Multiple regulatory levels influence cell integrity control by PKC ortholog Pck2 in fission yeast

Abstract

Fission yeast PKC ortholog Pck2 controls cell wall synthesis and is a major upstream activator of the cell integrity pathway (CIP) and its core component, MAP kinase Pmk1, in response to environmental stimuli. We show that in vivo phosphorylation of Pck2 at the conserved T842 activation loop during growth and in response to different stresses is mediated by the PDK ortholog Ksg1 and an autophosphorylation mechanism. However, T842 phosphorylation is not essential for Pmk1 activation, and putative phosphorylation at T846 might play an additional role for Pck2 catalytic activation and downstream signaling. These events together with turn motif autophosphorylation at T984 and binding to small GTPases Rho1 and/or Rho2 stabilize and render Pck2 competent to exert its biological functions. Remarkably, the TORC2 complex does not participate in catalytic activation of Pck2, but instead contributes to de novo Pck2 synthesis which is essential to activate the CIP in response to cell wall damage or glucose exhaustion. These results unveil a novel mechanism whereby TOR regulates PKC function at a translational level and add a new regulatory layer to MAPK signaling cascades.