Regulation of ureaamidolyase synthesis in Saccharomyces cerevisiae, RNA analysis, and cloning of the positive regulatory gene DURM

Summary

In S. cerevisiae, the synthesis of ureaamidolyase is subject to at least two different forms of regulation: nitrogen catabolite repression and induction by allophanate. Two positive regulatory genes DURM and DURL are involved in the induction process. We have measured the levels of mRNA homologous to the DUR2,1 gene in conditions of ureaamidolyase induction and in regulatory mutants. The amounts of DUR2,1 enzyme and messengers are well coordinated; moreover, the half life of DUR2,1 messengers is identical in the presence or absence of inducer.

These data suggest that the ureaamidolyase production is probably controlled at the level of transcription. From a pool of hybrid plasmids carrying Sau3A fragments representing the entire yeast genome, a 13 kb DNA fragment containing the regulatory gene DURM was cloned by complementation of a durM mutation which prevents the growth on allantoin as sole nitrogen source. Cells containing the cloned DNA recover the inducibility of ureaamidolyase by allophanate. Four RNA transcripts have homology to this 13 kb DNA fragment but the study of subcloned restriction endonuclease fragments allowed us to map the DURM regulatory gene within a 4 kilobase pair region. This fragment encodes a 1 kb transcript. The level of this RNA is the same in induced and non-induced cells.