ATM associates with and phosphorylates p53: mapping the region of interaction

Abstract

The human genetic disorder ataxia-telangiectasia (AT) is characterized by immunodeficiency, progressive cerebellar ataxia, radiosensitivity, cell cycle checkpoint defects and cancer predisposition. The gene mutated in this syndrome, ATM (for AT mutated), encodes a protein containing a phosphatidyl-inositol 3-kinase (PI-3 kinase)-like domain. ATM also contains a proline-rich region and a leucine zipper, both of which implicate this protein in signal transduction. The proline-rich region has been shown to bind to the SH3 domain of c-Abl, which facilitates its phosphorylation and activation by ATM. Previous results have demonstrated that AT cells are defective in the G1/S checkpoint activated after radiation damage and that this defect is attributable to a defective p53 signal transduction pathway. We report here direct interaction between ATM and p53 involving two regions in ATM, one at the amino terminus and the other at the carboxy terminus, corresponding to the PI-3 kinase domain. Recombinant ATM protein phosphorylates p53 on serine 15 near the N terminus. Furthermore, ectopic expression of ATM in AT cells restores normal ionizing radiation (IR)-induced phosphorylation of p53, whereas expression of ATM antisense RNA in control cells abrogates the rapid IR-induced phosphorylation of p53 on serine 15. These results demonstrate that ATM can bind p53 directly and is responsible for its serine 15 phosphorylation, thereby contributing to the activation and stabilization of p53 during the IR-induced DNA damage response.

Low frequency of CHEK2 1100delC allele in Australian multiple-case breast cancer families: functional analysis in heterozygous individuals

“…To ensure that the reduction in Thr68 phosphorylation of CHK2 was not due to an ATM defect upstream of CHK2, but was only a result of the decreased levels of CHK2, we investigated ATM phosphorylation of p53 at Ser15 as a measure of in vivo ATM activity (Banin et al, 1998;Canman et al, 1998;Khanna et al, 1998). Phosphorylation of p53 on Ser15 was similar between the control and heterozygous 1100delC LCLs, as was the level of ATM in these cells ( Figure 3B).…”

ATM: A mediator of multiple responses to genotoxic stress

“…ATM binds p53 in vivo, and directly phosphorylates it on Ser-15 in vitro (Banin et al, 1998;Khanna et al, 1998). The rapid de novo phosphorylation of Ser-15 in response to IR or DSBs, but not to UV, is ATM-dependent (Siliciano et al, 1997;Nakagawa et al, 1999).…”

Mechanisms of switching on p53: a role for covalent modification?

“…ATM had long been suspected of being an upstream component of the p53 pathway on the basis that the induction of p53 in cells from AT patients showed a signi®cant delay in the response to IR (Kastan et al, 1992;Khanna and Lavin, 1993). Recently ATM was shown to be a bona ®de protein kinase which can phosphorylate p53 at serine 15 in vitro (Banin et al, 1998;Canman et al, 1998;Khanna et al, 1998). Moreover, several lines of evidence supported the idea that ATM was indeed immediately upstream of p53 in the pathway responding to IR and radiomimetic drugs: (1) serine 15 phosphorylation correlates with stimulation of ATM protein kinase activity in cultured cells (Banin et al, 1998;Canman et al, 1998); (2) ATM physically complexes with p53 in cells from normal individuals but this is defective in AT cells (Khanna et al, 1998;Watters et al, 1997); (3) serine 15 phosphorylation is severely delayed in AT cells (Canman et al, 1998); and (4) ectopic expression of ATM in AT cells restores IR-induced phosphorylation of p53 at serine 15 whereas expression of antisense RNA in normal cells abrogates serine 15 phosphorylation (Khanna et al, 1998).…”