Worm Breeder's Gazette 11(3): 51
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.
Genetic analysis of dpy-29 has provided evidence that this locus defines a single, cryptic, sdc-like gene with separately mutable sex determination and dosage compensation functions. However, unlike sdc- 1 and sdc-2, mutations in dpy-29 exhibit two distinct classes of phenotypes, Tra or Dpy. The canonical dpy-29(Tra) allele, y52, causes transformation of XX animals into pseudomales, but shows no evidence of dosage compensation defects. In contrast, XX animals homozygous for a null allele of dpy-29 exhibit only the phenotypes associated with the disruption of dosage compensation (i.e. XX-specific lethality and dumpiness), but no masculinization. The masculinization caused by Tra alleles of dpy-29 can be suppressed in two ways. First, mutations that upset dosage compensation in XX animals (either in the dosage compensation function of dpy-29, or in the dosage compensation genes dpy-21, dpy-27, or dpy-28) suppress the transformation by dpy-29(Tra) alleles. Second, Tra alleles can be suppressed by mutations in her-1, suggesting that dpy-29(Tra) mutations disrupt the negative regulation of her-1 in XX animals ( DeLong, Plenefisch and Meyer, WBG 11(2): 118). Since her-1 is known to be regulated at the mRNA level (C. Trent and W. Wood, personal communication), we investigated whether y52 causes increased her-1 mRNA levels in XX animals as predicted by the genetics. Furthermore, since upsets in dosage compensation suppress this masculinization, we were interested in determining if this suppression was also evident in the level of her-1 mRNA In order to address these questions, we have measured steady state her-1 mRNA levels in various strains using Northern analysis. All mRNA was isolated from eggs in order to equalize for stage-specific differences in her-1 expression. Furthermore, all her-1 mRNA levels were normalized to act-1 mRNA levels to control for amounts of mRNA loaded. dpy-29(Tra) mutations cause overexpression of her-1 transcripts. mRNA isolated from the weak dpy-29(Tra) mutation y137, which causes 26% of the XX animals to be masculinized, showed 20-fold elevation of her-1 mRNA levels compared to N2. The strong dpy-29(Tra) mutation, y52, showed an approximately 50-fold elevation in her-1 levels. In order to obtain eggs from y52, the masculinization phenotype was suppressed by her-1(e1520), which blocks her-1 gene function but does not effect her-1 mRNA levels in XX or XO animals (C. Trent, personal communication). Disruptions in dosage compensation suppress overexpression of her-1 in dpy-29(y52) XX animals. Masculinization caused by dpy-29(Tra) mutations is suppressed by upsets in dosage compensation. Therefore, we were interested in determining if the suppression of dpy-29(Tra) by dosage compensation defects acted through her-1 or if it affected the sex determination pathway at a step downstream of her-1. We have found that disruptions in dosage compensation do in fact cause a substantial decrease in her-1 mRNA levels in dpy-29(Tra) animals, indicating the suppression is mediated either upstream of, or directly on, her-1. Both dpy-27(y57); 2) and dpy-28(y1ts); dpy-29(y62) have significantly lower her-1 mRNA levels as compared to the levels in dpy-29(y52) in the absence of a dosage compensation defect. This is particularly significant in the case of suppression by dpy-28(y1ts) because the animals were grown at the permissive temperature where the dosage compensation defect is slight. However, in both cases some residual expression of her-1 mRNA above N2 levels remained despite the complete phenotypic suppression of dpy-29(y52).In addition, we have examined the effects of loss-of-function mutations in both the dosage compensation(y180) and sex determination (y62) functions of dpy-29 on her-1 mRNA The balanced strain dpy-29(y52y180) /unc-61(e228) 11) showed a reduced her-1 mRNA level compared to the dpy-29(y62)/unc-61(e228) 11) strain, but has an equivalent level to that seen in N2. We conclude that the sex determination function of dpy-29 is required for the proper negative regulation of her-1 mRNA levels in XX animals. However, defects in dosage compensation suppress the masculinization of dpy-29(Tra) XX animals by restoring her-1 mRNA levels to nearly normal. [See Figure 1]