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Abstract:
We are studying the evolution of cell fate specification, using vulva development as a model system and compare C. elegans with Pristionchus pacificus. In P. pacificus, seven ventral epidermal cells P(1-4,9-11).p undergo programmed cell death during late embryogenesis.P(5-7.p) form the vulva and P8.p remains epidermal but influences the fate of P(5-7).p. We performed various genetic screens to isolate a number of mutants defective in vulva formation, many of which result in phenotypes unknown in C.elegans . It has previously been shown that the lin-39 homologue of P. pacificus prevents apoptosis of P(5-8).p and that mutations in Ppa-lin-39 result in a generation vulvaless phenotype. Double mutants of lin-39 with the cell death gene ced-3 indicated that Ppa lin-39 has an early role in vulva development in P. pacificus. It prevents cell death but is dispensable for vulva induction, which is in contrast to Cel lin-39. Several genetic screens were carried out, to isolate new generation vulvaless mutants. We have identified seven novel mutations not allelic to Ppa-lin-39. These new mutations fall into 3 new complementation groups, viz. ped-12, gev-1 and gev-2. ped-12 animals are egg-laying defective and haplo-insufficient, i.e. heterozygous animals show a protruding vulva phenotype. Males show pleiotropic defects like absence of rays and spicules and a malformed rectum. Double mutants of ped-12 with the cell death gene ced-3 resulted in a vulvaless phenotype indicating that Ppa ped-12 has both an early and a late role in vulva development in P. pacificus. To facilitate the mapping and molecular characterization of these mutants, we constructed a genetic map of Pristionchus pacificus. (Srinivasan et al. 2002). ped-12 mapped to Chromosome-I in P.pacificus. Fine mapping of ped-12placed it close to the BAC 21N20 containing the P.pacificus orthologue of the lin-2gene.To narrow down the region containing the gene , I looked for recombinants on both sides of the BAC 21N20. I obtained 3 recominants on 22D6, the neighbouring BAC on the left and 2 recombinants on 13C1, the BAC on the right. This indicates that ped-12 lies in this region. Currently shotgun sequencing of the BAC 21N20 is in progress to find suitable candidate genes.
