The differential regulation of podosome formation by Cytohesin-2 is mediated via α5β1 integrin and the small GTPase RhoA

Abstract

The interaction of a cell with the surrounding tissue is crucial not only to provide mechanical stability but also to enable adaptation of cellular responses to the environment. Recognition of extracellular matrix (ECM) is primarily exerted by integrin receptors, which consist of an α and a β chain. The high diversity of these integrin dimers allows for a variety of ligand specificities but differences in intracellular signaling cascades are only partially understood. <br > Within the cell, integrin-mediated adhesions are organized in specific adhesion structures. Among these, podosomes are characterized by their typical organization in an actin-rich core and an adhesive ring structure, which contains integrins and adhesion-related adaptor and signaling molecules. Apart from mediating adhesion, podosomes also are important during cell migration and invasion. Their formation is induced by growth factor or integrin signaling and depends very much on actin remodeling factors. Both actin dynamics and integrin signaling are regulated by numerous factors, including the cytohesin protein family. Cytohesins primarily act as guanine nucleotide exchange factors (GEFs) for Arf GTPases, but they are also involved in activation and recycling of integrins, as well as actin remodeling processes. One member of the cytohesin family, Cyth2, has recently been shown to be important for podosome formation in THP-1 cells. So far, though, it is unclear whether integrin-related functions of cytohesins also affect podosome formation. Therefore, this study aimed at identifying the role of different cytohesins in integrin-dependent podosome formation. <br > Using bone marrow-derived dendritic cells (BMDCs) from several cytohesin KO mice, we found that only Cyth2, but not Cyth1, Cyth3 or Cyth4, regulates podosome formation in a matrix-depending manner. Loss of Cyth2 impaired podosome formation on a fibronectin (FN) matrix, while Cyth2 KO BMDCs on fibrinogen or Icam-1 remained unaffected. Moreover, Cyth2 KO cells cultured on gelatin or collagen even increased their podosome numbers compared to wildtype cells. These effects were dependent on a differential involvement of α5β1 integrin and were mediated via the small GTPase RhoA. <br > Thus, our results show that Cyth2 is involved in regulation of integrin-dependent responses to environmental cues leading to differential formation of podosomes. Such specific adaptations of signaling pathways downstream of different integrins might affect invasion and migration processes of immune cells in certain ECM microenvironments in vivo.