Abstract
[Purpose] Heat shock protein 27 (HSP27) is implicated in diverse biological function. Cytoprotection by HSP27, such as suppression of apoptosis and prevention from stress-induced disruption of the cytoskeleton, was reported to be expressed via its interaction with cytochrome c and co-localization with actin filaments. We recently found that HSP27 is involved in a protective role against UVC-induced cell death in human cells. In the present study, to elucidate the molecular mechanisms underlying UVC resistance, we searched for HSP27-interacted proteins in UVC-resistant human cells by affinity column chromatography using HSP27 protein-bound Sepharose. [Methods]The HSP27 protein expressed as a fusion protein with GST (GST-HSP27) was purified from E.coli extracts using a GSH-Sepharose column and then conjugated with NHS-activated Sepharose. Cell lysates from human UVC- resistant cells were applied to the affinity column, and subjected to SDS-PAGE and mass spectrometrical analyses. [Results and Conclusion]Five proteins appeared to specifically bind to GST-HSP27-conjugated Sepharose compared with GST-conjugated Sepharose. Among the five candidates for HSP27-interacted proteins, annexin II and HSP70 proteins were much more co-immunoprecipitated with HSP27 in UVC-resistant cell lysates than in UVC-sensitive cell lysates. Furthermore, the protein amount of annexin II increased after UVC irradiation in the nuclear fraction of UVC-resistant cells. Thus, the two HSP27-interacted proteins may be involved in UVC resistance in human cells.
