Invasive matrix degradation at focal adhesions occurs via protease recruitment by a FAK-p130Cas complex.
Bottom Line: Importantly, we have found that this MT1-MMP targeting is dependent on an association with a FAK-p130Cas complex situated at FAs and is regulated by Src-mediated phosphorylation of Tyr 573 at the cytoplasmic tail of MT1.Disrupting the FAK-p130Cas-MT1 complex significantly impairs FA-mediated degradation and tumor cell invasion yet does not appear to affect invadopodia formation or function.These findings demonstrate a novel function for FAs and also provide molecular insights into MT1-MMP targeting and function.
Affiliation: Department of Biochemistry and Molecular Biology, and the Center for Basic Research in Digestive Diseases, Mayo Clinic and Graduate School, Rochester, MN 55905, USA.Show MeSH
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Mentions: Interaction between MT1-MMP and FAK is essential for FA-centric matrix degradation; however, whether this interaction is direct was unclear. To test this, we purified His-MT1 protein from Escherichia coli, and performed direct binding analysis with GST-PRR of FAK. No direct interaction was detected (unpublished data), suggesting the participation of an adaptor. One candidate known to bind directly to both MT1 (Gingras et al., 2008; Gonzalo et al., 2010) and to the second and third PXXP motifs of FAK is p130Cas (Harte et al., 1996; Mitra et al., 2005). However, whether these three proteins form a functional complex is not clear. To test this possibility, we transfected 293T cells with either MT1 alone or together with p130Cas and then performed pull-down assays with the purified GST-PRR of FAK. As shown in Fig. 4 a, p130Cas strongly binds to GST-PRR, and overexpression of p130Cas significantly enhances the amount of MT1-MMP in the pulldown, suggesting that p130Cas facilitates MT1–FAK interaction. To test if these three proteins interact endogenously, FAK was immunoprecipitated from HT-1080 cells, and both p130Cas and MT1 were detected in the Co-IP (Fig. 4 b). Importantly, the FAK–MT1 interaction was attenuated in p130Cas knockdown cells (Fig. 4 b), suggesting that p130Cas mediates the interaction between MT1-MMP and FAK.
Affiliation: Department of Biochemistry and Molecular Biology, and the Center for Basic Research in Digestive Diseases, Mayo Clinic and Graduate School, Rochester, MN 55905, USA.