How cells respond to changing matrix stiffness in 2D and 3D microenvironments: Role of Caveolin-1 and its phosphorylation

Abstract

Caveolae are omega-shaped structures that are found on eukaryotic plasma membranes and enriched in sphingolipids and sterols. Caveolae act as membrane reservoirs to buffer against changes in membrane tension, possibly explaining their abundance in cells experiencing mechanical stress. Caveolin-1 (Cav-1) is an important structural component of caveolae that is phosphorylated on its tyrosine-14. Cav-1 phosphorylation has also been shown to regulate its mechanotransductory properties and focal adhesion tension. In this study, we wanted to determine how WT MEFs respond to changing matrix stiffness in 2D and 3D microenvironments of different matrix stiffness and the role of Cav-1 and its Y14 phosphorylation in mediating this response. Our results show that in WT MEFs seeded on collagen coated 2D polyacrylamide gels of increasing stiffness, Cav-1 phosphorylation increases steadily. Adhesion dependent FAK, Akt and EGFR activation also increase as stiffness increases. Upon loss of Cav-1, this stiffness dependent activation of FAK, Akt and EGFR are all deregulated in cells on gels of stiffness 0.5 kPa, 2.5 kPa and 23 kPa, but maintained on glass. On the other hand, the response of WT MEFs embedded in 3D collagen gels to changing matrix stiffness is different. This is reflected in their differential regulation of Cav-1 phosphorylation, FAK, Akt and EGFR activation. Hence, Cav-1 and its phosphorylation could have different roles in 2D and 3D microenvironments and this could contribute differentially to regulating cellular function.