PubMed Health⌕ Search

Biomedical subjects

W Driever

Publications and source records attributed to W Driever.

At least 55 records · Page 3Linked to original sources
PubMed1996

A reference cross DNA panel for zebrafish (Danio rerio) anchored with simple sequence length polymorphisms.

The ultimate informativeness of the zebrafish mutations described in this issue will rest in part on the ability to clone these genes. However, the genetic infrastructure required for the positional cloning in zebrafish is still in its infancy. Here we report a reference cross panel of DNA, consisting of 520 F2 progeny (1040 meioses) that has been anchored to a zebrafish genetic linkage map by 102 simple sequence length polymorphisms. This reference cross DNA provides: (1) a panel of DNA from the cross that was used to construct the genetic linkage map, upon which polymorphic gene(s) and genetic markers can be mapped; (2) a fine order mapping tool, with a maximum resolution of 0.1 cM; and (3) a foundation for the development of a physical map (an ordered array of clones each containing a known portion of the genome). This reference cross DNA will serve as a resource enabling investigators to relate genes or genetic markers directly to a single genetic linkage map and avoid the problem of integrating different maps with different genetic markers, as must be currently done when using randomly amplified polymorphic DNA markers, or as has occurred with human genetic linkage maps.

Alleles↗
PubMed1996

Mutations affecting cell fates and cellular rearrangements during gastrulation in zebrafish.

One of the major challenges of developmental biology is understanding the inductive and morphogenetic processes that shape the vertebrate embryo. In a large-scale genetic screen for zygotic effect, embryonic lethal mutations in zebrafish we have identified 25 mutations that affect specification of cell fates and/or cellular rearrangements during gastrulation. These mutations define at least 14 complementation groups, four of which correspond to previously identified genes. Phenotypic analysis of the ten novel loci revealed three groups of mutations causing distinct effects on cell fates in the gastrula. One group comprises mutations that lead to deficiencies in dorsal mesodermal fates and affect central nervous system patterning. Mutations from the second group affect formation of ventroposterior embryonic structures. We suggest that mutations in these two groups identify genes necessary for the formation, maintenance or function of the dorsal organizer and the ventral signaling pathway, respectively. Mutations in the third group affect primarily cellular rearrangements during gastrulation and have complex effects on cell fates in the embryo. This group, and to some extent mutations from the first two groups, affect the major morphogenetic processes, epiboly, convergence and extension, and tail morphogenesis. These mutations provide an approach to understanding the genetic control of gastrulation in vertebrates.

Animals↗
PubMed1995

A behavioral screen for isolating zebrafish mutants with visual system defects.

Optokinetic and phototactic behaviors of zebrafish larvae were examined for their usefulness in screening for recessive defects in the visual system. The optokinetic response can be reliably and rapidly detected in 5-day larvae, whereas the phototactic response of larvae is variable and not robust enough to be useful for screening. We therefore measured optokinetic responses of mutagenized larvae as a genetic screen for visual system defects. Third-generation larvae, representing 266 mutagenized genomes, were examined for abnormal optokinetic responses. Eighteen optokinetic-defective mutants were identified and two mutants that did not show obvious morphological defects, no optokinetic response a (noa) and partial optokinetic response a (poa), were studied further. We recorded the electroretinogram (ERG) to determine whether these two mutations affect the retina. The b-wave of noa larvae was grossly abnormal, being delayed in onset and significantly reduced in amplitude. In contrast, the ERG waveform of poa larvae was normal, although the b-wave was reduced in amplitude in bright light. Histologically, the retinas of noa and poa larvae appeared normal. We conclude that noa larvae have a functional defect in the outer retina, whereas the outer retina of poa larvae is likely to be normal.

Animals↗
PubMed1995

Transparent things: cell fates and cell movements during early embryogenesis of zebrafish.

Development of an animal embryo involves the coordination of cell divisions, a variety of inductive interactions and extensive cellular rearrangements. One of the biggest challenges in developmental biology is to explain the relationships between these processes and the mechanisms that regulate them. Teleost embryos provide an ideal subject for the study of these issues. Their optical lucidity combined with modern techniques for the marking and observation of individual living cells allow high resolution investigations of specific morphogenetic movements and the construction of detailed fate maps. In this review we describe the patterns of cell divisions, cellular movements and other morphogenetic events during zebrafish early development and discuss how these events relate to the formation of restricted lineages.

Animals↗
PubMed1995

Axis formation in zebrafish.

