Bruce W. Draper

The maternal gene skn-1 encodes a protein that is distributed unequally in early C. elegans embryos

The autonomous or cell-intrinsic developmental properties of early embryonic blastomeres in nematodes are thought to result from the action of maternally provided determinants. After the first cleavage of the C. elegans embryo, only the posterior blastomere, P1, has a cell-intrinsic ability to produce pharyngeal cells. The product of the maternal gene skn-1 is required for P1 to produce pharyngeal cells. We show here that the Skn-1 protein is nuclear localized and that P1 appears to accumulate markedly higher levels of Skn-1 protein than its sister, the AB blastomere. We have examined the distribution of Skn-1 protein in embryos from mothers with maternal-effect mutations in the genes mex-1, par-1, and pie-1. These results suggest that mex-1(+) and par-1(+) activities are required for the unequal distribution of the Skn-1 protein and that pie-1(+) activity may function to regulate the activity of Skn-1 protein in the descendants of the posterior blastomere P1.

MEX-3 Is a KH Domain Protein That Regulates Blastomere Identity in Early C. elegans Embryos

After the first division of the C. elegans embryo, the posterior blastomere can produce numerous muscles while the anterior blastomere cannot. We show here that maternal-effect lethal mutations in the gene mex-3 cause descendants of the anterior blastomere to produce muscles by a pattern of development similar to that of a descendant of the wild-type posterior blastomere. mex-3 encodes a probable RNA-binding protein that is distributed unequally in early embryos and that is a component of germline-specific granules called P granules. We propose that MEX-3 contributes to anterior-posterior asymmetry by regulating one or more mRNAs involved in specifying the fate of the posterior blastomere.

The pie-1 and mex-1 genes and maternal control of blastomere identity in early C. elegans embryos

During C. elegans embryogenesis an 8-cell stage blastomere, called MS, undergoes a reproducible cleavage pattern, producing pharyngeal cells, body wall muscles, and cell deaths. We show here that maternal-effect mutations in the pie-1 and mex-1 genes cause additional 8-cell stage blastomeres to adopt a fate very similar to that of the wild-type MS blastomere. In pie-1 mutants one additional posterior blastomere adopts an MS-like fate, and in mex-1 mutants four additional anterior blastomeres adopt an MS-like fate. We propose that maternally provided pie-1(+) and mex-1(+) gene products may function in the early embryo to localize or regulate factors that determine the fate of the MS blastomere.

The maternal genes apx-1 and glp-1 and establishment of dorsal-ventral polarity in the early C. elegans embryo

The sister blastomeres ABp and ABa are equipotent at the beginning of the 4-cell stage in C. elegans embryos, but soon become committed to different fates. We show that the glp-1 gene, a homolog of the Notch gene of Drosophila, functions in two distinct cell-cell interactions that specify the ABp and ABa fates. These interactions both require maternal expression of glp-1. We show that a second maternal gene, apx-1, functions with glp-1 only in the specification of the ABp fate and that apx-1 can encode a protein homologous to the Delta protein of Drosophila. Our results suggest how interactions mediated by glp-1 and apx-1 contribute to the establishment of the dorsal-ventral axis in the early C. elegans embryo.

The zebrafish fgf24 mutant identifies an additional level of Fgf signaling involved in vertebrate forelimb initiation

The development of vertebrate limb buds is triggered in the lateral plate mesoderm by a cascade of genes, including members of the Fgf and Wnt families, as well as the transcription factor tbx5. Fgf8, which is expressed in the intermediate mesoderm, is thought to initiate forelimb formation by activating wnt2b, which then induces the expression of tbx5 in the adjacent lateral plate mesoderm. Tbx5, in turn, is required for the activation of fgf10, which relays the limb inducing signal to the overlying ectoderm. We show that the zebrafish fgf24 gene, which belongs to the Fgf8/17/18 subfamily of Fgf ligands, acts downstream of tbx5 to activate fgf10 expression in the lateral plate mesoderm. We also show that fgf24 activity is necessary for the migration of tbx5-expressing cells to the fin bud, and for the activation of shh, but not hand2, expression in the posterior fin bud.

A high-throughput method for identifying N-ethyl-N-nitrosourea (ENU)-induced point mutations in zebrafish.

Publisher Summary This chapter focuses on the high-throughput method for identifying n-ethyl-n-nitrosourea (ENU). Prior to screening for ENU-induced mutations in zebrafish, a library is generated consisting of cryopreserved sperm isolated from the F1 progeny of ENU-mutagenized males. Although it is possible to screen for mutations in fish that are kept alive, a cryopreserved library has several advantages. The zebrafish has become an important model system for vertebrate biology. Although forward genetic screens have uncovered the functions of many zebrafish genes, until recently no reliable, inexpensive, and high-throughput technology for targeted gene disruptions was developed. Zebrafish forward genetic screens have been—and continue to be—exceptionally productive. However, as the content of the zebrafish genome becomes available in the form of primary sequence information, it becomes increasingly evident that many essential genes have not been identified by this approach. Mutant phenotypes might be subtle or even undetectable in forward genetic screens because of the nature of the screen. For example, most genetic screens performed to date have focused on identifying phenotypes during the embryonic period while the embryo is still transparent and have therefore been easy to screen for morphological defects in the light microscope or following staining with tissue-specific markers.