BRK phosphorylates PSF promoting its cytoplasmic localization and cell cycle arrest.
BReast tumor Kinase (BRK) also known as protein kinase 6 (PTK6) is a non-receptor tyrosine kinase overexpressed in the majority of human breast carcinoma. The expression of BRK is a known prognostic marker of breast carcinoma. BRK has been shown to lie downstream of epidermal growth factor (EGF) signaling and mediate effects on cell proliferation and migration. To identify BRK substrates and interacting proteins, we undertook a proteomic approach. BRK immune complexes were purified from the BT-20 breast cancer cell line and analyzed by mass spectrometry. Herein, we report the identification of PSF, the polypyrimidine tract-binding (PTB) protein-associated splicing factor, as a BRK-interacting protein and substrate. BRK and PSF co-eluted in a large protein complex that was regulated by EGF stimulation. Furthermore, BRK and PSF co-immunoprecipitated in BT-20 cells and we defined the interaction as being an SH3 domain-polyproline interaction. The C-terminal tyrosines of PSF were the site of phosphorylation by BRK. Moreover, tyrosine phosphorylation of PSF was also observed upon EGF stimulation, consistent with a role of PSF and BRK downstream of the EGF receptor. Interestingly, the tyrosine phosphorylation promoted the cytoplasmic relocalization of PSF, impaired its binding to polypyrimidine RNA, and led to cell cycle arrest. Our findings show that BRK targets the PSF RNA-binding protein during EGF stimulation.
Terry Fox Molecular Oncology Group, Lady Davis Institute for Medical Research, Sir Mortimer B. Davis Jewish General Hospital, Canada.
Kiven E
KE
Lukong
Marc-Etienne
ME
Huot
Stéphane
S
Richard
D001943:Breast Neoplasms / Q000378:metabolism; D002453:Cell Cycle; D002460:Cell Line; D002465:Cell Movement; D049109:Cell Proliferation; D003593:Cytoplasm / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000378:metabolism / Q000494:pharmacology; D005260:Female; D006801:Humans; D009363:Neoplasm Proteins / Q000378:metabolism*; D000072670:PTB-Associated Splicing Factor; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D015398:Signal Transduction; D018909:src Homology Domains
D009363:Neoplasm Proteins; D000072670:PTB-Associated Splicing Factor; D016601:RNA-Binding Proteins; D004815:Epidermal Growth Factor; D011505:Protein-Tyrosine Kinases; C088375:PTK6 protein, human