Integrin α5β1 and ADAM-17 Interact in Vitro and Co-localize in Migrating HeLa Cells

Abstract

Tumor necrosis factor (TNF) ␣-converting enzyme (TACE/ADAM-17) has diverse roles in the proteolytic processing of cell surface molecules and, due to its ability to process TNF␣, is a validated therapeutic target for anti-inflammatory therapies. Unlike a number of other ADAM proteins, which interact with integrin receptors via their disintegrin domains, there is currently no evidence for an ADAM-17-integrin association. By analyzing the adhesion of a series of cell lines with recombinant fragments of the extracellular domain of ADAM-17, we now demonstrate a functional interaction between ADAM-17 and ␣ 5 ␤ 1 integrin in a trans orientation. Because ADAM-17-mediated adhesion was sensitive to RGD peptides and EDTA, and the integrin-binding site within ADAM-17 was narrowed down to the disintegrin/ cysteine-rich region, the two molecules appear to have a ligand-receptor relationship mediated by the ␣ 5 ␤ 1 ligand binding pocket. Intriguingly, ADAM-17 and ␣ 5 ␤ 1 were found to co-localize in both membrane ruffles and focal adhesions in HeLa cells. When confluent HeLa cell monolayers were wounded, ADAM-17 and ␣ 5 ␤ 1 redistributed to the leading edge and co-localized, which is suggestive of a cis orientation. We postulate that the interaction of ADAM-17 with ␣ 5 ␤ 1 may target or modulate its metalloproteolytic activity.

Mammary ductal morphogenesis requires paracrine activation of stromal EGFR via ADAM17-dependent shedding of epithelial amphiregulin

“…Phosphorylation of the cytoplasmic domain of ADAM17 appears to regulate processing of some substrates (Diaz-Rodriguez et al, 2002;Fan et al, 2003), whereas the cytoplasmic domain is dispensable for the processing of others (Reddy et al, 2000). Integrin ␣ 5 ␤ 1 may also influence ADAM17 activity (Bax et al, 2004), and Eve-1/Sh3d19, which binds to the cytoplasmic domain of various ADAMs, appears to promote the processing of EGFR ligands, including AREG (Tanaka et al, 2004). Interestingly, our microarray data indicate that Sh3d19 expression mirrors that of ADAM17 in developing mammary gland.…”

The Role of Metalloproteinase ADAM17 in Regulating ICOS Ligand–Mediated Humoral Immune Responses

“…Alternatively, ADAM17 may regulate cell surface expression of ICOSL independent of its proteolytic activity. In addition to the proteolytic function, some members of the ADAM family, including ADAM17, have been shown to support integrin binding via their disintegrin domains (38). Therefore, adhesive properties of ADAM17, through a yet-to-be-identified mechanism, might be responsible for controlling ICOSL cell surface levels.…”

The ADAM17–amphiregulin–EGFR Axis in Mammary Development and Cancer

“…Phosphorylation of the cytoplasmic domain of ADAM17 also appears to regulate the processing of some ADAM17 substrates [70,71], whereas the ADAM17 cytoplasmic domain appears to be dispensable for the processing of other substrates [72]. In addition, evidence suggests that integrin α 5 β 1 may affect ADAM17 activity [73], and the Adam and Eve inspired EVE-1/Sh3d19/PACSIN3 protein, which binds to the cytoplasmic domain of various ADAMs and which has comparable expression to that of ADAM17 during mammary development [28], appears to promote the processing of various EGFR ligands, including AREG [74]. Perhaps most interestingly, the prolonged activation of EGFR itself in human breast and epidermoid carcinoma cells causes substantial increases in processed (i.e., active) ADAM17 levels not by increasing its transcription or translation, but by enhancing its stability [75].…”