A mutation in Saccharomyces cerevisiae adenylate cyclase, Cyr1(K1876M), specifically affects glucose- and acidification-induced cAMP signalling and not the basal cAMP level

In the yeast Saccharomyces cerevisiae, the addition of glucose to derepress ed cells and intracellular acidification trigger a rapid increase in the cA MP level within 1 min. We have identified a mutation in the genetic backgro und of several related 'wild-type' laboratory yeast strains (e.g. ENY.cat80 -7A, CEN.PK2-1C) that largely prevents both cAMP responses, and we have cal led it lcr1 (for lack of cAMP responses). Subsequent analysis showed that l cr1 was allelic to CYR1/CDC35, encoding adenylate cyclase, and that it cont ained an A to T substitution at position 5627. This corresponds to a K1876M substitution near the end of the catalytic domain in adenylate cyclase. In troduction of the A5627T mutation into the CYR1 gene of a W303-1A wild-type strain largely eliminated glucose- and acidification-induced cAMP signalli ng and also the transient cAMP increase that occurs in the lag phase of gro wth. Hence, lysine(1876) of adenylate cyclase is essential for cAMP respons es in vivo. Lysine(1876) is conserved in Schizosaccharomyces pombe adenylat e cyclase. Mn2+-dependent adenylate cyclase activity in isolated plasma mem branes of the cyr1(met1876) (lcr1) strain was similar to that in the isogen ic wild-type strain, but GTP/Mg2+-dependent activity was strongly reduced, consistent with the absence of signalling through adenylate cyclase in vivo . Glucose-induced activation of trehalase was reduced and mobilization of t rehalose and glycogen and loss of stress resistance were delayed in the cyr 1(met1876) (lcr1) mutant. During exponential growth on glucose, there was l ittle effect on these protein kinase A (PKA) targets, indicating that the i mportance of glucose-induced cAMP signalling is restricted to the transitio n from gluconeogenic/respiratory to fermentative growth. Inhibition of grow th by weak acids was reduced, consistent with prevention of the intracellul ar acidification effect on cAMP by the cyr1(met1876) (lcr1) mutation. The m utation partially suppressed the effect of RAS2(val19) and GPA2(val132) on several PKA targets. These results demonstrate the usefulness of the cyr1(m et1876) (lcr1) mutation for epistasis studies on the signalling function of the cAMP pathway.