A search of the C. elegans genome database has revealed a total of 6 core 2 N-acetylglucosaminyltransferase (Core 2 GlcNAc-T) homologues. Of these six, no single homologue appears to be more significantly similar to its mammalian counterpart than any other. We have undertaken studies to determine the function in C. elegans of all of the members of this family. To this end, cDNA clones have been isolated by RT-PCR for all six homologues. Overall, the observed splice junctions are in good agreement with those predicted by Gene Finder. cDNA fragments have been used to conduct RNA interference experiments for C54C8.11, F22D6.11 and F22D6.12. gld-1
injected in parallel was used as the positive control. The core 2 GlcNAc-T RNAi's have yielded no obvious aberrant phenotypes. Fully coding cDNA constructs of all 6 family members are being derived and transgenic lines developed that express these gene-products under the control of the heat-shock promoter. Induced protein will be immunopurified and tested for enzyme activity. Tc1 mediated mutagenesis has been used to isolate a 2 bp frameshift allele (ev686
) of F44F4.6 ( gly-1
). This causes premature termination of the polypeptide prior to the putative catalytic domain and is presumably null. Phenotypic characterization of this strain has revealed a ray 1 defect in male tails. The defect shows incomplete penetrance and is temperature sensitive with respect to its penetrance. Work is in progress to determine if the defect is due to a mispositioning, fusion or loss of the ray.