Alteration of hsp82 gene expression by the gypsy transposon and suppressor genes in Drosophila melanogaster.

Abstract

Numerous spontaneous mutations at various loci inDrosophila melanogaster result from the insertion of retrotransposons. The mutant phenotypes of many of these insertion alleles are made less severe (suppressed) or more severe (enhanced) by mutations in certain allele-specific modifier genes. An example of these phenomena is provided by the 7.5-kb gypsy element, insertions of which are associated with nearly all of the alleles that are suppressed by mutations of suppressor of Hairy wing [su(Hw)] (Modolell et al. 1983). Some of the gypsy-insertion alleles suppressed by su (Hw) promoter effects. The identification and investigation of specific mechanisms, however, have been limited by uncertainties regarding the complex patterns of transcripts of the loci that have been studied.To overcome these difficulties, we have made use of a simple inducible gene with a single intron, the hsp82 heat shock gene. Without gypsy, the hsp82 gene is transcribed in response to heat shock to give two prominent RNAs, the full-length transcript and its spliced product. We investigated the effects on transcript formation, in transfected cells and in transformed Drosophila pupae, of inserting the gypsy element or specific fragments of it into the hsp82 intron. We also examined the effects of mutations in the modifier genes su(Hw), su(f), and su(w~). Here we present these observations and discuss their implications for the mechanisms of mutation by retrotransposon insertions and for the effects of modifier genes. ResultsThe

Nipped-A, the Tra1/TRRAP Subunit of the Drosophila SAGA and Tip60 Complexes, Has Multiple Roles in Notch Signaling during Wing Development

“…Northern blot assays were performed with formaldehyde-agarose gels and radioactively labeled single-stranded RNA probes as previously described (14). Total RNA was isolated from second-instar larvae with Trizol (Gibco BRL).…”

Drosophila retrotransposon promoter includes an essential sequence at the initiation site and requires a downstream sequence for full activity.

“…RNA was prepared from transfected cells as described (6). mRNA was selected by oligo(dT)-cellulose chromatography (10).…”

“…Drosophila Schneider 2 cells (-5 x 107) were transfected in 10 ml with 50 ,ug of gypsy-CAT DNA and 5 tg of heat shock protein hsp82-lacZ DNA (6) by the calcium phosphate method (7). Cells were harvested after 48 hr and extracted for CAT assays with '4C-labeled chloramphenicol (8).…”

Transposable elements P and gypsy in natural populations of Drosophila willistoni

“…These enzymes were chosen because the fragment pattern they generate provides information on the presence of complete or deleted elements, probable copy Sassi et al 735 number and the occurrence of restriction sites in the genomes of the flies based on the expected the restriction pattern the D. melanogaster elements. The probes used were the complete 2.9 kb sequence of the D. melanogaster P canonic element, contained in the pp 25.1 plasmid (O'Hare and Rubin, 1983) and a 6.9 kb fragment liberated from the pGGHS plasmid by the D. melanogaster gypsy retroelement restriction enzyme XhoI (Dorsett et al, 1989). For each sample, the DNA fragments were separated on 1% (w/v) agarose gel and transferred to a Hybond N + membrane (GE Healthcare) and hybridized to the random prime-labeled probes at 60°C in a mixture containing 0.1% (w/v) SDS, 5% dextran sulfate and a 20-fold dilution of liquid block (Gene Image kit, GE Healthcare) in 5X SSC.…”