Worm Breeder's Gazette 9(3): 61
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 have been continuing our analysis of ama-1 IV and envirous, the cloning of which has been reported in a previous edition (WBG, 9(1)). Using a conventional walking approach, we have expanded the area around the region of Drosophila homology to about 29 kb [Sulston and Coulson et al. were unable to find a cosmid contig, perhaps as a result of the paucity of HindIII sites]. About 20 kb of the ama-1 region is shown in Figure 1. Using restriction fragments spanning this region (bottom line of Figure I ), we have, barring any very large introns, delineated the approximate extent of the region encoding the 5.9 kb ama-1 transcript. Based on hybridization signal strength, ama-1 transcripts are about 150 times less abundant than unc-54 transcripts in total RNA. While doing northerns, we observed that a probe approximately 5 kb rightward of ama- I detected an abundant smear of transcripts at 1.2 - 1.4 kb. Since this resembled the pattern for a collagen probe, we sent a flanking sequence to Jim Kramer who very kindly screened the collagen gene bank (Cox et al., MCB, 4, 2389, 1984), identifying this as the collagen gene in CG41. CG41 extends an additional 2 kb more rightward than shown in Figure 1. This collagen gene has been assigned the gene name col-33.{Figure 1} The analogues of ama- I in yeast (RP02 1 ) and mouse have been sequenced (Allison et aL, Cell, 42, 599, 1985; M. Bartolemei, personal communication). Both were found to encode a C-terminal domain comprised of an heptapeptide, with consensus [tyr-ser-pro-thr-ser-pro- ser], tandemly repeated (26 times in yeast and 52 times in mouse). Loss of this domain during preparation of the pol II enzyme from yeast cells results in a characteristic 190 k degradation product, which is also observed for C. M. Golomb, personal communication), suggesting that worms may also have this strange tail. To test this, we have begun DNA sequencing the region that we suspect encodes the C-terminus of the ama- I product. Although preliminary, we have detected regions of strong homology between the ama- I sequence and the heptapeptide tail of RP02 1 (Figure 2), thereby orienting the ama- I transcription unit S' - 3' as shown left to right in Figure 1. We haven't yet observed the tandem array seen in yeast or mouse, although this motif may be present a little further 5', where hybridization to the Drosophila probe is much stronger (solid box in Figure 1). Sequencing in this area is currently in progress. {Figure 2} We are continuing to walk leftward from ama-1 so as to locate both the mDf4 breakpoint and dpy-13 (see Albert and Riddle, this issue).