Regulation of Non-Canonical MAP Kinase Signaling
Mitogen activated protein kinases (MAPKs) are signaling enzymes that control diverse processes in eukaryotic cells including proliferation, differentiation, and development. Many MAPKs are activated through well-studied canonical signaling pathways in which a MAPK kinase phosphorylates a T-X-Y motif in the activation loop of the MAPK. However, MAPKs can also be activated by non-canonical mechanisms. Smk1 is a meiosis-specific MAPK in the yeast S. cerevisiae that controls spore wall morphogenesis. The Smk1 pathway provides a model system to interrogate mechanisms of non-canonical MAPK signaling in the context of a developmental program. Previous genetic studies identified CAK1, AMA1 , IME2 and SSP2 as genes that regulate Smk1. Cak1 is a cyclin-dependent kinase (CDK)-activating kinase. Ama1 is a meiosis-specific targeting factor of the APC/C E3 ubiquitin ligase that controls exit from meiosis II (MII). Ime2 is a meiosis-specific CDK-like kinase. Ssp2 is a meiosis-specific protein required for spore morphogenesis, whose biochemical function has not previously been described. In this thesis we provide evidence for a novel spatiotemporal regulatory mechanism in which Cak1 phosphorylates Smk1 on its activation-loop T (T207) as the meiotic divisions are taking place. Smk1 autophosphorylates its activation-loop Y (Y209) as Ssp2 binds Smk1 at membranes that surround haploid products (prospore membranes; PSMs) during exit from meiosis II. Ssp2/Smk1 complex formation is regulated through multiple mechanisms, preventing premature MAPK activity. First, although both SMK1 and SSP2 are transcriptionally induced as cells enter meiosis I (MI), SMK1 mRNA is translated almost immediately, while SSP2 mRNA is translationally repressed until Ime2 triggers its derepression at MII. Next, Ssp2 localizes to PSMs, but inhibitory phosphates prevent it from binding Smk1. These phosphates are removed during meiotic exit, as PSMs close around each meiotic product in a step that requires Ama1. Finally, dephosphorylated Ssp2 binds Smk1, inducing autophosphorylation, thus coupling meiotic exit to Smk1 activation. We also show that RRM-like domains in the C-terminus of Ssp2 bind Smk1. Taken together, this work has defined novel mechanisms for spatiotemporally regulating the Smk1 MAPK during meiotic development.
Omerza, Gregory, "Regulation of Non-Canonical MAP Kinase Signaling" (2016). ETD Collection for Thomas Jefferson University. AAI10172490.