Abstract
During M phase, Endosulfine (Endos) family proteins are phosphorylated by Greatwall kinase (Gwl), and the resultant pEndos inhibits the phosphatase PP2A-B55, which would otherwise prematurely reverse many CDK-driven phosphorylations. We show here that PP2A-B55 is the enzyme responsible for dephosphorylating pEndos during M phase exit. The kinetic parameters for PP2A-B55's action on pEndos are orders of magnitude lower than those for CDK-phosphorylated substrates, suggesting a simple model for PP2A-B55 regulation that we call inhibition by unfair competition. As the name suggests, during M phase PP2A-B55's attention is diverted to pEndos, which binds much more avidly and is dephosphorylated more slowly than other substrates. When Gwl is inactivated during the M phase-to-interphase transition, the dynamic balance changes: pEndos dephosphorylated by PP2A-B55 cannot be replaced, so the phosphatase can refocus its attention on CDK-phosphorylated substrates. This mechanism explains simultaneously how PP2A-B55 and Gwl together regulate pEndos, and how pEndos controls PP2A-B55. DOI: http://dx.doi.org/10.7554/eLife.01695.001.
Highlights
Entry of cells into M phase of mitosis or meiosis requires the phosphorylation of thousands of sites on hundreds of proteins (Dephoure et al, 2008; Lindqvist et al, 2009; Dulla et al, 2010; Olsen et al, 2010)
Three independent lines of evidence indicate that the major activity contributing to the dephosphorylation of Greatwall kinase (Gwl)-phosphorylated Endos is a phosphatase that includes the three subunits of the PP2A-B55 heterotrimer
(1) Inhibitor specificities: all of the anti-Endos activity is sensitive to okadaic acid and calyculin A (Figure 2B, Figure 2—figure supplement 1), and the majority is highly sensitive to fostriecin (Figure 2C, Figure 2—figure supplement 2), suggesting that the catalytic subunit of the anti-Endos phosphatase is PP2A or its less abundant relatives PP4 or PP6
Summary
Entry of cells into M phase of mitosis or meiosis requires the phosphorylation of thousands of sites on hundreds of proteins (Dephoure et al, 2008; Lindqvist et al, 2009; Dulla et al, 2010; Olsen et al, 2010). Many of these sites are substrates of cyclin-dependent kinases (CDKs), most notably MPF (M phase-promoting factor; CDK1-Cyclin B). Gwl-phosphorylated Endos (pEndos) binds to and inactivates PP2A-B55, protecting a major class of CDK-governed
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