title: Pyk2 inhibition of p53 as an adaptive and intrinsic mechanism facilitating cell proliferation and survival.
abstract: Pyk2 is a cytoplasmic tyrosine kinase related to focal adhesion kinase (FAK). Compensatory Pyk2 expression occurs upon FAK loss in mice. However, the impact of Pyk2 up-regulation remains unclear. Previous studies showed that nuclear-localized FAK promotes cell proliferation and survival through FAK FERM domain-enhanced p53 tumor suppressor degradation (Lim, S. T., Chen, X. L., Lim, Y., Hanson, D. A., Vo, T. T., Howerton, K., Larocque, N., Fisher, S. J., Schlaepfer, D. D., and Ilic, D. (2008) Mol. Cell 29, 9-22). Here, we show that FAK knockdown triggered p53 activation and G(1) cell cycle arrest in human umbilical vein endothelial cells after 4 days. However, by 7 days elevated Pyk2 expression occurred with a reduction in p53 levels and the release of the G(1) block under conditions of continued FAK knockdown. To determine whether Pyk2 regulates p53, experiments were performed in FAK(-/-)p21(-/-) mouse embryo fibroblasts expressing endogenous Pyk2 and in ID8 ovarian carcinoma cells expressing both Pyk2 and FAK. In both cell lines, Pyk2 knockdown increased p53 levels and inhibited cell proliferation associated with G(1) cell cycle arrest. Pyk2 FERM domain re-expression was sufficient to reduce p53 levels and promote increased BrdUrd incorporation. Pyk2 FERM promoted Mdm2-dependent p53 ubiquitination. Pyk2 FERM effects on p53 were blocked by proteasomal inhibition or mutational-inactivation of Pyk2 FERM nuclear localization. Staurosporine stress of ID8 cells promoted endogenous Pyk2 nuclear accumulation and enhanced Pyk2 binding to p53. Pyk2 knockdown potentiated ID8 cell death upon staurosporine addition. Moreover, Pyk2 FERM expression in human fibroblasts upon FAK knockdown prevented cisplatin-mediated apoptosis. Our studies demonstrate that nuclear Pyk2 functions to limit p53 levels, thus facilitating cell growth and survival in a kinase-independent manner.
authors.affiliation: Department of Reproductive Medicine, Moores Cancer Center, University of California San Diego, La Jolla, California 92093, USA.
authors.forename: Ssang-Taek
authors.initials: ST
authors.lastname: Lim
authors.forename: Nichol L G
authors.initials: NL
authors.lastname: Miller
authors.forename: Ju-Ock
authors.initials: JO
authors.lastname: Nam
authors.forename: Xiao Lei
authors.initials: XL
authors.lastname: Chen
authors.forename: Yangmi
authors.initials: Y
authors.lastname: Lim
authors.forename: David D
authors.initials: DD
authors.lastname: Schlaepfer
mesh_terms: D021581:Active Transport, Cell Nucleus; D000222:Adaptation, Physiological; D000818:Animals; D001973:Bromodeoxyuridine / Q000378:metabolism; D002453:Cell Cycle / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002470:Cell Survival / Q000235:genetics; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D051418:Focal Adhesion Kinase 2 / Q000737:chemistry / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic; D055785:Gene Knockdown Techniques; D006801:Humans; D051379:Mice; D009154:Mutation; D017434:Protein Structure, Tertiary; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism; D019311:Staurosporine / Q000494:pharmacology; D013312:Stress, Physiological / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*
chemical_list: D050759:Cyclin-Dependent Kinase Inhibitor p21; D016159:Tumor Suppressor Protein p53; D051736:Proto-Oncogene Proteins c-mdm2; D051418:Focal Adhesion Kinase 2; D001973:Bromodeoxyuridine; D019311:Staurosporine