Glucose Inhibits Meiotic DNA Replication through SCF^Grr1p-Dependent Destruction of Ime2p Kinase

Abstract

In the budding yeast Saccharomyces cerevisiae, the cell division cycle and sporulation are mutually exclusive cell fates; glucose, which stimulates the cell division cycle, is a potent inhibitor of sporulation. Addition of moderate concentrations of glucose (0.5%) to sporulation medium did not inhibit transcription of two key activators of sporulation, IME1 and IME2, but did increase levels of Sic1p, a cyclin-dependent kinase inhibitor, resulting in a block to meiotic DNA replication. The effects of glucose on Sic1p levels and DNA replication required Grr1p, a component of the SCF Grr1p ubiquitin ligase. Sic1p is negatively regulated by Ime2p kinase, and several observations indicate that glucose inhibits meiotic DNA replication through SCF Grr1p -mediated destruction of this kinase. First, Ime2p was destabilized in the presence of glucose, and this turnover required Grr1p, a second component of SCF Grr1p , Cdc53p, and an SCF Grr1p -associated E2 enzyme, Cdc34p. Second, Ime2p-ubiquitin conjugates were detected under conditions of rapid Ime2p turnover, and conjugation of Ime2p to ubiquitin required GRR1. Third, a mutant form of Ime2p (Ime2 ⌬PEST ), in which a putative Grr1p-interacting sequence was deleted, was more stable than wild-type Ime2p. Finally, expression of the IME2⌬PEST allele bypassed the block to meiotic DNA replication caused by 0.5% glucose. In addition, Grr1p is required for later events in sporulation independently of its role in Ime2p turnover.

The Ras/cAMP Pathway and the CDK-Like Kinase Ime2 Regulate the MAPK Smk1 and Spore Morphogenesis in Saccharomyces cerevisiae

“…Indeed, glucose can increase Ime2 degradation in sporulating cells through its C terminus (Purnapatre et al 2005;Gray et al 2008) and the DC241 deletion stabilizes Ime2 in mitotic cells (Sari et al 2008). Gpa2 is a Ga-protein that transduces glucose signals from the Gpr1 receptor to adenylate cyclase and PKA (reviewed in Thevelein et al 2008).…”

“…IME2-DC241 exacerbates the smk1-2 phenotype: Ime2 consists of an amino-terminal phosphotransferase domain and a carboxy-terminal domain. Glucose has been shown to negatively regulate Ime2 via the carboxyterminal domain (Donzeau and Bandlow 1999;Purnapatre et al 2005). It has also been shown that IME2-DC241, which truncates the carboxy-terminal domain, can function in a dominant manner to promote skipped spore packaging and thus increase the frequency of asci containing dyads and triads (Sari et al 2008).…”

“…Shortly after Ime2 is produced it is activated through T-loop phosphorylation by the Cak1 CDK activating kinase (Schindler et al 2003), and its activity is further regulated by phosphorylation of the C-terminal domain as cells progress through meiotic development (Schindler and Winter 2006). In meiotic cells, motifs in the C terminus of Ime2 regulate protein stability in response to low levels of glucose (Purnapatre et al 2005;Gray et al 2008). Truncation of the carboxy-terminal third of Ime2 is hyperactivating (Kominami et al 1993;Sia and Mitchell 1995) and generates a stabilized protein that causes cells to form asci with reduced numbers of spores (dyads and triads) (Sari et al 2008).…”

Sporulation in the Budding Yeast Saccharomyces cerevisiae

“…In addition, Ime2 is also subject to direct regulation by glucose (Purnapatre et al 2005;Gray et al 2008). In the presence of glucose, Ime2 is rapidly degraded via the SCF ubiquitin ligase Grr1 and degradation signals in the Ime2 C terminus (Purnapatre et al 2005;Sari et al 2008). Thus, reintroduction of glucose early in sporulation can block further progression down this developmental pathway, at least in part, by inactivating Ime2.…”

“…Its initial induction requires activation by Ime1/ Ume6 and so is affected by nutritional controls acting on Ime1 (Pak and Segall 2002a). In addition, Ime2 is also subject to direct regulation by glucose (Purnapatre et al 2005;Gray et al 2008). In the presence of glucose, Ime2 is rapidly degraded via the SCF ubiquitin ligase Grr1 and degradation signals in the Ime2 C terminus (Purnapatre et al 2005;Sari et al 2008).…”