Structure-function relationship of yeast S-II in terms of stimulation of RNA polymerase II, arrest relief, and suppression of 6-azauracil sensitivity.
The yeast S-II null mutant is viable, but the mutation induces sensitivity to 6-azauracil. To examine whether the region needed for stimulation of RNA polymerase II and that for suppression of 6-azauracil sensitivity in the S-II molecule could be separated, we constructed various deletion mutants of S-II and expressed them in the null mutant using the GAL1 promoter to see if the mutant proteins suppressed 6-azauracil sensitivity. We also expressed these constructs in Escherichia coli, purified the mutant proteins to homogeneity, and examined if they stimulated RNA polymerase II. We found that a mutant protein lacking the first 147 amino acid residues suppressed 6-azauracil sensitivity but that removal of 2 additional residues completely abolished the suppression. A mutant protein lacking the first 141 residues had activity to stimulate RNA polymerase II, whereas removal of 10 additional residues completely abolished this activity. We also examined arrest-relief activity of these mutant proteins and found that there is a good correlation between RNA polymerase II-stimulating activity and arrest-relief activity. Therefore, at least the last 168 residues of S-II are sufficient for expressing these three activities.
Nakanishi
T
T
Faculty of Pharmaceutical Sciences, University of Tokyo, Japan.
Shimoaraiso
M
M
Kubo
T
T
Natori
S
S
D000963:Antimetabolites / Q000494:pharmacology*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D012319:RNA Polymerase II / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D017384:Sequence Deletion; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D035165:Transcription Factors, General; D035602:Transcriptional Elongation Factors; D014498:Uracil / Q000031:analogs & derivatives* / Q000494:pharmacology
D000963:Antimetabolites; D005656:Fungal Proteins; D014157:Transcription Factors; D035165:Transcription Factors, General; D035602:Transcriptional Elongation Factors; C018648:transcription factor S-II; D014498:Uracil; D012319:RNA Polymerase II; C100248:azauracil