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M Ekker

Publications and source records attributed to M Ekker.

At least 19 recordsLinked to original sources
PubMed2001

Evidence that GRIP, a PDZ-domain protein which is expressed in the embryonic forebrain, co-activates transcription with DLX homeodomain proteins.

The DLX homeodomain proteins control development of the basal ganglia and branchial arches. To identify co-factors that regulate DLX function we utilized the yeast two-hybrid assay, and found a DLX interacting protein (DIP2) which binds to the N-terminal region of DLX2 via a PDZ domain. DIP2 appears to be an alternatively spliced form of GRIP1, a protein known to bind AMPA glutamate receptors via PDZ domains. Thus, we named it GRIP1b. We provide evidence that GRIP1b can function as a transcriptional co-activator of DLX2 and DLX5. Glutamate receptors inhibit this co-activation. These results suggest that some PDZ proteins may regulate transcription via their interactions with homeodomain proteins. Furthermore, these results suggest a link between glutamate receptors, PDZ proteins and the DLX transcription factors, all of which are co-expressed in the developing basal ganglia.

Alternative Splicing↗
PubMed2001

Expression and regulation of mouse Mtsh1 during limb and branchial arch development.

The mouse genome contains at least two genes, Mtsh1 and Mtsh2, related in sequence to the Drosophila homeotic gene teashirt (tsh). In this paper, we report the characterization of Mtsh1 expression in the developing branchial arches and forelimbs during mouse embryogenesis. Mtsh1 was found predominantly transcribed in the mesenchymal tissue of branchial arches and forelimbs. Surgical removal of the epithelium of both forelimb and branchial arch significantly decreased the expression of Mtsh1 in the mesenchymal cells of these tissues. Upon implantation of FGF8-soaked beads into arches and limbs, Mtsh1 transcription was up-regulated. In contrast, when BMP4-soaked beads were implanted, Mtsh1 expression was inhibited. Together, these results suggest that mouse Mtsh1 gene may be involved in the outgrowth of limbs and arches and may be functioning downstream of BMP and FGF signaling pathways.

Animals↗
PubMed2001

fgfr3 and regionalization of anterior neural tube in zebrafish.

Here we describe the isolation of the zebrafish fgfr3 gene, its structure and chromosomal location. Expression in wild type embryos occurs in the axial mesoderm, the diencephalon, the anterior hindbrain and the anterior spinal cord. In the hindbrain, a differential expression of fgfr3 was detected at several levels of intensity, with the highest expression in the posterior rhombomere 1 that is morphologically distinct from the anterior part, which develops into the cerebellum. Further, analysis of fgfr3 expression in mutants deficient in the formation of the midbrain-hindbrain boundary (MHB), noi(-/-) and ace(-/-), demonstrated that in the absence of Pax2.1 and FGF8 activity, the expression domains of FGFR3 expand into the MHB, tegmentum, cerebellum and optic tectum, which are the affected structures in these mutants.

Amino Acid Sequence↗
PubMed2001

The LN54 radiation hybrid map of zebrafish expressed sequences.

To increase the density of a gene map of the zebrafish, Danio rerio, we have placed 3119 expressed sequence tags (ESTs) and cDNA sequences on the LN54 radiation hybrid (RH) panel. The ESTs and genes mapped here join 748 SSLp markers and 459 previously mapped genes and ESTs, bringing the total number of markers on the LN54 RH panel to 4226. Addition of these new markers brings the total LN54 map size to 14,372 cR, with 118 kb/cR. The distribution of ESTs according to linkage groups shows relatively little variation (minimum, 73; maximum, 201). This observation, combined with a relatively uniform size for zebrafish chromosomes, as previously indicated by karyotyping, indicates that there are no especially gene-rich or gene-poor chromosomes in this species. We developed an algorithm to provide a semiautomatic method for the selection of additional framework markers for the LN54 map. This algorithm increased the total number of framework markers to 1150 and permitted the mapping of a high percentage of sequences that could not be placed on a previous version of the LN54 map. The increased concentration of expressed sequences on the LN54 map of the zebrafish genome will facilitate the molecular characterization of mutations in this species.

