title: Phosphorylation of FOXO3a on Ser-7 by p38 promotes its nuclear localization in response to doxorubicin.
abstract: 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.
authors: Department of Surgery and Cancer, Imperial College London, Hammersmith Hospital Campus, London W12 0NN, United Kingdom.
authors: Ka-Kei
authors: KK
authors: Ho
authors: Victoria A
authors: VA
authors: McGuire
authors: Chuay-Yeng
authors: CY
authors: Koo
authors: Kyle W
authors: KW
authors: Muir
authors: Natalia
authors: N
authors: de Olano
authors: Evie
authors: E
authors: Maifoshie
authors: Douglas J
authors: DJ
authors: Kelly
authors: Ursula B
authors: UB
authors: McGovern
authors: Lara J
authors: LJ
authors: Monteiro
authors: Ana R
authors: AR
authors: Gomes
authors: Angel R
authors: AR
authors: Nebreda
authors: David G
authors: DG
authors: Campbell
authors: J Simon C
authors: JS
authors: Arthur
authors: Eric W-F
authors: EW
authors: Lam
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000235:genetics; D019943:Amino Acid Substitution; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D004317:Doxorubicin / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D020125:Mutation, Missense; D010766:Phosphorylation / Q000187:drug effects / Q000235:genetics; D011725:Pyridines / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D000903:Antibiotics, Antineoplastic; D004791:Enzyme Inhibitors; C499517:FOXO3 protein, human; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; C474033:FoxO3 protein, mouse; D007093:Imidazoles; D011725:Pyridines; D004317:Doxorubicin; D048051:p38 Mitogen-Activated Protein Kinases; C090942:4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole