Biomedical subjects

S Vriz

Publications and source records attributed to S Vriz.

17 recordsLinked to original sources

PubMed2001

Zebrafish keratin 8 is expressed at high levels in the epidermis of regenerating caudal fin.

Cytokeratins are structural proteins of the intermediate filament family and are mainly expressed in epithelial cells. In several vertebrates it has been shown that keratin 8 is expressed in simple epithelial tissues, some non-epithelial tissue and in hyper-proliferative tissues during development and tumor transformation. We previously cloned and characterised the zebrafish (Danio rerio) homologous cytokeratin 8 cDNA (zfk8) which was described as an epidermal marker during zebrafish development. It has been found that the zfk8 gene is normally expressed in simple epithelia in embryonic and mature zebrafish. Using whole-mount in situ hybridisation, we show in this report that expression of zfk8 is tightly linked to the regeneration of caudal fin and exclusively observed in epidermal cells. It is strongly expressed in the epidermis overlaying the inter-rays zone of regenerating caudal fin. Our results indicate that in zebrafish, cytokeratin 8 is a suitable epidermal marker during regeneration.

Animals↗

PubMed2000

Zebrafish Hsp40 and Hsc70 genes are both induced during caudal fin regeneration.

We report the characterization of a zebrafish DnaJ-like protein, homologue to the human Hsp40 protein hdj1, that we named zf-Hsp40. We have studied its expression during fin regeneration and we show that zf-Hsp40 mRNA level is drastically increased in all ray segments beneath the regenerated part, or blastema, in response to caudal fin amputation. In order to investigate whether zf-Hsp40 is part of a stress response system after injury, we studied the expression of the zebrafish Hsc70 transcript. Our results show a correlation between the expression of the two transcripts.

Amino Acid Sequence↗

PubMed1998

Hotfoot mouse mutations affect the delta 2 glutamate receptor gene and are allelic to lurcher.

Hotfoot (ho) is a recessive mouse mutation characterized by cerebellar ataxia associated with relatively mild abnormalities of the cerebellum. It has been previously mapped to Chromosome 6, and at least eight independent alleles have been reported. Here we show that the hotfoot phenotype is associated with mutations in the glutamate receptor ionotropic delta2 gene (Grid2). We have identified a 510-bp deletion in the Grid2 coding sequence in the ho4J allele, resulting in a deletion of 170 amino acids of the extracellular domain of the receptor. Analysis of a second allele, hoTgN37INRA, revealed a 4-kb deletion in the Grid2 transcript. The GRID2 protein in these hotfoot mutants probably has a reduced (or null) activity since the phenotype of hotfoot bears similarities with the previously described phenotype of Grid2 knockout mice. The exceptionally high number of independent alleles at the ho locus is an invaluable tool for investigating the function of the glutamate receptor ionotropic delta2 protein, which so far remains largely unknown.

Alleles↗

PubMed1997

Molecular characterization of a heat shock cognate cDNA of zebrafish, hsc70, and developmental expression of the corresponding transcripts.

To elucidate the potential role of the hsp70 gene family in developmental processes in vertebrates, we chose to study the expression of one of these genes in zebrafish. A zebrafish gastrula cDNA library was screened with a Pleurodeles waltl hsp70 cDNA probe. A 2.3-kb cDNA was thus isolated and sequenced. The predicted amino acid sequence contained an open reading frame encoding for a 649-amino acid polypeptide. Sequence analysis showed strong homology with hsp70-related gene sequences in other species; in particular, the strongest homology was found with the cognate members of this family. Tests of heat inducibility revealed that transcripts were expressed at normal temperature, but the level of transcript expression increased after heat shock. Moreover, experiments of the neosynthesis of total proteins in heat shock conditions and corresponding immunoblotting assays showed that 24-h-stage embryos are able to respond to heat shock. The quantity of 70 kDa proteins, recognized by a specific antibody of the HSP/C70 protein family, is expressed in control condition and increased significantly after heat shock. Furthermore, Northern blot analysis of transcript expression showed that the corresponding mRNAs were detected throughout embryonic development in the absence of any heat shock. Our clone, named hsc70, thus corresponded to a cognate member of the hsp70 gene family, expressed under normal conditions during development, but also heat inducible. The spatio-temporal pattern of transcripts during development was determined by in situ hybridization on wholemount embryos at different stages. As a maternal RNA, hsc70 mRNA was uniformly present in the embryo, up to the end of gastrulation. Later, a tissue-specific enrichment of hsc70 transcripts was detected in the central nervous system (CNS) and in a fraction of the somites. These results suggest that the hsc70 gene may be involved in developmental differentiation events.

