SELECTION AND ANALYSIS OF RARE 2ND-SITE SUPPRESSORS OF DROSOPHILA RNA-POLYMERASE-II MUTATION

We used a mutagenesis and selection procedure in Drosophila melanogast er to recover rare allele-specific suppressor mutations. More than 11 million flies mutant for one of five recessive-lethal mutations in the two largest subunits of RNA polymerase II were selected for additiona l mutations that restored viability. Forty one suppressor mutations we re recovered. At least 16 are extragenic, identifying a minimum of thr ee loci, two of which do not map near genes known to encode subunits o f RNA polymerase II. At most, 25 are intragenic, 4 reverting the initi al altered nucleotide back to wild type. Sequence analysis of interact ing mutations in the two largest subunits identified a discrete domain in each subunit. These domains might be contact points for the subuni ts. Finally, our selections were large enough to allow recovery of mul tiple independent changes in the same nucleotides yet mutations in oth er equally likely targets were not recovered. The mutations recovered are not random and might provide insights into possible mechanisms for mutagenesis in eukaryotes.