UV-mediated regulation of the anti-senescence factor Tbx2.
Several lines of evidence have implicated members of the developmentally important T-box gene family in cell cycle regulation and in cancer. Importantly, the highly related T-box factors Tbx2 and Tbx3 can suppress senescence through repressing the cyclin-dependent kinase inhibitors p19(ARF) and p21(WAF1/CIP1/SDII). Furthermore, Tbx2 is up-regulated in several cancers, including melanomas where it was shown to function as an anti-senescence factor, suggesting that this may be one of the mechanisms by which T-box proteins contribute to the oncogenic process. However, very little is known about whether Tbx2 is regulated by p21-mediated stress-induced senescence signaling pathways. In this study, using the MCF-7 breast cancer cell line known to overexpress Tbx2, we show that in response to stress induced by ultraviolet irradiation the Tbx2 protein is specifically phosphorylated by the p38 mitogen-activated protein kinase. Using site-directed mutagenesis and in vitro kinase assays, we have identified serine residues 336, 623, and 675 in the Tbx2 protein as the p38 target sites and show that these sites are phosphorylated in vivo. Importantly, we show by Western blotting, immunofluorescence, and reporter assays that this phosphorylation leads to increased Tbx2 protein levels, predominant nuclear localization of the protein, and an increase in the ability of Tbx2 to repress the p21(WAF1/CIP1/SDII) promoter. These results show for the first time that the ability of Tbx2 to repress the p21 gene is enhanced in response to a stress-induced senescence pathway, which leads to a better understanding of the regulation of the anti-senescence function of Tbx2.
Abrahams
Amaal
A
Divisions of Medical Biochemistry and Cell Biology, Faculty of Health Sciences, University of Cape Town, Observatory 7925, Cape Town, South Africa.
Mowla
Shaheen
S
Parker
M Iqbal
MI
Goding
Colin R
CR
Prince
Sharon
S
D021581:Active Transport, Cell Nucleus / Q000235:genetics / Q000528:radiation effects; D000818:Animals; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism; D019556:COS Cells; D002453:Cell Cycle / Q000235:genetics / Q000528:radiation effects*; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects*; D016922:Cellular Senescence / Q000235:genetics / Q000528:radiation effects*; D002522:Chlorocebus aethiops; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D020935:MAP Kinase Signaling System / Q000235:genetics / Q000528:radiation effects*; D008545:Melanoma / Q000235:genetics / Q000378:metabolism; D051379:Mice; D016297:Mutagenesis, Site-Directed; D041681:NIH 3T3 Cells; D010766:Phosphorylation; D020825:T-Box Domain Proteins / Q000235:genetics / Q000378:metabolism*; D014466:Ultraviolet Rays; D015854:Up-Regulation / Q000235:genetics / Q000528:radiation effects; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism
C493956:CDKN1A protein, human; C493957:Cdkn1a protein, mouse; C417743:Cdkn2a protein, mouse; D019941:Cyclin-Dependent Kinase Inhibitor p16; D050759:Cyclin-Dependent Kinase Inhibitor p21; C097348:T-Box Domain Protein 2; D020825:T-Box Domain Proteins; C106884:TBX3 protein, human; D048051:p38 Mitogen-Activated Protein Kinases