Cellular stress induces the tyrosine phosphorylation of caveolin-1 (Tyr(14)) via activation of p38 mitogen-activated protein kinase and c-Src kinase - Evidence for caveolae, the actin cytoskeleton, and focal adhesions as mechanical sensors of osmotic stress

Environmental stressors have been recently shown to activate intracellular mitogen-activated protein (MAP) kinases, such as p38 MAP kinase, leading to changes in cellular functioning. However, little is known about the downst ream elements in these signaling cascades. In this study, we show that cave olin-1 is phosphorylated on tyrosine 14 in NIH 3T3 cells after stimulation with a variety of cellular stressors (ie. high osmolarity, H2O2, and UV lig ht). To detect this phosphorylation event, we employed a phosphospecific mo noclonal antibody probe that recognizes only tyrosine 14-phosphorylated cav eolin-1. Since p38 MAP kinase and c-Src have been previously implicated in the stress response, we next assessed their role in the tyrosine phosphoryl ation of caveolin-1. Interestingly, we show that the p38 inhibitor (SB20358 0) and a dominant-negative mutant of c-Src (SRC-RF) both block the stress-i nduced tyrosine phosphorylation of caveolin-1 (Tyr(P)(14)), In contrast, in hibition of the p42/44 MAP kinase cascade did not affect the tyrosine phosp horylation of caveolin-1. These results indicate that extracellular stresso rs can induce caveolin-1 tyrosine phosphorylation through the activation of well established upstream elements, such as p38 MAP kinase and c-Src kinas e, However, heat shock did not promote the tyrosine phosphorylation of cave olin-1 and did not activate p38 MAP kinase, Finally, we show that after hyp erosmotic shock, tyrosine-phosphorylated caveolin-1 is localized near focal adhesions, the major sites of tyrosine kinase signaling. In accordance wit h this localization, disruption of the actin cytoskeleton dramatically pote ntiates the tyrosine phosphorylation of caveolin-1. Taken together, our res ults clearly define a novel signaling pathway involving p38 MAP kinase acti vation and caveolin-1 (Tyr(P)(14)). Thus, tyrosine phosphorylation of caveo lin-1 may represent an important downstream element in the signal transduct ion cascades activated by cellular stress.