Worm Breeder's Gazette 10(2): 108
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
We are cloning the mab-5 gene to study the molecular mechanism by which patterns of cell types arise along the anteroposterior body axis of C. elegans after hatching. As a brief review, mab-5 is required to give cells located in the posterior body region their normal identities. This gene influences the fates of epidermal, neural, and mesodermal cells in a cell-autonomous fashion. It affects sequences of cell division, cell death, fates of specific neurons, production of mating structures and sensilla, cell fusion, and also directions of cell migration. The simplest way to explain most of the differences between mab-5 mutants and the wild type is to postulate that this gene allows posterior cells to develop differently than they would if located further anteriorly. Differences between mab-5 and wild type males and hermaphrodites also suggest that mab-5 interacts with elements of the sex determination pathway to generate much sexual dimorphism in the posterior body region. We identified and cloned a closely linked Tc1 element in the strain N62. We obtained cosmids in this region from John Sulston and Alan Coulson (thanks!) and used them to probe Southern blots of mab-5 mutants. In the mab-5 (e2088) mutant, we detected a deletion of about 150 bp located approximately one cosmid length, or 35 Kb, from the Tc1 element. To test whether this deletion could be in the mab-5 gene, we carried out high resolution mapping experiments. We found that the Mab-5 phenotype and the deletion always cosegregated in 102 recombinants within the 0.5 mu sma-3 to unc-36 interval. Since we examined 38 crossovers in the approximately 315 Kb interval between mab-5 and unc-36 (A. Coulson, pers. comm.), we expect about one crossover per 8 Kb. Thus it seems quite likely that this deletion is the mab-5 mutation. We have sequenced wild type genomic DNA in this region and determined that part of a large open reading frame is deleted in e2088. By Northern analysis, we found that this open reading frame is part of a polyadenylated RNA present in larvae (we have yet to look at other developmental stages). Currently, we are doing Northern analysis of the mab-5 mutants and transformation experiments to provide further evidence that we have cloned mab-5. We have sequenced partial cDNAs containing this open reading frame isolated from a library kindly given to us by C. Thacker and M. Capecchi. By comparing the sequence to known protein sequences, we have found that the open reading frame contains a homeobox highly homologous to those of Drosophila pattern formation genes. This putative mab-5 homeodomain is most similar to that of the Antennapedia gene. Of the 60 amino acids in the mab-5 homeodomain, 44 are identical to those of Antennapedia, and only 4 of the substitutions are not conservative. The figure below shows these homeodomains using dashes to represent amino acids identical to those of mab-5 with conservative substitutions underlined. Assuming that this sequence corresponds to mab-5, this similarity indicates that homeodomains are used to generate global aspects of body pattern in animals other than Drosophila. At a more mechanistic level, since homeodomain proteins are thought to be transcriptional regulators, it also suggests that mab-5 exerts its many influences on cell division, differentiation, and migration by regulating gene expression. [See Figure 1]