Amino Acid Sequence↗

PubMed1997

Construction of a high-resolution genetic map encompassing the hotfoot locus.

Hotfoot (ho) is a mutation affecting posture and movement. We report a new allele associated with the insertion of a transgene and its high-resolution mapping. Analysis of the transgene revealed that two complete and two truncated copies are inserted at the ho locus. The ho locus cosegregated with D6Mit299 in 702 meioses and is confined to a 1.1-cM region between the markers D6Mit122 and D6Mit174. If the order and distances between markers are consistent with previously published mapping data, the position of the ho locus must be revised and placed approximately 30 cM from the centromere. This high-resolution genetic map is the first step towards the positional cloning of the ho mutation.

Alleles↗

PubMed1997

Cytokeratin 8 is a suitable epidermal marker during zebrafish development.

We have cloned and characterized the zebrafish (Danio rerio) homologous cytokeratin 8 (zf-K8) cDNA. This cytokeratin belongs to the gene family of intermediate filaments and it is a component of the cytoskeleton of epithelial cells. Gene expression analysis during embryonic development and at adult stages presented here revealed that zf-K8 mRNA is inherited maternally and that it is present in the oocyte, the zygote and in the cleavage stage embryo. After mid blastula transition this gene is expressed in all surface cells, notably in those of the enveloping layer (EVL) and of the periderm, as well as in a subpopulation of the deep cells (DEL) presumed to be intestinal progenitors. During later embryonic stage zf-K8 mRNA is strongly expressed in the developing pectoral fin. In adult zebrafish, the zf-K8 gene is not only expressed in simple epithelia such as the colorectal intestine, but also, in contrast to other vertebrates, it is present in stratified skin and differentiated fins. These observations suggest that the zf-K8 gene is an appropriate epidermal marker during zebrafish ontogenesis.

Amino Acid Sequence↗

PubMed1996

Zygotic expression of the zebrafish Sox-19, an HMG box-containing gene, suggests an involvement in central nervous system development.

The zebrafish Sox-19 belongs to the Sry subfamily of HMG (Hight Mobility Group) box genes and is closely related to the Sox sub-group B, comprising the mouse Sox-1, Sox-2 and Sox-3 genes, with respect to both HMG box homology (95.3%) and neural expression during embryogenesis. Analysis of Sox-19 expression during embryogenesis by whole-mount in-situ hybridization revealed interesting features. In early gastrula embryos, Sox-19 transcripts are detected within a circular area in the region of the presomptive central nervous system (CNS) and appears to be the earliest molecular marker of the CNS in vertebrates. In the developing brain, ZfSox-19 mRNA is distributed in the ventral region of the diencephalon, midbrain and hindbrain whereas the expression is excluded from the telencephalon. In spite of the ventral localisation of its mRNA, the expression of this ZfSox-19 gene is completely normal in cyclops embryos which implies that ZfSox-19 expression is independent of the presence of the floor plate.

Amino Acid Sequence↗

PubMed1995

The zebrafish Zf-Sox 19 protein: a novel member of the Sox family which reveals highly conserved motifs outside of the DNA-binding domain.

A cDNA encoding a zebrafish Sox protein (Sry-type high-mobility-group box) was isolated and sequenced. The sequence within the HMG box is close to those of the B subfamily comprising mouse Sox-1, Sox-2, Sox-3 and Sox-14. While much of the rest of the zebrafish protein is unique, there are blocks of amino acids showing considerable identity with regions of Sox-1, -2 and -3. These domains may represent conserved parts of the protein, required for interaction with other proteins, and strengthen the assignment of the zebrafish gene, termed Zf-sox 19, to the B subfamily.

Amino Acid Sequence↗

PubMed1995

Widespread expression of the eve1 gene in zebrafish embryos affects the anterior-posterior axis pattern.

