Spontaneous mutation in Drosophila melanogaster are frequently caused by insertions of retrotransposons. Mutant phenotypes result from insertions into both coding and noncoding regions, and the mutagenic effects caused by insertions into coding and noncoding regions are not well understood. Some phenotypes resulting from retrotransposoninsertions can be reversed by extrangenic suppressor and intragenic reversion mutations. The long term objective of this study is to understand how these mobile elements disrupt the expression of nearby genes and to elucidate the role of suppressors in modulating the mutagenic activity of retrotransposons. A suppressible mutation caused by insertion of the 412 element into the vermilion (v) gene of Drosophila is being investigated. In this case, the 412 element inserted into the 5' untranslated region of the first exon, and transposable element sequences are almost entirely eliminated from the mature RNA by splicing at sites near the ends of 412. Mutations at suppressor of sable (su(s)) cause increased accumulation of nearly wild-type sized v transcripts. One objective of this proposal is to investigate the mechanism of suppression. The proposed experiments will define whether su(s) mutations affect transcription, RNA stability or splicing. Other experiments will indicate whether specific 412 sequences interfere with v RNA accumulation and are required for the response to su(s) mutations or whether interference is due to the position of the 412 insertion. A second objective is to investigate the mechanism of intragenic reversion. In one v revertant, a second retrotransposon inserted into one end of the 412 element, altering the splicing pattern. The proposed experiments will ascertain whether the secondary insertion significantly improves the rate of splicing to remove transposon sequences. If so, the effect of changes in splice sites on the response to su(s) mutations will be determined. These goals will be accomplished by examining levels of precursor and mature v RNA produced by in vitro generated variants of the mutant and revertant alleles after transformation into the Drosophila germ line.***
