High-fidelity maintenance of genomic integrity in eukaryotes is ensured by cell 
cycle checkpoints and DNA repair. The checkpoint kinase, Chk2, has been 
implicated in both of these responses. In response to DNA damage, Chk2 is 
initially phosphorylated at Thr-68, which leads to its full activation. The 
fully activated Chk2 then phosphorylates downstream substrates of cell cycle 
control. However, the mechanism of inactivation of Chk2 is still unknown. 
Protein phosphatase type 2A (PP2A) plays an essential role in cell cycle 
regulation and induction of G2 arrest by a mechanism of 
phosphorylation/dephosphorylation with a variety of protein kinases. Data from 
our investigation provide evidence that, in response to cisplatin exposure, PP2A 
associates with Chk2 as a complex in cells and functions as a negative regulator 
of Chk2 activation by dephosphorylating p-Chk2. Results from immunostaining and 
coimmunoprecipitation demonstrate that Chk2 and PP2A can colocalize in cells, 
and the holoenzyme of PP2A (subunits A, B and C) coimmunoprecipitates with 
p-Chk2. Further, inhibition of PP2A by okadaic acid, an inhibitor of PP2A, and 
by small interfering RNA (siRNA) to PP2A results in enhanced Chk2 
phosphorylation, implicating a direct enzyme-substrate relationship. An in vitro 
PP2A dephosphorylation assay shows that PP2A dephosphorylates p-Chk2 in a 
cell-free system. These findings suggest that the protein serine/threonine 
kinase, Chk2, is activated after cisplatin exposure and negatively regulated by 
a tightly associated protein serine/threonine phosphatase, PP2A.