Worm Breeder's Gazette 12(2): 68 (January 1, 1992)

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.

Promoting her-1

Weiqing Li, Marc Perry, Bill Wood

Figure 1

MCD Biology University of Colorado, Boulder CO 80309-0347

The her-1 gene is required for normal male development in C. elegans. Previous work (Trent et al. M.o.D. 34:43,1991; Perry et al., C. elegans Mtg. Abstr. 1992, p.230) has shown that the gene includes four exons and gives rise to a rare larger and an abundant smaller transcript as shown below. To confirm that these RNA's have different promoters as suggested by RNase protection assays (Robertson et al., this issue) and to analyze their activities, we made reporter constructs by fusing the LacZ gene to each of two DNA fragments containing putative promoter regions (see below): a) the 2.5 kb 5' flanking region of exon 1 (putative P1 ),and b) the entire 3.4 kb intron 2 (putative P2 ).Each of the constructs was co-injected with cloned rol-6 ( su1006 )DNA into a him-8 ( e1489 )strain, which produces about 40% XO progeny. Stable transformant lines were obtained and stained with X-gal. For the P1 construct, no ß-galactosidase activity has been detected in the embryos, possibly because of the weakness of this promoter. For the P2 construct, staining is observed in up to 40% of the embryos, from about the 30-cell stage onward. Most cells are stained, suggesting that expression is not limited to certain cell lineages. These results prove the presence of a second promoter in the her-1 locus. When an integrated P2 construct was crossed into an N2 background to determine its activity in XX embryos, no ß-galactosidase activity was detected except in a few cells in late embryos. This result is also consistent with RNase-protection results (ibid.). When the same construct was introduced into the her-1 deletion strain him-8 ( e1489 ); her-1 ( y101 hv1 )(all hermaphrodites, but 40% XO), it showed staining similar to that seen in the him-8 ( e1489 )strain. We conclude that P2 activity is XO-specific and is independent of her-1 expression. We also tried to detect P1 activity by observing its biological function, transforming the him-8 ; her-1 strain with a construct (c) carrying the putative P1 region, the four exons, and only the first intron. Transformed animals produced strongly masculinized progeny including fertile males, indicating that the large transcript can be sufficient for male development.

[See Figure 1]

Figure 1