ILK supports RhoA/ROCK-mediated contractility of human intestinal epithelial crypt cells by inducing the fibrillogenesis of endogenous soluble fibronectin during the spreading process

Abstract

Background: Fibronectin (FN) assembly into an insoluble fibrillar matrix is a crucial step in many cell responses to extracellular matrix (ECM) properties, especially with regards to the integrin-related mechanosensitive signaling pathway. We have previously reported that the silencing of expression of integrin-linked kinase (ILK) in human intestinal epithelial crypt (HIEC) cells causes significant reductions in proliferation and spreading through concomitantly acquired impairment of soluble FN deposition. These defects in ILK-depleted cells are rescued by growth on exogenous FN. In the present study we investigated the contribution of ILK in the fibrillogenesis of FN and its relation to integrin-actin axis signaling and organization. Results: We show that de novo fibrillogenesis of endogenous soluble FN is ILK-dependent. This function seemingly induces the assembly of an ECM that supports increased cytoskeletal tension and the development of a fully spread contractile cell phenotype. We observed that HIEC cell adhesion to exogenous FN or collagen-I (Col-I) is sufficient to restore fibrillogenesis of endogenous FN in ILK-depleted cells. We also found that optimal engagement of the Ras homolog gene family member A (RhoA) GTPase/Rho-associated kinase (ROCK-1, ROCK-2)/myosin light chain (MLC) pathway, actin ventral stress fiber formation, and integrin adhesion complex (IAC) maturation rely primarily upon the cell's capacity to execute FN fibrillogenesis, independent of any significant ILK input. Lastly, we confirm the integrin α5β1 as the main integrin responsible for FN assembly, although in ILK-depleted cells αV-class integrins expression is needed to allow the rescue of FN fibrillogenesis on exogenous substrate. Conclusion: Our study demonstrates that ILK specifically induces the initiation of FN fibrillogenesis during cell spreading, which promotes RhoA/ROCK-dependent cell contractility and maturation of the integrin-actin axis structures. However, the fibrillogenesis process and its downstream effect on RhoA signaling, cell contractility and spreading are ILK-independent in human intestinal epithelial crypt cells.

Fibronectin adsorption on oxygen plasma-treated polyurethane surfaces modulates endothelial cell response

“…33 In literature, an increased HFN7.1 antibody binding to unfolded FN has been described. 14,59 In our study, especially samples A and D showed a high level of accessible cell binding domain indicating an extended FN conformation. However, our data suggest that the accessibility of the cell-binding-domain is not necessary and is solely associated with the conformational change, but also with the orientation of the protein.…”

Cell migration guided by long-lived spatial memory

“…In vivo, motile cells integrate various inputs from the extra-cellular matrix, and, then, adjust their migration mode 61 . Gagné et al 62 showed that ILK potentiated intestinal epithelial cell spreading and migration by allowing fibronectin fibrillogenesis. The assembly of new roads built up by the cells themselves can be also crucial in the context of collective cell migration where the remodelling of ECM by leader cells can provide a path for follower cells 63 .…”

“…Several works have shown that cells are able to produce and to remodel their extra-cellular matrix 35,37,62,[64][65][66] . In this study, we have shown that cells actually deposit fibronectin in the form of small puncta but other ECM components may play a role.…”

Cell migration driven by long-lived spatial memory

“…Two questions remain open: the nature of the footprint and the physical mechanism by which the cell is attracted back. Several works have shown that cells are able to produce and to remodel their extra-cellular matrix 35,61,63,64,65,66 . In this study, we have shown that cells actually deposit fibronectin in the form of small puncta but other ECM components may play a role.…”

“…Gagné et al . 61 showed that ILK potentiates intestinal epithelial cell spreading and migration by allowing fibronectin fibrillogenesis. The assembly of new roads built up by the cells themselves can be also crucial in the context of collective cell migration where the remodeling of ECM by leader cells can provide a path for follower cells 62 .…”