Recent advances in our understanding of axis formation and patterning in zebrafish relate the developmental mode of this aspiring genetic model organism to higher vertebrates. The effect of UV irradiation and lithium treatment, as well as detailed early lineage analyses, have shed some light on dorsoventral axis formation. However, the molecular mechanism of axis formation, as well as the identity of a fish Nieuwkoop center, are still open issues. A Vg1 homolog is expressed in zebrafish, and activin as well as the mouse nodal gene product have been demonstrated to induce mesoderm and ectopic axes, respectively, in zebrafish. The zebrafish organizer is defined by the expression domains of goosecoid, axial, and lim1. The cyclops gene is involved in maintaining goosecoid expression in axial mesoderm of the head. Large mutagenesis screens provide the basis for a genetic analysis of axis formation.

Animals↗
PubMed1995

Gridlock, a localized heritable vascular patterning defect in the zebrafish.

We are using the zebrafish, Danio rerio, to identify genes that generate and pattern the vertebrate vasculature. We have isolated a recessive mutation, gridlockm145 (grlm145) in which blood flow to the tail is impeded by a localized vascular defect. Using a novel microangiographic method, we show that the blockade is in the anterior trunk, where the paired lateral dorsal aortae normally merge to form the single midline aorta. Arterial-venous shunts and collateral vessels develop in most mutant embryos, bypassing the lesion and reconstituting caudal blood flow. The grl defect resembles coarctation of the aorta, a human congenital cardiovascular malformation of unknown aetiology, in the location of the lesion and its consequences and in the mutants' dependence on collateral vessels for survival.

Angiography↗
PubMed1994

Implications for dorsoventral axis determination from the zebrafish mutation janus.

THE mechanisms underlying the formation of dorsoventral polarity in the zebrafish Danio rerio are unknown. Here we describe the zebrafish recessive maternal-effect mutation janusm55. The mutant phenotype is a division of the blastoderm along the first cleavage plane into two detached half-sized blastoderms. Partial-axis bifurcation occurs in a subset of mutants. Analysis of goosecoid expression in the mutant embryos indicates that only one organizer region is present in each embryo. Furthermore, the position of this organizer region is random with respect to the first cleavage plane bisecting the two blastoderms. Finally, cell tracing in wild-type embryos demonstrates that there is no strict correlation of the dorsoventral axis with early cleavage planes in zebrafish. These findings support the notion that the establishment of the dorsoventral axis and the first cleavage planes are determined by separate mechanisms in the zebrafish embryo.

Animals↗
PubMed1994

Zebrafish: genetic tools for studying vertebrate development.

Zebrafish have entered the arena of vertebrate biology as a mainstream model system, and the use of genetic tools in this tropical fish should enhance our understanding of vertebrate development. The zebrafish system allows genetic experiments that are not possible in other vertebrates, and the mutations isolated thus far attest to its usefulness, complementing knowledge obtained from other model organisms.

Animals↗
PubMed1994

Efficient recovery of ENU-induced mutations from the zebrafish germline.

We studied the efficiency with which two chemical mutagens, ethyl methanesulfonate (EMS) and N-ethyl-N-nitrosourea (ENU) can induce mutations at different stages of spermatogenesis in zebrafish (Brachydanio rerio). Both EMS and ENU induced mutations at high rates in post-meiotic germ cells, as indicated by the incidence of F1 progeny mosaic for the albino mutation. For pre-meiotic germ cells, however, only ENU was found to be an effective mutagen, as indicated by the frequencies of non-mosaic mutant progeny at four different pigmentation loci. Several mutagenic regimens that varied in either the number of treatments or the concentration of ENU were studied to achieve an optimal ratio between the mutagenicity and toxicity. For the two most mutagenic regimens: 4 x 1 hr in 3 mM ENU and 6 x 1 hr in 3 mM ENU, the minimum estimate of frequencies of independent mutations per locus per gamete was 0.9-1.3 X 10(-3). We demonstrate that embryonic lethal mutations induced with ENU were transmitted to offspring and that they could be recovered in an F2 screen. An average frequency of specific-locus mutations of 1.1 X 10(-3) corresponded to approximately 1.7 embryonic lethal mutations per single mutagenized genome. The high rates of mutations achievable with ENU allow for rapid identification of large numbers of genes involved in a variety of aspects of zebrafish development.

Alleles↗
PubMed1994

Microtubule arrays of the zebrafish yolk cell: organization and function during epiboly.

