HSP27 expression regulates CCK-induced changes of the actin cytoskeleton in CHO-CCK-A cells

We investigated how heat shock protein 27 (HSP27) and its phosphorylation a re involved in the action of cholecystokinin (CCK) on the actin cytoskeleto n by genetic manipulation of Chinese hamster ovary (CHO) cells stably trans fected with the CCK-A receptor. In these cells, as in rat acini, CCK activa ted p38 mitogen-activated protein (MAP) kinase and increased the phosphoryl ation of HSP27. This effect could be blocked with the p38 MAP kinase inhibi tor SB-203580. Examination by confocal microscopy of cells stained with rho damine phalloidin showed that CCK dose-dependently induced changes of the a ctin cytoskeleton, including cell shape changes, which were coincident with actin cytoskeleton fragmentation and formation of actin filament patches i n the cells. To further evaluate the role of HSP27, CHO-CCK-A cells were tr ansfected with expression vectors for either wild-type (wt) or mutant (3A, 3G, and 3D) human HSP27. Overexpression of wt-HSP27 and 3D-HSP27 inhibited the effects on the actin cytoskeleton seen after high-dose CCK stimulation. In contrast, overexpression of nonphosphorylatable mutants, 3A- and 3G-HSP 27, or inhibition of phosphorylation of HSP27 by preincubation of wt-HSP27 transfected cells with SB-203580 did not protect the actin cytoskeleton. Th ese results suggest that phosphorylation of HSP27 is required to stabilize the actin cytoskeleton and to protect the cells from the effects of high co ncentrations of CCK.