Animals↗
PubMed2000

Differential expression of orthologous Dlx genes in zebrafish and mice: implications for the evolution of the Dlx homeobox gene family.

Dlx homeobox genes of vertebrates are often organised as physically linked pairs in which the two genes are transcribed convergently (tail-to-tail arrangement). Three such Dlx pairs have been found in mouse, human, and zebrafish and are thought to have originated from the duplication of an ancestral gene pair. These pairs include Dlx1/Dlx2, Dlx7/Dlx3, and Dlx6/Dlx5 (the zebrafish orthologue of Dlx5 is named dlx4). Expression patterns of physically linked Dlx genes overlap extensively. Furthermore, orthologous Dlx genes often show highly similar expression patterns. We analysed Dlx expression during the gastrula and early somitogenesis of the mouse and zebrafish. It was found that expression of the mouse Dlx6 gene takes place in the rostral ectoderm and presumptive olfactory and otic placodes with patterns similar to the previously reported expression of the physically linked Dlx5 gene. However, we observed only very weak expression of the mouse Dlx3 gene at the same stage. This contrasts with the expression of dlx genes in zebrafish where dlx3 and dlx7, but not dlx4 and dlx6 are expressed during gastrulation in the rostral ectoderm and presumptive placodes. Thus, Dlx expression patterns at early stages are better conserved between paralogous pairs of physically linked genes than between orthologous pairs. This suggests that early expression of Dlx genes existed prior to the duplications that led to the multiple pairs of physically linked genes but was differentially conserved in different paralogs in zebrafish and mice.

Animals↗
PubMed2000

Characterization of a zebrafish/mouse somatic cell hybrid panel.

We have characterized a collection of zebrafish/mouse somatic cell hybrids with 211 genes and markers chosen from the 25 zebrafish linkage groups. Most of the zebrafish genome is represented in this collection with 88% of genes/markers present in at least one hybrid cell line. Although most hybrids contain chromosomal fragments, there are a few instances where a complete or nearly complete zebrafish chromosome has been maintained in a mouse background, based on multiple markers covering the entire chromosome. In addition to their use in mapping studies, this collection of somatic cell hybrids should constitute an important tool as a source of specific chromosome fragments and for assessing the function of genome regions.

Animals↗
PubMed2000

A highly conserved enhancer in the Dlx5/Dlx6 intergenic region is the site of cross-regulatory interactions between Dlx genes in the embryonic forebrain.

Four Dlx homeobox genes, Dlx1, Dlx2, Dlx5, and Dlx6 are expressed in the same primordia of the mouse forebrain with temporally overlapping patterns. The four genes are organized as two tail-to-tail pairs, Dlx1/Dlx2 and Dlx5/Dlx6, a genomic arrangement conserved in distantly related vertebrates like zebrafish. The Dlx5/Dlx6 intergenic region contains two sequences of a few hundred base pairs, remarkably well conserved between mouse and zebrafish. Reporter transgenes containing these two sequences are expressed in the forebrain of transgenic mice and zebrafish with patterns highly similar to endogenous Dlx5 and Dlx6 expression. The activity of the transgene is drastically reduced in mouse mutants lacking both Dlx1 and Dlx2, consistent with the decrease in endogenous Dlx5 and Dlx6 expression. These results suggest that cross-regulation by Dlx proteins, mediated by the intergenic sequences, is essential for Dlx5 and Dlx6 expression in the forebrain. This hypothesis is supported by cotransfection and DNA-protein binding experiments. We propose that the Dlx genes are part of a highly conserved developmental pathway that regulates forebrain development.

Animals↗
PubMed2000

The zebrafish scyba gene encodes a novel CXC-type chemokine with distinctive expression patterns in the vestibulo-acoustic system during embryogenesis.

