Identification and characterization of rapidly accumulating sch9Δ suppressor mutations in Saccharomyces cerevisiae

Abstract

Abstract Nutrient sensing is important for cell growth, aging, and longevity. In Saccharomyces cerevisiae, Sch9, an AGC-family protein kinase, is a major nutrient sensing kinase homologous to mammalian Akt and S6 kinase. Sch9 integrates environmental cues with cell growth by functioning downstream of TORC1 and in parallel with the Ras/PKA pathway. Mutations in SCH9 lead to reduced cell growth in dextrose medium; however, reports on the ability of sch9Δ mutants to utilize non-fermentable carbon sources are inconsistent. Here, we show that sch9Δ mutant strains cannot grow on non-fermentable carbon sources and rapidly accumulate suppressor mutations, which reverse growth defects of sch9Δ mutants. sch9Δ induces gene expression of three transcription factors required for utilization of non-fermentable carbon sources, Cat8, Adr1, and Hap4, while sch9Δ suppressor mutations, termed sns1 and sns2, strongly decrease the gene expression of those transcription factors. Despite the genetic suppression interactions, both sch9Δ and sns1 (or sns2) homozygous mutants have severe defects in meiosis. By screening mutants defective in sporulation, we identified additional sch9Δ suppressor mutants with mutations in GPB1, GPB2, and MCK1. Using library complementation and genetic analysis, we identified SNS1 and SNS2 to be IRA2 and IRA1, respectively. Furthermore, we discovered that lifespan extension in sch9Δ mutants is dependent on IRA2 and that PKA inactivation greatly increases basal expression of CAT8, ADR1, and HAP4. Our results demonstrate that sch9Δ leads to complete loss of growth on non-fermentable carbon sources and mutations in MCK1 or genes encoding negative regulators of the Ras/PKA pathway reverse sch9Δ mutant phenotypes.