Phosphorylation of FOXO3a on Ser-7 by p38 promotes its nuclear localization in response to doxorubicin

TY - JOUR

T1 - Phosphorylation of FOXO3a on Ser-7 by p38 promotes its nuclear localization in response to doxorubicin

A1 - Ho,Ka-Kei

A1 - McGuire,Victoria A.

A1 - Koo,Chuay-Yeng

A1 - Muir,Kyle W.

A1 - de Olano,Natalia

A1 - Maifoshie,Evie

A1 - Kelly,Douglas J.

A1 - McGovern,Ursula B.

A1 - Monteiro,Lara J.

A1 - Gomes,Ana R.

A1 - Nebreda,Angel R.

A1 - Campbell,David G.

A1 - Arthur,J. Simon C.

A1 - Lam,Eric W. -F.

AU - Ho,Ka-Kei

AU - McGuire,Victoria A.

AU - Koo,Chuay-Yeng

AU - Muir,Kyle W.

AU - de Olano,Natalia

AU - Maifoshie,Evie

AU - Kelly,Douglas J.

AU - McGovern,Ursula B.

AU - Monteiro,Lara J.

AU - Gomes,Ana R.

AU - Nebreda,Angel R.

AU - Campbell,David G.

AU - Arthur,J. Simon C.

AU - Lam,Eric W. -F.

PY - 2012/1/6

Y1 - 2012/1/6

N2 - <p>FOXO3a is a forkhead transcription factor that regulates a multitude of important cellular processes, including proliferation, apoptosis, differentiation, and metabolism. Doxorubicin treatment of MCF-7 breast carcinoma cells results in FOXO3a nuclear relocation and the induction of the stress-activated kinase p38 MAPK. Here, we studied the potential regulation of FOXO3a by p38 in response to doxorubicin. Co-immunoprecipitation studies in MCF-7 cells demonstrated a direct interaction between p38 and FOXO3a. We also showed that p38 can bind and phosphorylate a recombinant FOXO3a directly in vitro. HPLC-coupled phosphopeptide mapping and mass spectrometric analyses identified serine 7 as a major site for p38 phosphorylation. Using a phosphorylated Ser-7 FOXO3a antibody, we demonstrated that FOXO3a is phosphorylated on Ser-7 in response to doxorubicin. Immunofluorescence staining studies showed that upon doxorubicin treatment, the wild-type FOXO3a relocalized to the nucleus, whereas the phosphorylation-defective FOXO3a (Ala-7) mutant remained largely in the cytoplasm. Treatment with SB202190 also inhibits the doxorubicin-induced FOXO3a Ser-7 phosphorylation and nuclear accumulation in MCF-7 cells. In addition, doxorubicin caused the nuclear translocation of FOXO3a in wild-type but not p38-depleted mouse fibroblasts. Together, our results suggest that p38 phosphorylation of FOXO3a on Ser-7 is essential for its nuclear relocalization in response to doxorubicin.</p>

AB - <p>FOXO3a is a forkhead transcription factor that regulates a multitude of important cellular processes, including proliferation, apoptosis, differentiation, and metabolism. Doxorubicin treatment of MCF-7 breast carcinoma cells results in FOXO3a nuclear relocation and the induction of the stress-activated kinase p38 MAPK. Here, we studied the potential regulation of FOXO3a by p38 in response to doxorubicin. Co-immunoprecipitation studies in MCF-7 cells demonstrated a direct interaction between p38 and FOXO3a. We also showed that p38 can bind and phosphorylate a recombinant FOXO3a directly in vitro. HPLC-coupled phosphopeptide mapping and mass spectrometric analyses identified serine 7 as a major site for p38 phosphorylation. Using a phosphorylated Ser-7 FOXO3a antibody, we demonstrated that FOXO3a is phosphorylated on Ser-7 in response to doxorubicin. Immunofluorescence staining studies showed that upon doxorubicin treatment, the wild-type FOXO3a relocalized to the nucleus, whereas the phosphorylation-defective FOXO3a (Ala-7) mutant remained largely in the cytoplasm. Treatment with SB202190 also inhibits the doxorubicin-induced FOXO3a Ser-7 phosphorylation and nuclear accumulation in MCF-7 cells. In addition, doxorubicin caused the nuclear translocation of FOXO3a in wild-type but not p38-depleted mouse fibroblasts. Together, our results suggest that p38 phosphorylation of FOXO3a on Ser-7 is essential for its nuclear relocalization in response to doxorubicin.</p>

U2 - 10.1074/jbc.M111.284224

DO - 10.1074/jbc.M111.284224

M1 - Article

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 2

VL - 287

SP - 1545

EP - 1555

ER -