Caveolin1 Tyr14 phosphorylation: Regulation and role in adhesion-dependent cellular responsiveness of normal and cancer cells

Abstract

Cell matrix-adhesion regulates cellular responses to changing biochemical and mechanical properties of the extracellular matrix (ECM). This is mediated in part by regulation of membrane trafficking through caveolae to control anchorage-dependent signaling. Caveolae are mechanosensitive plasma membrane invaginations that support signaling and endocytosis. They are formed by Caveolin1 (Cav1) protein in mammalian cells. Tyr-14 phosphorylation of Cav1 (pY14Cav1) is crucial for its role both in caveolae and non-caveolar focal adhesions. Our studies reveal endogenous pY14Cav1 to localize to both caveolae and focal adhesions in mouse fibroblasts. We find total cellular pY14Cav1 levels to be stimulated by cell-matrix adhesion and increasing matrix stiffness, similar to activation of FAK. pY14Cav1 levels in caveolae isolated from adherent and suspended cells are comparable, suggesting cell-matrix adhesion to not regulate caveolar pY14Cav1. This further suggests the focal adhesion-associated pY14Cav1 to be regulated by adhesion. Interestingly, inhibition of FAK activation using a small molecule inhibitor abrogates re-adhesion-mediated recovery of pY14Cav1. FAK inhibition across increasing stiffnesses shows its regulation of pY14Cav1 to be more prominent at higher stiffness. Together, these studies establish the FAK-pY14Cav1 crosstalk at focal adhesions, and its regulation by cell-matrix adhesion. Apart from their role in caveolar endocytosis and focal adhesion function in non-transformed cells, pY14Cav1 is also thought to have a regulatory role in cancers. The role of Cav1 as a tumour suppressor or promoter in cancers is contested, and could in part be dependent on its Tyr-14 phosphorylation. We find that adhesion regulates pY14Cav1 levels in some, but not all, Cav1-expressing cancer cell lines. Changes in pY14Cav1 levels could hence contribute to the cancer cell phenotype. Protein tyrosine phosphatase (PTP) levels and activation could help mediate the same. si-RNA-mediated targeting of PTPs increases pY14Cav1 levels in T24 bladder cancer cells to support anchorage-independent signaling. Taken together, these studies reveal a distinct adhesion-dependent regulation and role for Cav1 Tyr-14 phosphorylation in normal and cancer cells.