Transcriptional activation of p53 target genes, due to DNA damage, causes either apoptosis or survival by cell cycle arrest and DNA repair. However, the regulators of the choice between cell death and survival signaling have not been completely elucidated. Here, we report that human adenocarcinoma cells (MCF-7) survive UV-induced DNA damage by heat shock protein 27 (Hsp27)-assisted Akt/p21 phosphorylation/translocation. Protein levels of the p53 target genes, such as p21, Bcl-2, p38MAPK, and Akt, showed a positive correlation to Hsp27 level during 48 hours postirradiation, whereas p53 expression increased initially but started decreasing after 12 hours. Hsp27 prevented the G 1 -S phase cell cycle arrest, observed after 8 hours of post-UV irradiation, and PARP-1 cleavage was inhibited. Conversely, silencing Hsp27 enhanced G 1 -S arrest and cell death. Moreover, use of either Hsp27 or Akt small interference RNA reduced p21 phosphorylation and enhanced its retention in nuclei even after 48 hours postirradiation, resulting in enhanced cell death. Our results showed that Hsp27 expression and its direct chaperoning interaction increases Akt stability, and p21 phosphorylation and nuclear-to-cytoplasm translocation, both essential effects for the survival of UV-induced DNA-damaged cells. We conclude that the role of Hsp27 in cancer is not only for enhanced p53 proteolysis per se, rather it is also a critical determinant in p21 phosphorylation and translocation. Mol Cancer Res; 8(10); 1399-412. ©2010 AACR.

Small heat shock proteins (sHsps) protect the heart from chemotherapeutics-induced heart failure, by inhibiting p53-dependant apoptosis. However, mechanism of such protection has not been elucidated yet. Here we test a hypothesis that serine phosphorylation of sHsps is essential to inhibit the Doxorubicin-induced p53-dependent apoptotic pathway. Three transgenic mice (TG) lines with cardiomyocyte specific overexpression of human heat shock protein 27 (hHsp27), namely, wild type (MHC-hHsp27), S82A single mutant (MHC-mut-hHsp27(S82A) and tri-mutant (MHC-mut-hHsp27(S15A/S78A/S82A)) were generated. TG mice were treated with Dox (6mg/kg body weight; once in a week; 4 weeks) along with age-matched non-transgenic (Non-TG) controls. The Dox-treated MHC-hHsp27 mice showed improved survival and cardiac function (both MRI and echocardiography), in terms of contractility (%EF) and left ventricular inner diameter (LVID), compared to the Dox-treated Non-TG mice. However, both MHC-mut-hHsp27(S82A) and MHC-mut-hHsp27(S82A/S15A/S76A) mutants overexpressing TG mice did not show such a cardioprotection. Furthermore, transactivation of p53 was found to be attenuated only in Dox-treated MHC-hHsp27 mice-derived cardiomyocytes in vitro, as low p53 was detected in the nuclei, not in mutant hHsp27 overexpressing cardiomyocytes. Similarly, only in MHC-hHsp27 overexpressing cardiomyocytes, low Bax, higher mTOR phosphorylation and low apoptotic PARP-1 cleavage (89kDa fragment) were detected. Pharmacological inhibition of p53 was more effective in mutant-TG mice, compared to MHC-hHsp27 mice. We conclude that phosphorylation of overexpressed Hsp27 at S82 and its association with p53 is essential for the overall cardioprotective effect of Hsp27 against Dox-induced dilated cardiomyopathy. Only phosphorylated Hsp27 protect the heart by inhibiting p53 transactivation.