelial growth factor (VEGF)-dependent angiogenesis [Proc. Natl Acad. Sci. USA 99 (2002) 10470]. Subsequently, we have isolated three other full-length neuropilin genes (nrp1b, nrp2a, and nrp2b) and find that they map to independent zebrafish linkage groups. The nrp1s and nrp2s had differential spatio-temporal gene expression profiles with nrp1a being most prominent in the gut, brain, retina, hypochord, motorneurons, fin bud and mandibular cartilage, nrp1b in the brain, dorsal aorta, melanophores, ventral fin, and heart, nrp2a in the brain, retina, heart, and caudal vessels, and nrp2b in the brain, retina, gut, fin bud, melanophores, heart, and caudal vessels. In addition, we have identified an alternatively-spliced transcript of the nrp1b gene (denoted as nrp1b(s)) which is predicted to encode a soluble form of Nrp1b, containing only the a, b, and c extracellular domains. Transcript expression of nrp1b(s) was different from full-length nrp1b transcript, with prominence in the brain, developing mouth, heart, and fin bud. The NRP1s were tested for VEGF-binding ability. Both 125 kDa Nrp1a and 145 kDa Nrp1b bound 125I-labelled VEGFA165. In summary, two nrp1 and two nrp2 genes, with expression patterns similar to higher vertebrates, have been isolated from zebrafish.

Alternative Splicing↗

PubMed2004

Identification and expression pattern of mago nashi during zebrafish development.

In a search for zebrafish genes expressed during early stages of development, we have identified two ESTs encoding proteins related to Drosophila mago nashi. Zebrafish mago nashi codes for a small protein with no clearly identified functional domains, and which is highly conserved during evolution. This paper describes the identification and a detailed gene expression analysis of zebrafish mago nashi during development. Our results demonstrate that mago nashi encodes a maternal transcript detected in both blastomeres and yolk cell at the 1-2 cell stages, and in the blastoderm during segmentation. We show that a putative microtubule-mediated transport of mago nashi mRNA from the vegetal hemisphere into animal blastomeres determines the localization of the transcript in the animal pole, immediately after fertilization. Furthermore, the microtubule array contained into the yolk cell seems to be responsible for the high level of mago nashi transcript detected in the central blastomeres at the 8-16 cell stages. Zygotic mago nashi is expressed into the dorsal-marginal region during gastrulation, and starting from somitogenesis to 24 hpf, the expression domain becomes progressively restricted to the developing neural tube and paraxial structures, and ventrally to the pronephric ducts.

Amino Acid Sequence↗