Chemokines, in addition to their characterized functions as immune modulators, also play a role in developmental processes such as neural cell migration. Although, chemokines have been described in human, mouse and other vertebrate species, they have yet to be characterized in zebrafish. In this paper, we report the isolation and expression analysis of scyba, a zebrafish gene encoding a CXC-type chemokine protein. During early segmentation, scyba transcripts are detected in the midbrain region and the otic placodes. At later developmental stages, scyba expression is restricted to a subset of hindbrain commissural neurons and to the hair-cell sensory patches of the otic vesicle and lateral-line neuromasts.

Amino Acid Sequence↗
PubMed2000

Distal-less-related homeobox genes of vertebrates: evolution, function, and regulation.

Homeobox genes of the Distal-less family have been identified in virtually all metazoan groups where they play roles in the ontogeny of these animals. The vertebrate Distal-less related genes (Dlx genes) are thought to have arisen as a result of a tandem gene duplication event followed by a number of larger genomic scale duplications and thus represent an interesting model with which to study the evolution of clustered gene families. Dlx genes are involved in the development of the forebrain, branchial arches, sensory organs, and limbs. Here we describe the current state of knowledge of the Dlx genes in terms of their developmental expression, how this expression is regulated and how the products of these genes function, once expressed. We highlight a number of recent studies that have shed light on the transcriptional regulation of this gene family. These findings have not only contributed to our understanding of the selective pressures involved in the maintenance of familial gene clustering in genomes, but also to our understanding of how genes may diverge in function during the course of evolution as a result of divergence of regulatory mechanisms.

Animals↗
PubMed1999

Radiation hybrid mapping of the zebrafish genome.

The zebrafish is an excellent genetic system for the study of vertebrate development and disease. In an effort to provide a rapid and robust tool for zebrafish gene mapping, a panel of radiation hybrids (RH) was produced by fusion of irradiated zebrafish AB9 cells with mouse B78 cells. The overall retention of zebrafish sequences in the 93 RH cell lines that constitute the LN54 panel is 22%. Characterization of the LN54 panel with 849 simple sequence length polymorphism markers, 84 cloned genes and 122 expressed sequence tags allowed the production of an RH map whose total size was 11,501 centiRays. From this value, we estimated the average breakpoint frequency of the LN54 RH panel to correspond to 1 centiRay = 148 kilobase. Placement of a group of 235 unbiased markers on the RH map suggests that the map generated for the LN54 panel, at present, covers 88% of the zebrafish genome. Comparison of marker positions in RH and meiotic maps indicated a 96% concordance. Mapping expressed sequence tags and cloned genes by using the LN54 panel should prove to be a valuable method for the identification of candidate genes for specific mutations in zebrafish.

Animals↗
PubMed1999

A role for DNA methylation in gastrulation and somite patterning.

DNA methylation constitutes an important epigenetic factor in the control of genetic information. In this study, we analyzed expression of the DNA methyltransferase gene and examined DNA methylation patterns during early development of the zebrafish. Maternal transcripts of the zebrafish DNA methyltransferase gene (MTase) are ubiquitously present at high levels in early embryos with overall levels decreasing after the blastula stage. At 24 h, methyltransferase mRNA is predominantly found in the brain, neural tube, eyes, and differentiating somites. Expression of MTase in the somites is highest in the anterior cells of the somites. Despite the high levels of MTase mRNA in blastula-stage embryos, we observe DNA hypomethylation at the blastula and gastrula stages compared to sperm or older embryos. Zebrafish embryos treated with 5-azacytidine (5-azaC) and 5-aza-2-deoxycytidine (5-azadC), nucleotide analogs known to induce cellular differentiation and DNA hypomethylation in mammalian cells, exhibit DNA hypomethylation and developmental perturbations. These defects are specifically observed in embryos treated at the beginning of the blastula period, just prior to midblastula transition. The most common phenotype is the loss of tail and abnormal patterning of somites. Head development is also affected in some embryos. Histological and in situ hybridization analyses reveal whole or partial loss of a differentiated notochord and midline muscle in treated embryos. When examined during gastrulation, 5-azaC-treated embryos have a shortened and thickened axial mesoderm. We propose that DNA methylation is required for normal gastrulation and subsequent patterning of the dorsal mesoderm.

