Worm Breeder's Gazette 10(3): 46
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.
The deg-1(u38) mutation results in swelling and degeneration of certain neurons (including the PVC cells, whose deaths impart a tail touch insensitive phenotype). The swelling of the affected cells suggests some type of membrane defect which eventually allows water to enter the cell. We have found that the deg-1 gene product may in fact be an integral membrane protein. This conclusion is based on cDNA sequence analysis (described below). Localization of the deg-1 gene product to the cell membrane is consistent with the idea that the deg- 1 induced degenerations result from a membrane defect. We have been sequencing three cDNA's obtained from the Ahringer/Kimble 'L1' library. All three hybridize to the same genomic fragment of the deg-1 gene under stringent conditions. The three cDNA's appear to represent transcripts which have undergone different sets of splicing events. All.three contain long stretches of identical sequence, which supports the conclusion that they represent transcripts of the same gene, rather than transcripts of other members of a hypothesized deg-1 gene family (see our abstract in last WBG). One of the cDNA's, described in June at the Boston meeting, contains a deduced open reading frame encoding at least 340 amino acids. Hydrophobicity plots of the translated protein indicate two possible membrane spanning segments, located between residues 191 to 231 and residues 275 to 316. Eleven of the fifteen cysteine residues present in the sequence are located in the first 191 residues, upstream of the first hydrophobic region. It is possible that the first 191 amino acids are part of an extracellular domain, since intracellular conditions do not favor cross-linking of cysteines. This cDNA does not contain a poly A tail. The second cDNA may be the most likely to represent a mature transcript, since it contains a poly A region of about sixty bases following a poly A addition site. This cDNA contains an open reading frame encoding 338 amino acids, which is nearly identical to that of the first cDNA, with two important exceptions. The first difference is a 72 base insertion that would result in a novel 24 amino acid sequence following amino acid 176 of the protein encoded by the first cDNA. Second, the termination codons and poly A tail of the second cDNA occur at the end of the second possible membrane spanning region encoded by the first cDNA. This would result in a protein at least twenty five amino acids shorter than that encoded by the first cDNA. The two cDNA's are probably identical at their 5' ends. However, it is possible that both are incomplete due to the usual imperfections of cDNA libraries. The third cDNA appears to represent an incompletely spliced transcript. It contains relatively short open reading frames ( some of which encode sequences found in the proteins translated from the first two cDNA's). These are flanked by A/T rich intron-like regions. Consensus splice sites can be found at several intron/exon junctions. Comparison of the sequences of the three cDNA clones suggests that deg-1 transcripts may be processed to produce different splicing patterns. In the case of the first two cDNA's, splicing differences would result in different exon usage and different protein products. It is also possible that the differences between the cDNA's arose as artifacts of library construction or subsequent cloning events. We plan to perform RNA protection experiments to see if multiple forms of deg-1 transcripts can in fact be isolated from wild type worms and from deg-1 mutants. The putative deg-1 protein sequences do not show significant homology to proteins in the data bases screened so far.