Suppressors of the cold-sensitivity of the yeast Rpb3-K9E substitution
We performed a genome-wide selection for spontaneous mutations that correct the cold-sensitive growth defect caused by a Lys-to-Glu substitution in residue 9 (K9E) of the Rpb3 subunit of brewer's yeast RNA polymerase II. This substitution was selected for readthrough of a snoRNA (SNR13) terminator placed in the intron of the ACT1/CUP1 reporter gene. The suppressor mutations could identify a protein that is interacting with the Rpb3 subunit of RNAP II that influences transcription termination. From 20 cold-resistant phenotypic revertants, one contains an R317G substitution in HRP1 and Q90E substitution in ADE4. Eight contain a nonsense-to-missense substitution in the ade2-1 allele and one is a true revertant of this nonsense codon to Glu at position 64 of the ade2-1 allele. Eight contain an r.35G>T substitution in one of three identical ochre (UAA)-suppressor serine tRNA genes (SUP16, 17, or 19). Two strains have no identified suppressor mutation. We infer that the cold-sensitive growth defect of Rpb3-K9E is enhanced by the ade2-1 substitution.