Abnormalities, Drug-Induced↗
PubMed1999

Saving zebrafish mutants.

At present, the zebrafish Danio rerio is the only vertebrate species for which a large-scale mutagenesis effort to identify developmental genes has been reported. Several laboratories are now intensely pursuing the molecular characterization of the genes affected by these mutations. One important criterion for the identity of the mutated gene is the rescue of the mutant phenotype by a wild-type (wt) copy of the gene. Until recently, most rescue attempts were carried out by injecting wt messenger RNA (mRNA) into fertilized eggs. A report by Yan and collaborators shows the partial rescue of floatinghead mutants by injection of genomic fragments cloned in either bacterial artificial chromosomes or bacteriophage lambda vectors. Combined with other ongoing efforts to characterize the zebrafish genome, this approach of mutant rescue opens interesting avenues for a systematic functional analysis of vertebrate genes.

Animals↗
PubMed1999

The role of tolloid/mini fin in dorsoventral pattern formation of the zebrafish embryo.

A highly conserved TGF-&bgr; signaling pathway is involved in the establishment of the dorsoventral axis of the vertebrate embryo. Specifically, Bone Morphogenetic Proteins (Bmps) pattern ventral tissues of the embryo while inhibitors of Bmps, such as Chordin, Noggin and Follistatin, are implicated in dorsal mesodermal and neural development. We investigated the role of Tolloid, a metalloprotease that can cleave Chordin and increase Bmp activity, in patterning the dorsoventral axis of the zebrafish embryo. Injection of tolloid mRNA into six dorsalized mutants rescued only one of these mutants, mini fin. Through chromosomal mapping, linkage and cDNA sequence analysis of several mini fin alleles, we demonstrate that mini fin encodes the tolloid gene. Characterization of the mini fin mutant phenotype reveals that Mini fin/Tolloid activity is required for patterning ventral tissues of the tail: the ventral fin, and the ventroposterior somites and vasculature. Gene expression studies show that mfn mutants exhibit reduced expression of ventrally restricted markers at the end of gastrulation, suggesting that the loss of ventral tail tissues is caused by a dorsalization occurring at the end of gastrulation. Based on the mini fin mutant phenotype and the expression of tolloid, we propose that Mini fin/Tolloid modifes the Bmp activity gradient at the end of gastrulation, when the ventralmost marginal cells of the embryo are in close proximity to the dorsal Chordin-expressing cells. At this time, unimpeded Chordin may diffuse to the most ventral marginal regions and inhibit high Bmp activity levels. In the presence of Mini fin/Tolloid, however, Chordin activity would be negatively modulated through proteolytic cleavage, thereby increasing Bmp signaling activity. This extracellular mechanism is amplified by an autoregulatory loop for bmp gene expression.

Animals↗
PubMed1998

Zebrafish hox clusters and vertebrate genome evolution.

HOX genes specify cell fate in the anterior-posterior axis of animal embryos. Invertebrate chordates have one HOX cluster, but mammals have four, suggesting that cluster duplication facilitated the evolution of vertebrate body plans. This report shows that zebrafish have seven hox clusters. Phylogenetic analysis and genetic mapping suggest a chromosome doubling event, probably by whole genome duplication, after the divergence of ray-finned and lobe-finned fishes but before the teleost radiation. Thus, teleosts, the most species-rich group of vertebrates, appear to have more copies of these developmental regulatory genes than do mammals, despite less complexity in the anterior-posterior axis.

Animals↗
PubMed1998

Ventral and lateral regions of the zebrafish gastrula, including the neural crest progenitors, are established by a bmp2b/swirl pathway of genes.

