lin 25 is required for proper vulval cell fate determination (Ferguson and Horvitz. 1985. Genetics 110 17-72). In animals carrying null mutations in
lin-25 ,P6 .padopts a vulval fate whereas all the other VPCs adopt the 3 fate (Ferguson et al., 1987. Nature 326. 259-267 and our own observations). Iin 25 mutations recessively suppress the multivulva phenotype of Iet-60 (gf)and
lin-1 (lf)mutations but not the multivulva phenotype of
lin-12 (d)allele. In a previous gazette article we described our genetic characterization of
lin-25 (WBG 11 No. 5, 77); here we report our progress towards cloning
lin-25 and some preliminary results from genetic mosaic analysis.
lin-25 cloning. We have identified a 10kb fragment that completely rescues the
lin-25 mutant phenotypes (figure 1). When a plasmid containing 10kb subcloned from YL559 was injected at a concentration of 15mg/ml into animals homozygous for the temperature sensitive
lin-25 mutation,
n545 ,more than 80% of the F1 Rollers (n=100) (grown at the restrictive temperature) were rescued for the egg-laying defect. All the lines generated were also rescued. Deletion of 2kb from either end of the 10kb fragment abolished the rescuing ability.
lin-25 genetic mosaics. Temperature shift experiments indicate that
lin-25 is required for the proper development of more than one component of the egg-laying system (Ferguson et al., 1987 and our own observations). We are scoring
lin-25 mosaic animals, therefore, both for the vulval lineage defect and for the ability to lay eggs. To generate mosaics mutant for
lin-25 in the AB lineage we constructed a strain of the genotype
dpy-17 unc-36 ncl-1 ;
unc-42 lin-25 (0);ctDp11 (ctDp11 is-fusion between parts of chromosomes III and V; it complements an of these markers, Hunter and Wood. 1992. Nature 355. 551-555). We picked Unc NonDpy segregants, which were mosaics that had lost ctDp11 in AB or ABp. Half of them were Egl- (n=25). We have followed the vulval lineages in just two such mosaics so far (one AB mosaic and one ABp): both of these had wild-type vulval lineages and were Egl+. To generate P1 mosaics we constructed the strain
sup-18 ncl-1 unc-36 ;
unc-42 lin-25 (0);ctDp11 ;
sup-10 (
n983).We picked wild-type segregants, which were mosaics that had lost ctDpl11 in P1 :
sup-18 is a recessive suppressor of
sup-10 (
n983),whose focus is in P1 .We have found two P1 mosaics so far; both of these were Egl-. There are several possible explanations for these results at the moment. Iin-25 might be required in two different tissues for proper egg-laying the VPCs (for proper fate determination) and a tissue derived from P1 (for some other, as yet unknown, aspect of egg-laying). The wild-type vulval lineages observed in the AB and ABp mosaics could be because of maternal rescue (
lin-25 homozygotes coming from a heterozygote are sometimes Egl+). A second possibility is that focus of
lin-25 for proper vulval cell fate determination might be a tissue derived from both AB and P1 such as
hyp7 .The focus for
lin-15 for VPC fate determination is thought to be
hyp7 (Herman and Hedgecock 1990. Nature 348. 169-171). We are currently examining the vulval lineages in more mosaic animals to distinguish these and other possibilities. [See Figure]