The role of SNX26 in EGFR-mediated circular dorsal ruffle formation

Abstract

A large number of plasma membrane structures induced dynamic changes by a variety of mechanical and chemical stimuli. Circular dorsal ruffles (CDRs) are dynamic and transient actin-rich ring-like membrane structures that arise from the dorsal side of mammalian cells treated with various growth factors. Physiological functions of CDRs are implicated in the down-regulation of growth factor receptors and the migration of cells. However, the precise mechanism by which the dynamics and formation of CDRs are regulated is largely unknown. Here, we show that Sorting nexin 26 (SNX26), a brain-enriched RhoGAP containing protein with multiple signaling domains, interacts with actin and regulates the formation of CDRs via the RhoGAP domain in COS-7 cells. SNX26 interacts with actin via its PX and RhoGAP domains, but not SH3 and PRD domains. Overexpression of SNX26 full-length and its RhoGAP domain-containing deletion mutants decreased the filamentous actin content in COS-7 cells by ~50%. Overexpression of N-terminal comprising of PX, SH3 and Rho GAP domains inhibited CDR formation induced by epidermal growth factor (EGF) in cells. On the other hand, N-term R350I, which is the GAP defective mutant form of N-term, increased the number of cells forming CDRs in response to EGF. Interestingly, although PX and PX Y100A (defective mutant of phospholipid binding affinity) domains are capable of binding to actin, they cannot affect the formation of CDR. Moreover SNX26 has the capability for intra- or inter-molecular interaction through PX and RhoGAP domains. These results indicate that SNX26, acting as a RhoGAP, regulates actin cytoskeleton and modulates CDR formation by EGF treatment in cells. Furthermore its function might be regulated by intra/inter-molecular interaction event. Taken together, it has raised possibility that SNX26, acting as a regulator of cytoskeletal remodeling, has a key role in EGFR-mediated CDR formation via Rho protein signaling pathway in cells.