WIP1 dephosphorylation of p27Kip1 Serine 140 destabilizes p27Kip1 and reverses anti-proliferative effects of ATM phosphorylation

Abstract

The phosphoinositide-3-kinase like kinases (PIKK) such as ATM and ATR play a key role in initiating the cellular DNA damage response (DDR). One key ATM target is the cyclin-dependent kinase inhibitor p27Kip1 that promotes G1 arrest. ATM activates p27Kip1-induced arrest in part through phosphorylation of p27Kip1 at Serine 140. Here we show that this site is dephosphorylated by the type 2C serine/threonine phosphatase, WIP1 (Wildtype p53-Induced Phosphatase-1), encoded by the PPM1D gene. WIP1 has been shown to dephosphorylate numerous ATM target sites in DDR proteins, and its overexpression and/or mutation has often been associated with oncogenesis. We demonstrate that wildtype, but not phosphatase-dead WIP1, efficiently dephosphorylates p27Kip1 Ser140 both in vitro and in cells and that this dephosphorylation is sensitive to the WIP1-specific inhibitor GSK 2830371. Increased expression of wildtype WIP1 reduces stability of p27Kip1 while increased expression of similar amounts of phosphatase-dead WIP1 has no effect on p27Kip1 protein stability. Overexpression of wildtype p27Kip1 reduces cell proliferation and colony forming capability relative to the S140A (constitutively non-phosphorylated) form of p27. Thus, WIP1 plays a significant role in homeostatic modulation of p27Kip1 activity following activation by ATM.