Determinants for Activation of the Atypical AGC Kinase Greatwall during M Phase Entry

Abstract

The atypical AGC kinase Greatwall (Gwl) mediates a pathway that prevents the precocious removal of phosphorylations added to target proteins by M phase-promoting factor (MPF); Gwl is thus essential for M phase entry and maintenance. Gwl itself is activated by M phase-specific phosphorylations that are investigated here. Many phosphorylations are nonessential, being located within a long nonconserved region, any part of which can be deleted without effect. Using mass spectrometry and mutagenesis, we have identified 3 phosphorylation sites (phosphosites) critical to Gwl activation (pT193, pT206, and pS883 in Xenopus laevis) located in evolutionarily conserved domains that differentiate Gwl from related kinases. We propose a model in which the initiating event for Gwl activation is phosphorylation by MPF of the proline-directed sites T193 and T206 in the presumptive activation loop. After this priming step, Gwl can intramolecularly phosphorylate its C-terminal tail at pS883; this site probably plays a role similar to that of the tail/Z motif of other AGC kinases. These events largely (but not completely) explain the full activation of Gwl at M phase.

This article is related to:

Stable Government of Mitosis by Greatwall: the Emperor's Best Servant

ACKNOWLEDGMENTS

James Maller (University of Colorado School of Medicine, Aurora, CO) has been remarkably generous with reagents, advice, laboratory space, and comments on the manuscript. We also thank J. Kuang (M. D. Anderson Cancer Center, Houston, TX) for the gift of MPM-2 antibody.

This study was supported by NIH grant GM048430 to M.L.G.