Ime2, a Meiosis-Specific Kinase in Yeast, Is Required for Destabilization of Its Transcriptional Activator, Ime1

Abstract

In the budding yeast Saccharomyces cerevisiae, entry into meiosis and its successful completion depend on two positive regulators, Ime1 and Ime2. Ime1 is a transcriptional activator that is required for transcription of IME2, a serine/threonine protein kinase. We show that in vivo Ime2 associates with Ime1, that in vitro Ime2 phosphorylates Ime1, and that in living cells the stability of Ime1 depends on Ime2. Diploid cells with IME2 deleted show an increase in the level of Ime1, whereas haploid cells overexpressing IME2 show a decrease in the stability of Ime1. Furthermore, the level of Ime1 depends on the kinase activity of Ime2. Using a mutation in one of the ATPase subunits of the proteasome, RPT2, we demonstrate that Ime1, amino acids 270 to 360, is degraded by the 26S proteasome. We also show that Ime2 itself is an extremely unstable protein whose expression in vegetative cultures is toxic. We propose that a negative-feedback loop ensures that the activity of Ime1 will be restricted to a narrow window.Successful progression and completion of the mitotic cell cycle depends on transcriptional and proteolytic regulation. These two processes determine the availability of cyclins and cyclin-dependent kinase (CDK) inhibitors that govern the sequential activation of CDKs (18,28,31,35). Initiation and progression through the meiotic cycle should also be subjected to transcriptional and proteolytic regulation. Indeed, in budding yeast a transcriptional cascade governs initiation and progression through the meiotic cell cycle (4). Yet there is no direct evidence concerning proteolysis of either positive or negative meiotic regulators. This report focuses on the regulated degradation of one of the two positive regulators of meiosis in Saccharomyces cerevisiae, Ime1, by the other, Ime2.IME1 encodes a transcriptional activator (30, 47) that is necessary for the transcription of meiosis-specific genes (48). Ime1 is tethered to promoters of early meiosis-specific genes, such as IME2, by a specific DNA-binding protein, Ume6 (39). Diploid cells with deletions of IME1 arrest at G 1 prior to the initiation of premeiotic DNA replication (22). The transcription of IME1 is regulated by nutrients. In vegetative cultures with glucose as the sole carbon source, IME1 is silent, but in the presence of acetate, low levels of IME1 mRNA are observed (22). Under meiotic conditions, i.e., nitrogen depletion and the presence of a nonfermentable carbon source such as acetate, transcription of IME1 is induced transiently in MATa/ MAT␣ diploids (22). It is not known whether this transient transcription reflects transient availability of the Ime1 protein.In addition, the IME1 promoter is subject to positive autoregulation (40,43,44), as well as negative-feedback regulation by both Ime1 and Ime2 (43,48,49).Another important regulator of meiosis and sporulation is the serine/threonine protein kinase Ime2 (12,24,34,48,49). Diploid cells with deletions of IME2 show a 5-to 12-h delay in the transcription of early meiosis-specific genes, a reduction...

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“…This observation is consistent with previous studies showing that IME2 is regulated through negative feedback (Mitchell et al 1990). Negative feedback occurs in part through cell autonomous mechanisms; for example, Ime2p targets Ime1p for degradation, thus repressing transcription of its own gene (Guttmann-Raviv et al 2002). In addition, negative feedback in the context of colonies could proceed through cell nonautonomous mechanisms; for example, signals secreted by one cell could regulate sporulation in neighboring cells.…”

A nutrient dependant switch explains mutually exclusive existence of meiosis and mitosis initiation in budding yeast

“…These results are consistent with the experimental results; Ime1 is in undetectable levels, and Ime2 is absent in the nucleus during vegetative conditions (Sagee et al, 1998). According to experiments, expression of Ime2 in vegetative cultures is toxic (Guttmann-Raviv et al, 2002). The Rpd3/Sin3 complex on the promoters shows high value in our model output.…”

Arg-Pro-X-Ser/Thr Is a Consensus Phosphoacceptor Sequence for the Meiosis-Specific Ime2 Protein Kinase in Saccharomyces cerevisiae

“…Ime2 also appears to promote early gene transcription in an Ime1-independent fashion. Later in the program Ime2 is also required for Ime1 destabilization and the downregulation of early meiotic genes (8). Ime2 can phosphorylate Ime1 in vitro, although the phosphoacceptor site(s) in Ime1 have not been identified, and therefore the mechanism by which Ime2 down-regulates Ime1 is yet to be determined.…”