ADAM12-mediated focal adhesion formation is differently regulated by beta1 and beta3 integrins.

ADAM12, adisintegrin and metalloprotease, has been demonstrated to be upregulated in human malignant tumors and to accelerate the malignant phenotype in a mouse model for breast cancer. ADAM12 is a substrate for beta1 integrins and may affect tumor and stromal cell behavior through its binding to beta1 integrins. Here, we report that cells deficient in beta1 integrin or overexpressing beta3 integrin can bind to recombinant full-length human ADAM12 via beta3 integrin.

Hierarchy of ADAM12 binding to integrins in tumor cells.

ADAMs (a disintegrin and metalloprotease) comprise a family of cell surface proteins with protease and cell-binding activities. Using different forms and fragments of ADAM12 as substrates in cell adhesion and spreading assays, we demonstrated that alpha9beta1 integrin is the main receptor for ADAM12. However, when alpha9beta1 integrin is not expressed--as in many carcinoma cells--other members of the beta1 integrin family can replace its ligand binding activity.

Regulation of ADAM12 cell-surface expression by protein kinase C epsilon.

The ADAM (a disintegrin and metalloprotease) family consists of multidomain cell-surface proteins that have a major impact on cell behavior. These transmembrane-anchored proteins are synthesized as proforms that have (from the N terminus): a prodomain; a metalloprotease-, disintegrin-like-, cysteine-rich, epidermal growth factor-like, and transmembrane domain; and a cytoplasmic tail. The 90-kDa mature form of human ADAM12 is generated in the trans-Golgi through cleavage of the prodomain by a furin-peptidase and is stored intracellularly until translocation to the cell surface as a constitutively active protein.

Leukotriene D4-induced calcium signaling in human intestinal epithelial cells.

LTD4 induces a calcium signal that consists of a mobilization from internal stores regulated by PTX-insensitive G protein, Rho, and, an influx across the plasma membrane regulated by PTX-sensitive Gi protein in human intestinal epithelial cells. The LTD4 induced mobilization of Ca2+ is mediated by a PH domain dependent association between PLC-gamma 1 and G beta gamma subunits. This interaction requires Src kinase activity as well as the association of this kinase with PLC-gamma 1, suggesting a G beta gamma mediated recruitment of proteins to the plasma membrane and formation of a signaling complex which is essential for the downstream Ca2+ signal.

ADAM12/syndecan-4 signaling promotes beta 1 integrin-dependent cell spreading through protein kinase Calpha and RhoA.

The ADAMs (a disintegrin and metalloprotease) comprise a large family of multidomain proteins with cell-binding and metalloprotease activities. The ADAM12 cysteine-rich domain (rADAM12-cys) supports cell attachment using syndecan-4 as a primary cell surface receptor that subsequently triggers beta(1) integrin-dependent cell spreading, stress fiber assembly, and focal adhesion formation. This process contrasts with cell adhesion on fibronectin, which is integrin-initiated but syndecan-4-dependent.

Leukotriene D4 induces association of active RhoA with phospholipase C-gamma1 in intestinal epithelial cells.

It has been previously suggested that leukotriene-induced Ca2+ signalling is mediated through a Rho-dependent process, but neither direct activation of Rho nor a mechanism underlying such signalling has been reported. Accordingly, we used the Rhotekin binding assay to assess RhoA activation in intestinal epithelial cells and observed that RhoA was activated by leukotriene D4 (LTD4). We also found that, within 15 s, activation of RhoA by LTD4 led to an increased association of RhoA with G-protein betagamma (Gbetagamma) and phospholipase C-gamma1 (PLC-gamma1) in the plasma membrane, as evidenced by the results of co-immunoprecipitation, glutathione S-transferase (GST) pulldown assays, and confocal microscopy.