Cooperative Signaling between α6β4Integrin and ErbB-2 Receptor Is Required to Promote Phosphatidylinositol 3-Kinase-dependent Invasion

Abstract

We previously demonstrated that ␤ 4 integrin subunit overexpression increases in vitro invasiveness of NIH3T3 cells that have been transformed by ErbB-2 oncogene. We used this model to identify domains within the large ␤ 4 cytoplasmic domain that are involved in the interaction of ␣ 6 ␤ 4 with ErbB-2, invasion, and phosphatidylinositol 3-kinase (PI3K) activation. For this purpose, we expressed deletion mutants of ␤ 4 that lacked either all or portions of the ␤ 4 cytoplasmic domain in NIH3T3/ErbB-2 cells. We also used an ecto-domain mutant in which most of the extracellular domain of ␤ 4 was replaced with a c-Myc tag. These transfectants were examined for their ability to invade Matrigel and their ability to activate PI3K, as well as for the ability of ␣ 6 ␤ 4 to co-immunoprecipitate with ErbB-2. The results obtained revealed that a region of the ␤ 4 cytoplasmic domain between amino acids 854 and 1183 is critical for the ability of ␣ 6 ␤ 4 integrin to increase invasion. Interestingly, the extracellular domain of ␤ 4 is not necessary for ␣ 6 ␤ 4 to stimulate invasion. The association of ␣ 6 ␤ 4 with ErbB-2 is dependent upon the ␤ 4 cytoplasmic domain and can occur in the absence of ␣ 6 ␤ 4 heterodimerization. Finally, we observed strong activation of PI3K with ␤ 4 wild type and with those ␤ 4 deletion mutants that were able to stimulate invasion upon the expression in NIH3T3/ErbB-2 cells. In conclusion, our results establish that there is cooperation between ␣ 6 ␤ 4 and ErbB-2 in promoting PI3K-dependent invasion and implicate a specific region of the ␤ 4 cytoplasmic domain (amino acids 854 -1183) in this event.Integrins are the major family of cell surface receptors that mediate attachment to the extracellular matrix. The interaction between integrins and their ligands is involved in the regulation of many cellular functions, including embryonic development, cell proliferation, as well as tumor growth and metastasis. Integrins are composed of ␣ and ␤ transmembrane subunits that heterodimerize to form different receptors. A single ␣ subunit (e.g. ␣ v or ␣ 6 ) can associate with different ␤ subunits, (␤ 1 , ␤ 3 , ␤ 5 , or ␤ 1 , ␤ 4 , respectively), promoting different ligand binding specificity (1-3). The ␣ 6 ␤ 4 integrin is a receptor for various isoforms of the basement membrane component laminin (4 -6), and its expression is restricted to epithelia, endothelia and peripheral nerves (7-9). In many epithelia, ␣ 6 ␤ 4 is found in hemidesmosomes where it plays an essential role in their organization (10,11). This integrin can also interact with F-actin and promote the migration of invasive carcinoma cells (12, 13).The intracellular portion of the ␤ 4 subunit is much larger (1,000 amino acids) than that of all the other known ␤ subunits, and it does not exhibit apparent sequence homology with them (14 -16). Increasing evidence indicates that the ability of ␣ 6 ␤ 4 to regulate cell proliferation, motility, and invasion is dependent upon signal transduction events that are mediated by the ␤ 4 cytoplasmic domain (17-19)...

EMT, the cytoskeleton, and cancer cell invasion

“…Effectors of these signaling cascades include MAPK, NFκB, Jun, β-catenin and others and together they modulate cell proliferation, cell survival, cell migration and invasion. Among the mitogenic integrins are α 6 β 4 or α v β 3 -integrins which cooperate with diverse growth factor receptors, including EGFR, ErbB2, and c-Met [108][109][110][111]. On the other hand, growth inhibitory signals can be transmitted, for example via α 2 β 1 -integrin, which in turn activates p38-MAPKmediated cell-cycle inhibition [112,113].…”

Biologic and therapeutic role of HER2 in cancer

“…These alterations, together with changes in integrin and cadherin function frequently observed in tumor cells with activated HER, facilitate the communication between tumor cells and their escape from control by the microenvironment. In particular, the creation of new interactions of HER family members with integrins, such as HER2 and a 6 b 4 ( Figure 1) and focal adhesion kinases recruits the PI3 K pathway (Campiglio et al, 1994;Tagliabue et al, 1996;Gambaletta et al, 2000), inducing the release of second messengers (phosphoinositides and calcium). These molecules in turn recruit multiprotein complexes that play a role in reorganization of actin cytoskeleton, and thus in tumor cell migration (Dittmar et al, 2002;Feldner and Brandt, 2002).…”

The Met Receptor and α6β4 Integrin Can Function Independently to Promote Carcinoma Invasion

“…Advances include the observation that ␣ 6 ␤ 4 is localized to the leading edge of migrating carcinoma cells where it can contribute to the formation and stabilization of actin protrusions (16,17). In addition, there is evidence from several laboratories indicating that ␣ 6 ␤ 4 stimulates the activity of phosphoinositide 3-OH kinase (PI3K) 1 in invasive carcinoma cells and that PI3K is essential for migration and invasion (7,18). Interestingly, it has been suggested that ␣ 6 ␤ 4 activates PI3K and mediates invasion through its ability to cooperate with specific growth factor receptors (1,4,18).…”