TRANSPOSON INSERTIONS CAUSING CONSTITUTIVE SEX-LETHAL ACTIVITY IN DROSOPHILA-MELANOGASTER AFFECT SXL SEX-SPECIFIC TRANSCRIPT SPLICING

Sex-lethal (Sxl) gene products induce female development in Drosophila melanogaster and suppress the transcriptional hyperactivation of X-li nked genes responsible for male X-chromosome dosage compensation. Cont rol of Sxl functioning by the dose of X-chromosomes normally ensures t hat the female-specific functions of this developmental switch gene ar e only expressed in diplo-X individuals. Although the immediate effect of X-chromosome dose is on Sxl transcription, during most of the life cycle ''on'' vs. ''off'' reflects alternative Sxl RNA splicing, with the female (productive) splicing mode maintained by a positive feedbac k activity of SXL protein on Sxl pre-mRNA splicing. ''Male-lethal'' (S xl(M)) gain-of-function alleles subvert Sxl control by X-chromosome do se, allowing female Sxl functions to be expressed independent of the p ositive regulators upstream of Sxl. As a consequence, Sxl(M) haplo-X a nimals ( chromosomal males) die because of improper dosage compensatio n, and Sxl(M) chromosomal females survive the otherwise lethal effects of mutations in upstream positive regulators. Five independent sponta neous Sxl(M) alleles were shown previously to be transposon insertions into what was subsequently found to be the region of regulated sex-sp ecific Sxl RNA splicing. We show that these five alleles represent thr ee different mutant types: Sxl(M1), Sxl(M3), and Sxl(M4). Sxl(M1) is a n insertion of a roo element 674 bp downstream of the translation-term inating male-specific exon. Sxl(M3) is an insertion of a hobo transpos on (not 297 as previously reported) into the 3' splice site of the mal e exon, and Sxl(M4) is an insertion of a novel transposon into the mal e-specific exon itself. We show that these three gain-of-function muta nts differ considerably in their ability to bypass the sex determinati on signal,with Sxl(M4) being the strongest and Sxl(M1) the weakest. Th is difference is also reflected in effects of these mutations on sex-s pecific RNA splicing and on the rate of appearance of SXL protein in m ale embryos. Transcript analysis of double-mutant male-viable Sxl(M) d erivatives in which the Sxl(M) insertion is cis to loss-of-function mu tations, combined with other results reported here, indicates that the constitutive character of these Sxl(M) alleles is a consequence of an alteration of the structure of the pre-mRNA that allows some level of female splicing to occur even in the absence of functional SXL protei n. Surprisingly, however, most of the constitutive character of Sxl(M) alleles appears to depend on the mutant alleles' responsiveness, perh aps greater than wild-type, to the autoregulatory splicing activity of the wild-type SXL proteins they produce.