Proteolysis of Cortactin by Calpain Regulates Membrane Protrusion during Cell Migration

Abstract

Calpain 2 regulates membrane protrusion during cell migration. However, relevant substrates that mediate the effects of calpain on protrusion have not been identified. One potential candidate substrate is the actin binding protein cortactin. Cortactin is a Src substrate that drives actin polymerization by activating the Arp2/3 complex and also stabilizes the cortical actin network. We now provide evidence that proteolysis of cortactin by calpain 2 regulates membrane protrusion dynamics during cell migration. We show that cortactin is a calpain 2 substrate in fibroblasts and that the preferred cleavage site occurs in a region between the actin binding repeats and the alpha-helical domain. We have generated a mutant cortactin that is resistant to calpain proteolysis but retains other biochemical properties of cortactin. Expression of the calpain-resistant cortactin, but not wild-type cortactin, impairs cell migration and increases transient membrane protrusion, suggesting that calpain proteolysis of cortactin limits membrane protrusions and regulates migration in fibroblasts. Furthermore, the enhanced protrusion observed with the calpain-resistant cortactin requires both the Arp2/3 binding site and the Src homology 3 domain of cortactin. Together, these findings suggest a novel role for calpain-mediated proteolysis of cortactin in regulating membrane protrusion dynamics during cell migration.

Podosome-type adhesions and focal adhesions, so alike yet so different

“…Since cortactin is a major component of PTA, its degradation by calpain may play a key role in their Perrin , 2006 et al disassembly. In addition, PTA turnover depends on degradation of WASP by calpain ( ) and on its stabilization by WIP ( Calle , 2006 et al ).…”

Extracellular Signal-regulated Kinase (ERK) Regulates Cortactin Ubiquitination and Degradation in Lung Epithelial Cells

“…Cortactin binds to filamentous actin to act as a regulator of cytoskeleton dynamics (1, 2, 6), thus understanding cortactin stability is essential in regulating cell motility. Prior studies have demonstrated that calpain 2 mediates proteolysis of cortactin in fibroblasts (37) and platelets (38). Perrin et al showed that calpain 2 cleaves cortactin between the actin binding repeats and the ␣-helical domain (37).…”

“…Phosphorylation is a molecular signature that often leads to recruitment of the ubiquitination E3 ligase complex to a target protein (34 -36). Several studies have shown that calpain 2 regulates cortactin degradation (37,38); however, cortactin degradation through the ubiquitin proteolytic system has not been studied. Here, we show for the first time that ␤-Trcp, 2 an E3 ligase component, is sufficient to mediate elimination of cortactin by the ubiquitin-proteasome system.…”

Regulation of cell polarity by controlled proteolytic systems

“…Knock down of calpain 2 by siRNA results in an increase in both the speed and frequency of the occurrence of the protrusion and retraction events at the membrane (Franco et al, 2004). Overexpression of a proteolysis-resistant form of cortactin results in defects similar to those observed in calpain 2 knockdown cells (Perrin et al, 2006).…”

“…Most efforts have been directed at understanding polarized molecular sorting as the main mechanism underlying the differential accumulation of a limiting factor in a polarized cell (Bradke and Dotti, 1997). More recently, the concept has emerged of spatially restricted protein degradation as an alternative and complementary mechanism responsible for the enrichment of polarizing factors (Perrin et al, 2006;Schwamborn et al, 2007b;Wang et al, 2003;Yan et al, 2006). This review focuses on two of the most conspicuous proteolytic systems in cells, the calpain and ubiquitin proteasome systems, and their roles in the acquisition of polarity of both migrating cells and neurons.…”