Large-scale screening of yeast mutants for sensitivity to the IMP dehydrogenase inhibitor 6-azauracil.

Mutations in several genes encoding components of the RNA polymerase II elongation machinery render S. cerevisiae cells sensitive to the drug 6-azauracil (6AU), an inhibitor of IMP dehydrogenase and orotidylate decarboxylase. It is thought that a reduction in nucleotide levels following drug treatment causes transcriptional elongation to be more dependent on a fully functional RNA polymerase.

Functional distinctions between IMP dehydrogenase genes in providing mycophenolate resistance and guanine prototrophy to yeast.

IMP dehydrogenase (IMPDH) catalyzes the rate-limiting step in the de novo synthesis of GTP. Yeast with mutations in the transcription elongation machinery are sensitive to inhibitors of this enzyme such as 6-azauracil and mycophenolic acid, at least partly because of their inability to transcriptionally induce IMPDH. To understand the molecular basis of this drug-sensitive phenotype, we have dissected the expression and function of a four-gene family in yeast called IMD1 through IMD4.

Use of an in vivo reporter assay to test for transcriptional and translational fidelity in yeast.

Eukaryotic RNA polymerase II and Escherichia coli RNA polymerase possess an intrinsic ribonuclease activity that is stimulated by the polymerase-binding proteins SII and GreB, respectively. This factor-activated hydrolysis of nascent RNA has been postulated to be involved in transcription elongation as well as removal of incorrect bases misincorporated into RNA. Little is known about the frequency of misincorporation by RNA polymerases in vivo or about the mechanisms involved in improving RNA polymerase accuracy.