Cell Surface Transglutaminase Promotes RhoA Activation via Integrin Clustering and Suppression of the Src-p190RhoGAP Signaling Pathway

Janiak A, Zemskov EA and Belkin AM
Source: Mol Biol Cell
Publication Date: (2006)
Issue: 17(4): 1606-19
Research Area:
Cancer Research/Cell Biology
Cells used in publication:
Embryonic fibroblast, mouse (MEF) immort
Species: mouse
Tissue Origin: embryo
Nucleofector® I/II/2b
Tissue transglutaminase (tTG) is a multifunctional protein which serves as cross-linking enzyme and integrin-binding adhesion coreceptor for fibronectin on the cell surface. Previous work showed activation of small GTPase RhoA via enzymatic transamidation by cytoplasmic tTG. Here, we report an alternative nonenzymatic mechanism of RhoA activation by cell surface tTG. Direct engagement of surface tTG with specific antibody or the fibronectin fragment containing modules I6II1,2I7-9 increases RhoA-GTP levels. Integrin-dependent signaling to RhoA and its downstream target ROCK is amplified by surface tTG. tTG expression on the cell surface elevates RhoA-GTP levels in nonadherent and adherent cells, delays maximal RhoA activation upon cell adhesion to fibronectin and accelerates a rise in RhoA activity after binding soluble integrin ligands. These data indicate that surface tTG induces integrin clustering regardless of integrin-ligand interactions. This notion is supported by visualization of integrin clusters, increased susceptibility of integrins to chemical cross-linking and biochemical detection of large integrin complexes in cells expressing tTG. In turn, integrin aggregation by surface tTG inhibits Src kinase activity and decreases activation of the Src substrate, p190RhoGAP. Moreover, pharmacological inhibition of Src kinase reveals inactivation of Src signaling as primary cause of elevated RhoA activity in cells expressing tTG. Together, these findings show that surface tTG amplifies integrin-mediated signaling to RhoA/ROCK via integrin clustering and down-regulation of the Src-p190RhoGAP regulatory pathway.