Worm Breeder's Gazette 10(3): 11
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 described earlier the isolation of ct31 as a gamma-ray-induced, semi-dominant, feminizing mutation that suppresses the masculinization of XX animals caused by the her-1(gf) allele n695, and reported that ct31 increases the penetrance of the X-linked hypomorph lin-15(n765), suggesting that ct31 might cause a decrease in X-chromosome expression (Burgess et al., WBG May, 1986). ct31 XX animals are Dpy, Egl hermaphrodites; XO animals are Dpy, nonfertile, partially feminized males. Here we present evidence that ct31 lowers expression of the X- linked act-4 gene and that it is probably a gain-of-function mutation involving duplication of a region of the X chromosome. Assay of act-4 mRNA in ct31 and N2 L1 hermaphrodites using an RNA dot blot assay ( Donahue et al., PNAS 84:7600,1987) showed that the level of this transcript in ct31 animals is only 46% + 4% of the level in N2 animals. This finding supports the genetic evidence cited above that ct31 causes a decrease in expression of X-linked genes. Three-factor mapping data places ct31 in the lin-2 - unc-9 interval on X. Deficiencies of this region have not been obtained, but it is covered by the duplication mnDp10. XX hermaphrodites of genotype mnDp10/+; ct31 lin15, which have two mutant and one wild-type copy of the ct31 locus, are more severely Dpy and Egl and generally sicker and more slow-growing than ct31 homozygotes. When the former animals are allowed to self-fertilize, one quarter of the progeny arrest as L1 larvae. We surmise that these arrested larvae are the class carrying two copies of mnDp10 (ct31/ct31/+/+). Since the ct31 phenotype appears to be enhanced by additional copies of the wildtype locus, these results are consistent with ct31 being a gain-of-function mutation. However, given the large size of mnDp10, and the fact that it carries at least one other locus known to affect X expression (sdc- 1), more definitive evidence on this point must await isolation and characterization of deficiencies in the region. The high reversion rate of ct31 described earlier (Burgess et al., 1986) suggested that it might involve an unstable chromosomal rearrangement. In an attempt to test this suggestion, we used a probe derived from the cloned myo-2 gene, which is located on the physical map at a position that should be near the ct31 locus, to carry out quantitative Southern blot analyses. The copy number of the myo-2 gene in the ct31 strain is clearly higher than in N2 or in the her-1(n695) parent strain of ct31. The elevated copy number of myo-2 appears identical to that measured in a strain homozygous for mnDp10. Seven independent nonDpy ct31 revertants do not show the elevated myo-2 copy number, correlating the visible ct31 phenotype and the apparent duplication. Preliminary experiments using as probes cosmid clones from the myo-2 contig ( obtained from A. Coulson and J. Sulston) indicate that at least 40 kb including and to the left of the myo-2 gene is duplicated in the ct31 mutant. Experiments are in progress to characterize further the extent and nature of the rearrangement.