HSP27 regulates p53 transcriptional activity in doxorubicin-treated fibroblasts and cardiac H9c2 cells: p21 upregulation and G2/M phase cell cycle arrest

C. D. Venkatakrishnan, Kathy Dunsmore, Hector Wong, Sashwathi Roy, Chandan K. Sen, Altaf Wani, Jay L. Zweier, Govindasamy Ilangovan

Research output: Contribution to journalArticle

Abstract

Treatment of cancer patients with anthracyclin-based chemotherapeutic drugs induces congestive heart failure by a mechanism involving p53. However, it is not known how p53 aggravates doxorubicin (Dox)-induced toxicity in the heart. On the basis of in vitro acute toxicity assay using heat shock factor-1 (HSF-1) wild-type (HSF-1+/+) and HSF-1-knockout (HSF-1-/-) mouse embryonic fibroblasts and neonatal rat cardiomyocyte-derived H9c2 cells, we demonstrate a novel mechanism whereby heat shock protein 27 (HSP27) regulates transcriptional activity of p53 in Dox-treated cells. Inhibition of p53 by pifithrin-α (PFT-α) provided different levels of protection from Dox that correlate with HSP27 levels in these cells. In HSF-1+/+ cells, PFT-α attenuated Dox-induced toxicity. However, in HSF-1-/- cells (which express a very low level of HSP27 compared with HSF-1+/+ cells), there was no such attenuation, indicating an important role of HSP27 in p53-dependent cell death. On the other hand, immunoprecipitation of p53 was found to coimmunoprecipitate HSP27 and vice versa (confirmed by Western blotting and matrix-assisted laser desorption/ionization time of flight), demonstrating HSP27 binding to p53 in Dox-treated cells. Moreover, upregulation of p21 was observed in HSF-1+/+ and H9c2 cells, indicating that HSP27 binding transactivates p53 and enhances transcription of p21 in response to Dox treatment. Further analysis with flow cytometry showed that increased expression of p21 results in G2/M phase cell cycle arrest in Dox-treated cells. Overall, HSP27 binding to p53 attenuated the cellular toxicity by upregulating p21 and prevented cell death.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Heart and Circulatory Physiology
Volume294
Issue number4
DOIs
StatePublished - Apr 2008
Externally publishedYes

Fingerprint

HSP27 Heat-Shock Proteins
M Phase Cell Cycle Checkpoints
G2 Phase
Doxorubicin
Up-Regulation
Fibroblasts
Shock
Hot Temperature
Protein Binding
Cell Death
Immunoprecipitation
Cardiac Myocytes
Flow Cytometry
Lasers
Heart Failure
Western Blotting

Keywords

  • Cell survival
  • DNA repair
  • Oxidative stress
  • Redox signaling

ASJC Scopus subject areas

  • Physiology

Cite this

HSP27 regulates p53 transcriptional activity in doxorubicin-treated fibroblasts and cardiac H9c2 cells : p21 upregulation and G2/M phase cell cycle arrest. / Venkatakrishnan, C. D.; Dunsmore, Kathy; Wong, Hector; Roy, Sashwathi; Sen, Chandan K.; Wani, Altaf; Zweier, Jay L.; Ilangovan, Govindasamy.

In: American Journal of Physiology - Heart and Circulatory Physiology, Vol. 294, No. 4, 04.2008.

Research output: Contribution to journalArticle

Venkatakrishnan, C. D. ; Dunsmore, Kathy ; Wong, Hector ; Roy, Sashwathi ; Sen, Chandan K. ; Wani, Altaf ; Zweier, Jay L. ; Ilangovan, Govindasamy. / HSP27 regulates p53 transcriptional activity in doxorubicin-treated fibroblasts and cardiac H9c2 cells : p21 upregulation and G2/M phase cell cycle arrest. In: American Journal of Physiology - Heart and Circulatory Physiology. 2008 ; Vol. 294, No. 4.
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abstract = "Treatment of cancer patients with anthracyclin-based chemotherapeutic drugs induces congestive heart failure by a mechanism involving p53. However, it is not known how p53 aggravates doxorubicin (Dox)-induced toxicity in the heart. On the basis of in vitro acute toxicity assay using heat shock factor-1 (HSF-1) wild-type (HSF-1+/+) and HSF-1-knockout (HSF-1-/-) mouse embryonic fibroblasts and neonatal rat cardiomyocyte-derived H9c2 cells, we demonstrate a novel mechanism whereby heat shock protein 27 (HSP27) regulates transcriptional activity of p53 in Dox-treated cells. Inhibition of p53 by pifithrin-α (PFT-α) provided different levels of protection from Dox that correlate with HSP27 levels in these cells. In HSF-1+/+ cells, PFT-α attenuated Dox-induced toxicity. However, in HSF-1-/- cells (which express a very low level of HSP27 compared with HSF-1+/+ cells), there was no such attenuation, indicating an important role of HSP27 in p53-dependent cell death. On the other hand, immunoprecipitation of p53 was found to coimmunoprecipitate HSP27 and vice versa (confirmed by Western blotting and matrix-assisted laser desorption/ionization time of flight), demonstrating HSP27 binding to p53 in Dox-treated cells. Moreover, upregulation of p21 was observed in HSF-1+/+ and H9c2 cells, indicating that HSP27 binding transactivates p53 and enhances transcription of p21 in response to Dox treatment. Further analysis with flow cytometry showed that increased expression of p21 results in G2/M phase cell cycle arrest in Dox-treated cells. Overall, HSP27 binding to p53 attenuated the cellular toxicity by upregulating p21 and prevented cell death.",
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T2 - p21 upregulation and G2/M phase cell cycle arrest

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AU - Dunsmore, Kathy

AU - Wong, Hector

AU - Roy, Sashwathi

AU - Sen, Chandan K.

AU - Wani, Altaf

AU - Zweier, Jay L.

AU - Ilangovan, Govindasamy

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AB - Treatment of cancer patients with anthracyclin-based chemotherapeutic drugs induces congestive heart failure by a mechanism involving p53. However, it is not known how p53 aggravates doxorubicin (Dox)-induced toxicity in the heart. On the basis of in vitro acute toxicity assay using heat shock factor-1 (HSF-1) wild-type (HSF-1+/+) and HSF-1-knockout (HSF-1-/-) mouse embryonic fibroblasts and neonatal rat cardiomyocyte-derived H9c2 cells, we demonstrate a novel mechanism whereby heat shock protein 27 (HSP27) regulates transcriptional activity of p53 in Dox-treated cells. Inhibition of p53 by pifithrin-α (PFT-α) provided different levels of protection from Dox that correlate with HSP27 levels in these cells. In HSF-1+/+ cells, PFT-α attenuated Dox-induced toxicity. However, in HSF-1-/- cells (which express a very low level of HSP27 compared with HSF-1+/+ cells), there was no such attenuation, indicating an important role of HSP27 in p53-dependent cell death. On the other hand, immunoprecipitation of p53 was found to coimmunoprecipitate HSP27 and vice versa (confirmed by Western blotting and matrix-assisted laser desorption/ionization time of flight), demonstrating HSP27 binding to p53 in Dox-treated cells. Moreover, upregulation of p21 was observed in HSF-1+/+ and H9c2 cells, indicating that HSP27 binding transactivates p53 and enhances transcription of p21 in response to Dox treatment. Further analysis with flow cytometry showed that increased expression of p21 results in G2/M phase cell cycle arrest in Dox-treated cells. Overall, HSP27 binding to p53 attenuated the cellular toxicity by upregulating p21 and prevented cell death.

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