Specific tyrosine phosphorylation sites on cortactin regulate Nck1-dependent actin polymerization in invadopodia

Department of Anatomy and Structural Biology, Albert Einstein College of Medicine of Yeshiva University, Bronx, NY 10461, USA.
Journal of Cell Science (Impact Factor: 5.43). 11/2010; 123(Pt 21):3662-73. DOI: 10.1242/jcs.068163
Source: PubMed


Invadopodia are matrix-degrading membrane protrusions in invasive carcinoma cells enriched in proteins that regulate actin polymerization. The on-off regulatory switch that initiates actin polymerization in invadopodia requires phosphorylation of tyrosine residues 421, 466, and 482 on cortactin. However, it is unknown which of these cortactin tyrosine phosphorylation sites control actin polymerization. We investigated the contribution of individual tyrosine phosphorylation sites (421, 466, and 482) on cortactin to the regulation of actin polymerization in invadopodia. We provide evidence that the phosphorylation of tyrosines 421 and 466, but not 482, is required for the generation of free actin barbed ends in invadopodia. In addition, these same phosphotyrosines are important for Nck1 recruitment to invadopodia via its SH2 domain, for the direct binding of Nck1 to cortactin in vitro, and for the FRET interaction between Nck1 and cortactin in invadopodia. Furthermore, matrix proteolysis-dependent tumor cell invasion is dramatically inhibited in cells expressing a mutation in phosphotyrosine 421 or 466. Together, these results identify phosphorylation of tyrosines 421 and 466 on cortactin as the crucial residues that regulate Nck1-dependent actin polymerization in invadopodia and tumor cell invasion, and suggest that specifically blocking either tyrosine 421 or 466 phosphorylation might be effective at inhibiting tumor cell invasion in vivo.

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Available from: Marco Magalhaes, Oct 29, 2015
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    • "This model is in agreement with previous findings on Arp2/3 complex in growth cone lamellipodia (Korobova and Svitkina, 2008; Yang et al., 2012; San Miguel-Ruiz and Letourneau, 2014) and on cortactin's regulatory role in Arp2/3-dependent actin branch formation and stabilization in nonneuronal cells (Weed et al., 2000; Uruno et al., 2001; Weaver et al., 2001, 2002; Bryce et al., 2005; Kempiak et al., 2005; Kowalski et al., 2005; Ammer and Weed, 2008; Cai et al., 2008); however, Src was not specifically investigated in these studies. This hypothesis is supported by in vitro findings that actin assembly can be enhanced via Src, cortactin, N-WASP, Nck, and Arp2/3 (Tehrani et al., 2007), as well as by studies on actin polymerization during invadopodium assembly of carcinoma cells (Oser et al., 2010). Previous studies in nonneuronal cells suggest that Src-mediated tyrosine phosphorylation of cortactin promotes cell migration and cancer metastasis (Huang et al., 1998, 2003; Bourguignon et al., 2001; Wang et al., 2011). "
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    • "In terms of signal transduction, growth factor receptor tyrosine kinase and integrin initiated upstream events have been shown to promote invadopodia formation through phosphorylation of cortactin via a Src and Arg dependent pathway (Stylli et al., 2008; Oser et al., 2010; Destaing et al., 2011; Mader et al., 2011; MacGrath and Koleske, 2012). b1 integrin has been shown to promote metastasis, invadopodia maturation, and matrix degradation through Arg (Beaty et al., 2013). "
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    • "The invadopodium precursor is then anchored by binding to PI(3,4)P 2 and further stabilized by β1 integrin-mediated adhesion to the ECM (Beaty et al., 2013; Sharma et al., 2013). Invadopodium maturation begins as β1 integrin activates Arg, which phosphorylates cortactin on Y421 to recruit Nck1 (Beaty et al., 2013; Oser et al., 2010). Talin localizes to the structure and recruits a complex of moesin and NHE-1 through a direct binding interaction with moesin (Beaty et al., 2014). "
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