In zebrafish (Danio rerio), meroblastic cleavages generate an embryo in which blastomeres cover the animal pole of a large yolk cell. At the 500-1000 cell stage, the marginal blastomeres fuse with the yolk cell forming the yolk syncytial layer. During epiboly the blastoderm and the yolk syncytial layer spread toward the vegetal pole. We have studied developmental changes in organization and function during epiboly of two distinct microtubule arrays located in the cortical cytoplasm of the yolk cell. In the anuclear yolk cytoplasmic layer, an array of microtubules extends along the animal-vegetal axis to the vegetal pole. In the early blastula the yolk cytoplasmic layer microtubules appear to originate from the marginal blastomeres. Once formed, the yolk syncytial layer exhibits its own network of intercrossing mitotic or interphase microtubules. The microtubules of the yolk cytoplasmic layer emanate from the microtubule network of the syncytial layer. At the onset of epiboly, the external yolk syncytial layer narrows, the syncytial nuclei become tightly packed and the network of intercrossing microtubules surrounding them becomes denser. Soon after, there is a vegetal expansion of the blastoderm and of the yolk syncytial layer with its network of intercrossing microtubules. Concomitantly, the yolk cytoplasmic layer diminishes and its set of animal-vegetal microtubules becomes shorter. We investigated the involvement of microtubules in epiboly using the microtubule depolymerizing agent nocodazole and a stabilizing agent taxol. In embryos treated with nocodazole, microtubules were absent and epibolic movements of the yolk syncytial nuclei were blocked. In contrast, the vegetal expansion of the enveloping layer and deep cells was only partially inhibited. The process of endocytosis, proposed to play a major role in epiboly of the yolk syncytial layer (Betchaku, T. and Trinkaus, J. P. (1986) Am. Zool. 26, 193-199), was still observed in nocodazole-treated embryos. Treatment of embryos with taxol led to a delay in all epibolic movements. We propose that the yolk cell microtubules contribute either directly or indirectly to all epibolic movements. However, the epibolic movements of the yolk syncytial layer nuclei and of the blastoderm are not coupled, and only movements of the yolk syncytial nuclei are absolutely dependent on microtubules. We hypothesize that the microtubule network of the syncytial layer and the animal-vegetal set of the yolk cytoplasmic layer contribute differently to various aspects of epiboly. Models that address the mechanisms by which the two microtubule arrays might function during epiboly are discussed.

Animals↗
PubMed1993

Vesicular stomatitis virus G glycoprotein pseudotyped retroviral vectors: concentration to very high titer and efficient gene transfer into mammalian and nonmammalian cells.

The restricted host-cell range and low titer of retroviral vectors limit their use for stable gene transfer in eukaryotic cells. To overcome these limitations, we have produced murine leukemia virus-derived vectors in which the retroviral envelope glycoprotein has been completely replaced by the G glycoprotein of vesicular stomatitis virus. Such vectors can be concentrated by ultracentrifugation to titers > 10(9) colony-forming units/ml and can infect cells, such as hamster and fish cell lines, that are ordinarily resistant to infection with vectors containing the retroviral envelope protein. The ability to concentrate vesicular stomatitis virus G glycoprotein pseudotyped vectors will facilitate gene therapy model studies and other gene transfer experiments that require direct delivery of vectors in vivo. The availability of these pseudotyped vectors will also facilitate genetic studies in nonmammalian species, including the important zebrafish developmental system, through the efficient introduction and expression of foreign genes.

Adenoviruses, Human↗
PubMed1993

Characterization of a cell line derived from zebrafish (Brachydanio rerio) embryos.

During the last decade, zebrafish (Brachydanio rerio) have emerged as a novel and attractive system to study embryogenesis and organogenesis in vertebrates. The main reason is that both extensive genetic studies and detailed embryologic analysis are possible using this small tropical fresh water teleost. However, in vitro analysis using cell culture or molecular genetics are still far less advanced than in other vertebrate systems. Here we report the generation and characterization of a fibroblast like cell line, ZF4, derived from 1-day-old zebrafish embryos. The hyperploid cell line has been stable in multiple passages for more than 2 yr now and is the first zebrafish cell line that can be maintained in conventional medium containing mammalian serum. Using a series of plasmids for expression of a marker gene, we evaluate in ZF4 cells the relative strength of expression from several different viral, fish, and mammalian promoters. Stable integration can be obtained by using G418 selection. We hope that our cell line will be a useful tool for the analysis of gene regulation in zebrafish.

