Abl Kinases Are Required for Invadopodia Formation and Chemokine-induced Invasion

Abstract

The Abl tyrosine kinases, Abl and Arg, play a role in the regulation of the actin cytoskeleton by modulating cell-cell adhesion and cell motility. Deregulation of both the actin cytoskeleton and Abl kinases have been implicated in cancers. Abl kinase activity is elevated in a number of metastatic cancers and these kinases are activated downstream of several oncogenic growth factor receptor signaling pathways. However, the role of Abl kinases in regulation of the actin cytoskeleton during tumor progression and invasion remains elusive. Here we identify the Abl kinases as essential regulators of invadopodia assembly and function. We show that Abl kinases are activated downstream of the chemokine receptor, CXCR4, and are required for cancer cell invasion and matrix degradation induced by SDF1␣, serum growth factors, and activated Src kinase. Moreover, Abl kinases are readily detected at invadopodia assembly sites and their inhibition prevents the assembly of actin and cortactin into organized invadopodia structures. We show that active Abl kinases form complexes with membrane type-1 matrix metalloproteinase (MT1-MMP), a critical invadopodia component required for matrix degradation. Further, loss of Abl kinase signaling induces internalization of MT1-MMP from the cell surface, promotes its accumulation in the perinuclear compartment and inhibits MT1-MMP tyrosine phosphorylation. Our findings reveal that Abl kinase signaling plays a critical role in invadopodia formation and function, and have far-reaching implications for the treatment of metastatic carcinomas.Podosomes and invadopodia are specialized protrusive structures consisting of a core assembly of F-actin-and actinbinding proteins that form on the ventral surface of migratory and invading cells. These structures are observed in physiological and pathological processes that involve remodeling of the extracellular environment and are found in endothelial cells during extracellular matrix (ECM) 5 degradation (1), transmigrating monocytic cells (2, 3), osteoclasts during bone reabsorption (4), and cancer cells during invasion and metastasis (5). Although podosomes and invadopodia are structurally distinct, they share many common features such as the enrichment of integrins, actin regulatory proteins, matrix metalloproteinases (MMPs), and tyrosine kinases (6 -8).Carcinoma cells utilize invadopodia to degrade the ECM during tumor invasion and metastasis (8). Invadopodia assembly occurs through sequential steps that begin with the assembly of precursor structures containing actin, cortactin, Tsk5, N-WASP, and other actin regulatory proteins, and progress into mature structures with matrix degradation activity (9). Invadopodia were first described in cells transformed with oncogenic v-Src (10), and endogenous Src kinases have been shown to promote podosome/invadopodia formation in response to growth factors and chemokines (1, 11-13). Src phosphorylates several invadopodia components including cortactin, N-WASP, and Tsk5/FISH (14). Cortactin regulates the formation ...

Sos1 Regulates Macrophage Podosome Assembly and Macrophage Invasive Capacity

“…In studies in which we demonstrated a role of Abl in macrophage podosome assembly and function, we highlighted the fact that, based on the evidence that the Abl/Arg family of cytoplasmic tyrosine kinases also regulate invadopodia formation and function (19,20), pharmacologic targeting of Abl may result in inhibition of macrophage infiltration into tumors and tumor cell invasive capacity (18). Sos1 can play the same role in macrophages and tumor cells.…”

“…Together with the evidence that the myeloid-specific Src kinases Hck and Fgr bind to both b1 and b2 integrins and Abl (17), these findings point to a key role of an integrin/Src kinase/ Abl pathway in macrophage podosome assembly. Importantly, recent studies demonstrated that Src and Abl/Arg cross talk in invadopodia formation in response to chemokine and growth factor receptors in carcinoma cells (19,20), thus highlighting the existence of a common signaling module regulating macrophage podosome and tumor cell invadopodia assembly.…”

“…In addition, the Rho GTPase GEF p21-activated kinase 1-interacting exchange factor (aPIX) was reported to localize to podosomes and to regulate their stabilization in human macrophages (51). Considering the role of Src kinases and Abl in regulating both macrophage podosomes (18) and cancer cell invadopodia assembly (19,20), we reasoned that Sos1, whose tyrosine phosphorylation by Abl dictates the acquisition of its Rac GEF activity (28), was a potentially critical candidate in regulating Rac-dependent podosome assembly downstream this Src/Abl signaling pathway. Notably, Eps8, an activator of the Rac-GEF activity of Sos1, was shown to interact with the phosphoprotein paladin, a podosome-associated protein, in mesenchymal cells (29).…”

“…In addition, we reported that the cytoplasmic tyrosine kinase Abl, which forms a tripartite complex with the Src kinases Hck or Fgr bound to integrins (17), redistributes to podosomes, and its silencing or pharmacologic inhibition results in podosome disassembly and reduction of macrophage threedimensional migration and ECM degradative capacity (18). Notably, Abl has been also implicated in invadopodia formation in neoplastic cells (19,20).…”

Signaling inputs to invadopodia and podosomes

“…In cancer cells, epidermal growth factor (EGF) is a well-described inducer of invadopodia formation (Mader et al, 2011;Yamaguchi et al, 2005). TGF-b (Mandal et al, 2008;Pignatelli et al, 2012), heparin binding (HB)-EGF (Díaz et al, 2013;Hayes et al, 2012), VEGF (Lucas et al, 2010) and hepatocyte growth factor/scatter factor (HGF) (Rajadurai et al, 2012) increase invadopodia numbers, whereas stromal cell derived factor 1a (SDF1a) has been shown to increase ECM degradation by breast cancer cells (Smith-Pearson et al, 2010). Many of these growth factor pathways converge on common signaling hubs, especially Src (1) Invadopodia/ podosome initiation.…”