The zygotic expression of the eve1 gene is restricted to the ventral and lateral cells of the marginal zone. At later stages, the mRNAs are localized in the most posterior part of the extending tail tip. An eve1 clone (pcZf14), containing a poly-A tail, has been isolated. In order to address eve1 gene function, pcZf14 transcript injections into zebrafish embryos have been performed. The injection into uncleaved eggs of a synthetic eve1 mRNA (12 pg), which encodes a protein of approximately 28 kd, produces embryos with anterior-posterior (A-P) axis defects and the formation of additional axial structures. The first category of 24 h phenotypes (87%) mainly displays a gradual decrease in anterior structures. This is comparable to previous phenotypes observed following Xhox3 messenger injection either in Xenopus or in zebrafish that have been classified according to the index of axis deficiency (zf-IAD). These phenotypes result in anomalies of the development of the neural keel, from microphthalmia to acephaly. The second category (13%) corresponds to the phenotypes described above together with truncal or caudal supernumerary structures. Additional truncal structures are the most prominent of these duplicated phenotypes, displaying a "zipper" shape of axial structures including neural keels and notochords. Caudal duplication presents no evident axis supernumerary structures. The observation of these phenotypes suggests an important role for the eve1 gene in mesodermal cell specification and in the development of the posterior region, and more particularly of the most posterior tail tip where endogenous eve1 messengers are found.

Amino Acid Sequence↗

PubMed1995

The SRY protein, like HMG 1, recognizes (CA)n sequences, an abundant repeat sequence in vertebrates.

The sex-determining region of the Y chromosome gene, sry is expressed in the foetal mouse for a brief period, just before testis differentiation, which could be consistent with negative autoregulation. SRY is a DNA binding protein which can bind to cruciform DNA and to linear DNA with a sequence specificity. We have examined if the Sry gene contain DNA binding sites for the SRY protein itself. We have found that in an in vitro assay, the SRY protein binds to several sites of the Sry gene and especially to a (CA)25 sequence and to a (CAG)30 repeat. These binding suggest that the function of SRY and in a general way HMG-box proteins may be mediated by an interaction with repeat sequences.

Animals↗

PubMed1992

Comparative analysis of the intracellular localization of c-Myc, c-Fos, and replicative proteins during cell cycle progression.

In eukaryotic cells, nucleus-cytoplasm exchanges play an important role in genomic regulation. We have analyzed the localization of four nuclear antigens in different growth conditions: two replicative proteins, DNA polymerase alpha and proliferating cell nuclear antigen (PCNA), and two oncogenic regulatory proteins, c-Myc and c-Fos. A kinetic study of subcellular localization of these proteins has been done. In cultures in which cells were sparse, these proteins were detected in the nucleus. When proliferation was stopped by the high density of culture cells or by serum starvation, these proteins left the nucleus for the cytoplasm with different kinetics. DNA polymerase alpha is the first protein to leave the nucleus, with the PCNA protein, c-Fos, and c-Myc leaving the nucleus later. In contrast, during serum stimulation c-Fos and c-Myc relocalize into the nucleus before the replicative proteins. We also noticed that in sparse cell cultures, 10% of the cells exhibit a perinuclear staining for the DNA polymerase alpha, PCNA, and c-Myc proteins but not for c-Fos. This peculiar staining was also observed as an initial step to nuclear localization after serum stimulation and in vivo in Xenopus embryos when the G1 phase is reintroduced in the embryonic cell cycle at the mid-blastula stage. We suggest that such staining could reflect specific structures involved in the initiation of the S phase.

3T3 Cells↗

PubMed1991

Posttranscriptional regulation of c-myc RNA during early development of Xenopus laevis.

The remarkable stability of c-myc during oogenesis contrasts with its degradation during the early developmental period in Xenopus laevis. Three evolutionary conserved motifs found in the 3'-untranslated region of Xenopus c-myc RNAs have been analyzed for a possible role in c-myc RNA degradation. No specific degradation was observed when these sequences were cloned downstream of a reporter gene and the corresponding RNAs were injected into fertilized eggs. The relation between polyadenylation and degradation of c-myc mRNA has been examined during early development. c-myc is adenylated during early oogenesis, and a dramatic de-adenylation occurs in full grown oocytes. Consequently, the de-adenylation of c-myc mRNA that occurs in eggs might be a requirement for its degradation after fertilization, but is not sufficient to trigger its degradation.

Animals↗

PubMed1990

Enhanced c-myc gene expression during forelimb regenerative outgrowth in the young Xenopus laevis.

Analysis of the expression of the c-myc protooncogene has been carried out in the forelimb regenerate of the Xenopus laevis froglet. Northern blot hybridization analysis revealed the presence of a 2.5-kilobase c-myc transcript in the regenerate forelimb at a level at least 7-fold more than the one found in nonregenerating forelimbs or stumps of regenerating forelimbs. In situ hybridization analyses confirmed the relative abundance of c-myc RNA in the regenerate forelimb and provided evidence of spatial localization of high levels of c-myc RNA in specific cell layers. The deepest layers of the wound epithelium of epidermal origin showed a strong signal, whereas virtually no c-myc RNA was detected in the outermost layers. Labeling was also observed in mesenchymal cells of the blastema where it was relatively evenly distributed. This pattern of c-myc RNA in the regenerate might indicate that the expression of c-myc plays a role in the regulation of the continued proliferation of specific cells of the regenerate, whereas repression of this gene in the epidermis correlates with terminal differentiation of keratinocytes.