Animals↗
PubMed1990

Autonomous determination of anterior structures in the early Drosophila embryo by the bicoid morphogen.

A small number of maternal effect genes determine anterior-posterior pattern in the Drosophila embryo. Embryos from females mutant for the maternal gene bicoid lack head and thorax. bcd mRNA becomes localized to the anterior tip of the egg during oogenesis and is the source for the morphogen gradient of bcd protein. Here we show that in vitro transcribed bicoid mRNA that has its own leader sequences substituted by the Xenopus beta-globin 5' untranslated sequences is translated more efficiently than bicoid mRNA with the natural 5' mRNA leader when tested in vitro and in Drosophila Schneider cells. When injected into bicoid mutant embryos, only the bcd mRNA with the beta-globin leader sequence, substituted for the natural leader, is able to induce anterior development. We used P-transformation to show that sequences in the 5' leader are neither necessary for localization of the transcript nor for the translational block of the bcd mRNA during oogenesis. For our injection experiments, we used only one of the identified splicing forms of bcd mRNA. The bcd protein species derived from this mRNA is able to induce anterior development at any position along the anterior-posterior axis. Thus bicoid protein can induce development of head and thorax independent of any other specifically localized morphogenetic factor. Our findings further support the notion that the concentration gradient of bcd protein, and not the existence of different forms of bcd protein, is responsible for specifying subregions of the embryo.

Animals↗
PubMed1989

Rescue of bicoid mutant Drosophila embryos by bicoid fusion proteins containing heterologous activating sequences.

The maternal gene bicoid (bcd) determines pattern in the anterior half of the Drosophila embryo. It is reported here that the injection of bcd mutant embryos with messenger RNAs that encode proteins consisting of heterologous acidic transcriptional activating sequences fused to the DNA-binding portion of the bcd gene product, can completely restore the anterior pattern of the embryo.

Animals↗
PubMed1989

Determination of spatial domains of zygotic gene expression in the Drosophila embryo by the affinity of binding sites for the bicoid morphogen.

The maternal gene bicoid is a key component of the system that determines the pattern of the anterior half of Drosophila embryos. The bicoid protein forms a concentration gradient in early embryos, and is known to bind DNA. Specific binding sites are now shown to confer expression in a region of the embryo that depends on their affinity for bicoid protein: sites of high affinity allow expression further down the bicoid protein gradient than sites of low affinity.

Animals↗
PubMed1989

Multiple steps in the localization of bicoid RNA to the anterior pole of the Drosophila oocyte.

The anterior region of the Drosophila embryonic pattern is determined by a gradient of the bicoid (bcd) protein. The correct formation of this gradient requires the localization of bcd RNA to the anterior pole of the egg. Here we use a wholemount in situ technique to examine the process of bcd RNA localization during oogenesis and embryogenesis. While bcd protein becomes distributed in a gradient that extends throughout the anterior two thirds of the early embryo, bcd RNA remains restricted to a much smaller region at the anterior pole. The difference between these distributions indicates that the shape of the protein gradient must depend to some extent on the posterior movement of the protein after it has been synthesized. Four distinct phases of bcd RNA localization can be distinguished during oogenesis. Between stages 6 and 9 of oogenesis, the RNA accumulates in a ring at the anterior end of the oocyte. During the second phase, in stage 9-10a follicles, the RNA also localizes to the apical regions of the nurse cells, demonstrating that the nurse cells possess an intrinsic polarity. As the nurse cells contract during stages 10b-11, all of the bcd RNA becomes localized to the cortex at the anterior end of the oocyte. During a final phase that must occur between stage 12 of oogenesis and egg deposition, the RNA becomes localized to a spherical region that occupies a slightly dorsal position at the anterior pole.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

A gradient of bicoid protein in Drosophila embryos.

The maternal gene bicoid (bcd) organizes anterior development in Drosophila. Its mRNA is localized at the anterior tip of the oocyte and early embryo. Antibodies raised against bcd fusion proteins recognize a 55-57 kd doublet band in Western blots of extracts of 0-4 hr old embryos. This protein is absent or reduced in embryonic extracts of nine of the 11 bcd alleles. The protein is concentrated in the nuclei of cleavage stage embryos. It cannot be detected in oocytes, indicating temporal control of bcd mRNA translation. The bcd protein is distributed in an exponential concentration gradient with a maximum at the anterior tip, reaching background levels in the posterior third of the embryo. The gradient is probably generated by diffusion from the local mRNA source and dispersed degradation.

Alleles↗