Protein phosphatase 2A (PP2A) is a very important family of holoenzyme complexes that functions within a diversity of signaling pathways inside the cell (9
). PP2A consists of either a core complex containing a catalytic (C) subunit and scaffolding (A) subunit (29
) or a trimer containing the AC core with one of many possible regulatory (B) subunits bound to it (30
). The known B subunits have been divided into four gene families based on sequence homology: the B (B55 or PR55), B′ (B56 or PR61), B" (PR48/59/72/130), and B
(PR93/110) families (25
). Each of these many B subunits can combine with the PP2A core to form complexes with distinct activities and substrate specificities. As such, PP2A is able to perform various functions in multiple regulatory pathways, depending on which B subunit is bound.
In the past, PP2A was thought to have primarily dull housekeeping functions inside the cell. Recent studies, however, suggest that PP2A may have more-active regulatory roles and may actually function as a tumor suppressor under certain conditions. It is believed that a small subset of B subunits is most likely responsible for promoting this function of PP2A. In support of this view, at least two B56 subunit family members have been implicated in conferring tumor-suppressive functions on the holoenzyme. The B56 family consists of five different genes, α (PPP2R5A), β (PPP2R5B), γ (PPP2R5C), δ (PPP2R5D), and
). In addition, the B56γ gene encodes four differentially spliced forms, PP2A B56γ1, -γ2, -γ3 and -γ4 (17
). B56δ-specific PP2A was shown to function in a mitotic checkpoint in Xenopus laevis
) and B56γ3-specific PP2A in blocking the proliferation of lung cancer cell lines (3
). Importantly, evidence from our laboratory indicates that B56γ-PP2A participates in the activation of the tumor suppressor protein p53 after DNA damage (13
p53 is a highly regulated tumor suppressor protein that is very important in cancer suppression. In response to genotoxic stress, p53 is activated through a series of posttranslational modifications (2
). Once activated, it acts as a transcription factor, eliciting the transcription of genes that induce cell cycle arrest or programmed cell death (23
). Our studies have shown that, under cell growth conditions, p53 is phosphorylated at Thr55 by TAF1, which helps to keep the protein inactive, and upon genotoxic stress, B56γ-PP2A complexes dephosphorylate p53 at this residue, leading to p53 activation, p21 expression, and G1
cell cycle arrest (12
). Interestingly, in the course of our studies, we observed an enhanced interaction between B56γ and p53 upon DNA damage; however, its significance in p53 activation and in PP2A tumor suppressor function remains unknown.
In the present study, we show that the p53-B56γ interaction is required for p53 and B56γ-PP2A cooperative tumor suppression. Mechanistically, we show that the kinase activity of ATM is required for Thr55 dephosphorylation in response to DNA damage. ATM is an important kinase involved in cellular responses to DNA double-strand breaks. Once activated, ATM directly phosphorylates p53 at Ser15 and promotes Ser20 phosphorylation indirectly by activating Chk2 kinase. We show that Ser15, but not Ser20, mutant p53 is unable to interact with B56γ and significantly reduces the ability of B56γ3 to inhibit cell proliferation and transformation, suggesting that Ser15 phosphorylation primes p53 for the p53-B56γ interaction and Thr55 dephosphorylation by PP2A. Taken together, our results demonstrate the importance of the Ser15-mediated p53-B56γ interaction in the activation of p53 by B56γ-PP2A and in PP2A tumor suppressor function. In addition, our results also provide a functional link between ATM and PP2A tumor suppressor activity in response to DNA damage.