Worm Breeder's Gazette 11(2): 31
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.
A contig of five cosmids that span 150kb around the Ubq-1 locus on chromosome III was mapped by cos end-labeling and partial cleavage using four different restriction enzymes; cleavage of the cos site was achieved with a crude preparation of lambda DNA terminase (Chow, S., Daub, E., and Murialdo, H., [1987 ] Gene 60, 277-289). We isolated a number of restriction fragments containing potential coding regions using cDNA probes from C. elegans embryo poly(A)+ RNA. Northern analysis with one of these fragments, B3255 obtained from cosmid ZK331, revealed that this presumptive gene encodes a single 3. 1kb mRNA. This message is expressed in both control and heat-shocked embryos. Restriction mapping of the contig placed B3255 at 50kb to the left of Ubq-1. Genomic Southern analysis of C. elegans (N2) and C. Briggsae DNAs using B3255 as a probe under low stringency conditions suggests that this gene is unique. Sequencing of this gene so far reveals a region of DNA encoding 365 amino acids which has 32% similarity to the hamster elongation factor EF-2 (hamEF2). No C. elegans splice consensus sequences are found in this EF-2-like (EF2-L) sequence whereas the hamEF2 gene has 5 introns ranging from 100-130bp long in the corresponding region. The size of the EF2-L mRNA is consistent with that of hamEF2, rat, Drosophila and human EF2 mRNAs. Due to the relatively low degree of amino acid sequence identity between EF2-L and hamEF2 compared to the greater than 80% identity between the corresponding hamster, Drosophila, and human EF-2 sequences, we believe that this gene product is not likely to be C. elegans EF-2. However, further sequence analysis is in progress in order to compare the N-terminal portion of this protein with the sequence motifs found in this region of EF-2, which have been shown to be involved in GDP and GTP binding during the elongation step of protein synthesis. The target of ADP-ribosylation by diphtheria toxin comprising amino acids from positions 704-717 in hamEF2 is highly conserved in many organisms. His-715 in hamEF2 is specifically post-translationally modified to diphthamide in hamEF2. It will be interesting to see whether the C. elegans EF2-L protein also contains this sequence. We are also in the process of carrying out a PCR analysis of C. elegans cDNA and genomic libraries using oligonucleotides from highly conserved regions between hamEF2 and Drosophila EF-2 to see if we can detect the C. elegans EF-2 gene. [See Figure 1]