A bone morphogenetic protein (BMP) signaling pathway is implicated in dorsoventral patterning in Xenopus. Here we show that three genes in the zebrafish, swirl, snailhouse, and somitabun, function as critical components within a BMP pathway to pattern ventral regions of the embryo. The dorsalized mutant phenotypes of these genes can be rescued by overexpression of bmp4, bmp2b, an activated BMP type I receptor, and the downstream functioning Smad1 gene. Consistent with a function as a BMP ligand, swirl functions cell nonautonomously to specify ventral cell fates. Chromosomal mapping of swirl and cDNA sequence analysis demonstrate that swirl is a mutation in the zebrafish bmp2b gene. Interestingly, our analysis suggests that the previously described nonneural/neural ectodermal interaction specifying the neural crest occurs through a patterning function of swirl/bmp2b during gastrulation. We observe a loss in neural crest progenitors in swirl/bmp2b mutant embryos, while somitabun mutants display an opposite, dramatic expansion of the prospective neural crest. Examination of dorsally and ventrally restricted markers during gastrulation reveals a successive reduction and reciprocal expansion in nonneural and neural ectoderm, respectively, in snailhouse, somitabun, and swirl mutant embryos, with swirl/bmp2b mutants exhibiting almost no nonneural ectoderm. Based on the alterations in tissue-specific gene expression, we propose a model whereby swirl/bmp2b acts as a morphogen to specify different cell types along the dorsoventral axis.

Animals↗
PubMed1998

A microsatellite genetic linkage map for zebrafish (Danio rerio).

We have constructed a zebrafish genetic linkage map consisting of 705 simple sequence-length polymorphism markers (SSLPs). The map covers 2350 centimorgans (cM) of the zebrafish genome with an average resolution of 3.3 cM. It is a complete map in genetic mapping terms (there is one linkage group for each of the 25 chromosomes), and it has been confirmed by somatic-cell hybrids and centromere-mapping using half-tetrad analysis. The markers are highly polymorphic in the zebrafish strains used for genetic crosses and provide a means to compare genetic segregation of developmental mutations between laboratories. These markers will provide an initial infrastructure for the positional cloning of the nearly 600 zebrafish genes identified as crucial to vertebrate development,and will become the anchor for the physical map of the zebrafish genome.

Animals↗
PubMed1998

Involvement of the sonic hedgehog, patched 1 and bmp2 genes in patterning of the zebrafish dermal fin rays.

The signaling molecule encoded by Sonic hedgehog (shh) participates in the patterning of several embryonic structures including limbs. During early fin development in zebrafish, a subset of cells in the posterior margin of pectoral fin buds express shh. We have shown that regulation of shh in pectoral fin buds is consistent with a role in mediating the activity of a structure analogous to the zone of polarizing activity (ZPA) (Akimenko and Ekker (1995) Dev. Biol. 170, 243-247). During growth of the bony rays of both paired and unpaired fins, and during fin regeneration, there does not seem to be a region equivalent to the ZPA and one would predict that shh would play a different role, if any, during these processes specific to fish fins. We have examined the expression of shh in the developing fins of 4-week old larvae and in regenerating fins of adults. A subset of cells in the basal layer of the epidermis in close proximity to the newly formed dermal bone structures of the fin rays, the lepidotrichia, express shh, and ptc1 which is thought to encode the receptor of the SHH signal. The expression domain of ptc1 is broader than that of shh and adjacent blastemal cells releasing the dermal bone matrix also express ptc1. Further observations indicate that the bmp2 gene, in addition to being expressed in the same cells of the basal layer of the epidermis as shh, is also expressed in a subset of the ptc1-expressing cells of the blastema. Amputations of caudal fins immediately after the first branching point of the lepidotrichia, and global administration of all-trans-retinoic acid, two procedures known to cause fusion of adjacent rays, result in a transient decrease in the expression of shh, ptc1 and bmp2. The effects of retinoic acid on shh expression occur within minutes after the onset of treatment suggesting direct regulation of shh by retinoic acid. These observations suggest a role for shh, ptc1 and bmp2 in patterning of the dermoskeleton of developing and regenerating teleost fins.

Animals↗