Worm Breeder's Gazette 11(4): 35
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.
From Chris Link's lambda library of DNAs from WS9-6, a male-female Caenorhabditis strain, I isolated an apparent mec-3 homologue. A 4kb EcoRI fragment contains exons corresponding to all the exons of mec-3 in C. elegans, as well as four long conserved regions upstream of the start codons. The regions of sequence similarity between the C. elegans and WS9-6 mec-3 genes are very much like the regions of similarity between C. elegans and C. briggsae described by Ding Xue and Marty Chalfie in the last newsletter (including their revision of the coding sequence), except that the overall level of similarity between the C. elegans and WS9-6 genes seems higher. There are four long upstream regions of conservation of 71,30,30 and 24 bp, and a few shorter ones. In the entire coding sequence, there are 29 amino acid substitutions, 19 of which are between the LIM and homeodomains. There are no amino acid changes in the homeodomain. In introns, there is no apparent similarity between the sequences. Also, the WS9-6 introns are generally shorter: the first intron, 395 bp in C. elegans, is only 73 bp in WS9-6. The fact that the intron sequences have completely drifted suggests that the conserved upstream regions have been preserved by selection. The four conserved upstream regions (hereafter termed regions I, II, III and IV) have distinctive features that may relate to how mec-3 is regulated. Regions I and II each have sequences very similar to the POU consensus binding site, and region III contains a close relative to the ISL-1 binding site (ISL-1 is a transcription factor for the insulin gene and is closely related to mec-3.) Region IV contains no obvious sequence features, but can be deleted to cause expression of a mec-3-lacZ fusion in extra cells (Way, WBG 11,2). A provisional organization scheme for this region would thus be that regions I and II mediate unc-86 binding and establishment expression, region III mediates mec-3 binding and maintenance expression, and region IV mediates negative regulation by unidentified proteins. With unc-86 protein from Mike Finney, I have shown that a restriction fragment containing regions I, II and III can be 'band- shifted'. I have not yet done a footprint to identify the actual unc- 86 binding sites. (Since C. elegans and C. briggsae are both hermaphrodite species while WS9-6 is male/female, it would appear that the male/female sex pattern is derived from the male/hermaphrodite pattern. It would be interesting to see which of the sex-determination genes of C. elegans have been altered in WS9-6 to account for this change.)