Amputation, Surgical↗

PubMed1990

Genes and mechanisms involved in early embryonic development in Xenopus laevis.

Our laboratory is studying genes involved in the regulation of the balance between cell growth and differentiation during embryonic development in Xenopus. We have analyzed the developmental expression of the proto-oncogenes c-myc, and KiRas 2B, the proliferating cell nuclear antigen (PCNA), and the tumor suppressor gene p53. These genes, usually expressed during cell proliferation, are expressed in the oocyte in large quantities, but the majority of their maternal RNAs are degraded by the gastrula stage. The expression of c-myc and the localization of the protein indicate that c-myc has the characteristics expected for a gene involved in the regulation of the mid-blastula transition, when zygotic expression is turned on in the embryo. Its expression during late development or during regeneration indicates that it enables the cells to remain competent for cycling during organogenesis. In vitro systems that reproduce the principal cellular functions during early development are used as model systems to understand the mechanisms involved in early embryogenesis.

Animals↗

PubMed1989

Differential expression of two Xenopus c-myc proto-oncogenes during development.

Two distinct Xenopus c-myc cDNA clones have been characterized from an oocyte cDNA library. This allowed a comparison of the c-myc protein sequence across the vertebrate phylum to be made and prominent conservations to be identified. The majority of the sequence differences between the two Xenopus c-myc cDNAs are in the 5' and 3' untranslated regions. Sequence-specific oligonucleotide probes from the 5' untranslated region were used to demonstrate the differential expression of the two c-myc mRNAs during development. One of the mRNAs corresponds to the Xenopus c-myc gene previously reported expressed as a stable maternal mRNA uncoupled from cell division during oogenesis (c-myc I). It is the major mRNA species expressed during oogenesis and is expressed again from the zygotic genome in post-gastrula embryos. In contrast, the second c-myc mRNA (c-myc II) is expressed only from the maternal genome during oogenesis. Primer extension experiments show that in the oocyte the transcriptional initiation sites for c-myc I and c-myc II are at different distances from the translational start site. The 'oocyte-specific' and 'somatic-type' developmental regulation of c-myc is reminiscent of polymerase III 5S RNA gene expression in Xenopus, and may provide new insights into the developmental regulation of genes transcribed by RNA polymerase II.

Amino Acid Sequence↗

PubMed1989

Analysis of 3'-untranslated regions of seven c-myc genes reveals conserved elements prevalent in post-transcriptionally regulated genes.

We have characterized the complete sequence of two c-myc cDNAs from the amphibian Xenopus laevis, and could thus compare the 3'-non-coding sequences of 7 myc cDNAs from 6 species spread over 350 million years of evolution. Although the size of these sequences is heterogeneous, we identified three completely conserved sequences of 10, 11 and 12 contiguous nucleotides. We observed that two of these elements may be contained in conserved stem-loop structures previously implicated in mRNA turnover. The length of these motifs, their existence in conserved predicted structures, and their presence in regulated eukaryote mRNA with a frequency greater than predicted by chance, suggest that they are functionally important.

Animals↗

PubMed1988

Proto-oncogenes and embryonic development.

The role of proto-oncogenes in embryonic development was investigated using one of the most characterized vertebrates, the amphibian Xenopus laevis. Genes which belong to the major proto-oncogene families have been detected in Xenopus genome. The developmental control of the myc gene was assayed using a characterized Xenopus myc probe and specific antibodies. The myc gene is highly expressed as a stable maternal mRNA in oocyte, and an unfertilized egg contains 5 X 10(5)-fold the myc RNA content of a proliferative somatic cell. The myc RNA store is evenly distributed in the oocyte and the egg. Fertilization triggers a post-transcriptional control of the gene and the RNA store is progressively degraded to a constitutive value of 10 to 30 myc RNA copies registered per gastrula embryonic cell. The 62K myc protein is accumulated late in oogenesis. This uncoupling of myc expression and cell proliferation appears as a specific developmental regulation of the myc gene, adapted to the series of rapid cell cleavages occurring after fertilization.

Animals↗