Roles of POPX2 phosphatase in the regulation of apoptosis

Abstract

TGF-β activated kinase 1 (TAK1) is a MAPK kinase kinase that has emerged as an important regulator in immune response, embryo development, tumorigenesis and cell death. In this study, we have identified TAK1 as a potential target of POPX2 by pull-down mass spectrometry. POPX2 is a PP2C family serine/threonine phosphatase. Association of TAK1 with POPX2 is confirmed by co-immunoprecipitation assay. An in vitro binding assay further indicates direct interaction between POPX2 and TAK1. We have found that POPX2 negatively regulates the activity of TAK1, which promotes genotoxic stress-induced apoptosis in response to VP-16. Increased activity of TAK1 in POPX2-knockdown cells results in increased IKKβ activity and subsequent transcription of anti-apoptotic genes mediated by NF-κB, thus contributing to resistance to genotoxin. In addition, we also investigated the influence of matrix stiffness in cancer progression. The rigidity of matrix plays crucial roles in tumorigenesis. Physical signals provided by the substrates are sensed and conveyed by focal adhesions and actin cytoskeletons to the interior of the cell to elicit responses. Since POPX2 has been implicated in the regulation of stress fibers and focal adhesions, the phosphatase may affect responses of cancer cells to the surrounding environment. By culturing cancer cells on polyacrylamide gels of different rigidities, we found a positive correlation between increased cell spreading and enhanced formation of stress fibers. We also observed increased number of focal adhesions with increased substrate stiffness. Interestingly, POPX2 may regulate cell response to matrix stiffness by promoting cell spreading, formation of stress fibers and focal adhesions. Apart from regulating cell morphology, higher substrate stiffness also increases cell growth. This is partly due to higher rigidity-dependent mitogenic activities of FAK, ERK, STAT3 and less basal apoptosis. Moreover, cells with reduced POPX2 levels induced by siRNA show higher growth and are more resistant to genotoxin.