Integrin-linked kinase is localized to cell-matrix focal adhesions but not cell-cell adhesion sites and the focal adhesion localization of integrin-linked kinase is regulated by the PINCH-binding ANK repeats

Abstract

Integrin-linked kinase (ILK) is a ubiquitously expressed protein serine/threonine kinase that has been implicated in integrin-, growth factor- and Wnt-signaling pathways. In this study, we show that ILK is a constituent of cell-matrix focal adhesions. ILK was recruited to focal adhesions in all types of cells examined upon adhesion to a variety of extracellular matrix proteins. By contrast, ILK was absent in E-cadherin-mediated cell-cell adherens junctions. In previous studies, we have identified PINCH, a protein consisting of five LIM domains, as an ILK binding protein. We demonstrate in this study that the ILK-PINCH interaction requires the N-terminal-most ANK repeat (ANK1) of ILK and one (the C-terminal) of the two zinc-binding modules within the LIM1 domain of PINCH. The ILK ANK repeats domain, which is capable of interacting with PINCH in vitro, could also form a complex with PINCH in vivo. However, the efficiency of the complex formation or the stability of the complex was markedly reduced in the absence of the C-terminal domain of ILK. The PINCH binding defective ANK1 deletion ILK mutant, unlike the wild-type ILK, was unable to localize and cluster in focal adhesions, suggesting that the interaction with PINCH is necessary for focal adhesion localization and clustering of ILK. The N-terminal ANK repeats domain, however, is not sufficient for mediating focal adhesion localization of ILK, as an ILK mutant containing the ANK repeats domain but lacking the C-terminal integrin binding site failed to localize in focal adhesions. These results suggest that focal adhesions are a major subcellular compartment where ILK functions in intracellular signal transduction, and provide important evidence for a critical role of PINCH and integrins in regulating ILK cellular function.

Pinch2 is a novel regulator of myelination in the Central Nervous System

“…The IPP protein complex arises from the association of different proteins, including the exclusive binding of either Pinch1 or Pinch2 to Ilk (Li et al, 1999, Zhang et al, 2002a). Thus, the loss of one Pinch protein does not preclude the assembly of an alternative complex with the existing Pinch, Ilk and Parvin (Fukuda et al, 2003).…”

Chronic Venous Disease in Pregnant Women Causes an Increase in ILK in the Placental Villi Associated with a Decrease in E-Cadherin

“…Within the cell, ILK is crucial for anchoring the actin filaments (microfilaments) to the integrins, also mediating signal transduction between intracellular and extracellular compartments [18]. Regarding the mediation of ILK in extracellular signaling, it is widely accepted that ILK is located in the cell-matrix focal adhesions but not in cell-to-cell adhesion sites [40]. Thus, ILK is essential for the formation of focal complexes and actine anchoring, being also involved in the assembly of fibronectin-bound fibrillar adhesions [41].…”

Integrin-linked kinase (ILK): the known vs. the unknown and perspectives

“…Early studies showed that deletion of ANK1 preventing PINCH from binding to ILK abolishes, in turn, the docking of ILK to FAs [ 4 ]. Going further, the fourth LIM domain of PINCH facilitates interaction with Nck-2, enabling the formation of a complex between ILK and Nck-2 and therefore create a physical link between the integrin-mediated ILK signaling pathway and growth factor-mediated or small GTPase-mediated signaling pathways [ 84 ].…”

“…Although ILK was initially described as a serine/threonine–protein kinase, there had still been considerable controversy surrounding the issue of the exact molecular mechanism of signal transduction by ILK [ 2 ] until it was proven that ILK is a pseudokinase [ 3 ]. It has been primarily detected at focal adhesion (FA) sites [ 4 ], confirmed by co-localization with focal adhesion markers like vinculin and paxillin [ 5 ]. Subsequently, association with its specific binding partners, such as particularly interesting cys-his-rich protein (PINCH) [ 4 ] and β-Parvin [ 6 , 7 ] within FAs has been revealed.…”

“…It has been primarily detected at focal adhesion (FA) sites [ 4 ], confirmed by co-localization with focal adhesion markers like vinculin and paxillin [ 5 ]. Subsequently, association with its specific binding partners, such as particularly interesting cys-his-rich protein (PINCH) [ 4 ] and β-Parvin [ 6 , 7 ] within FAs has been revealed. ILK combines functions of a signal transductor and a scaffold protein by interacting with the cytoplasmic domains of the β1 and β3 subunits of integrin receptors, then facilitating further protein recruitment within the ILK–PINCH–Parvin complex (IPP) [ 8 – 11 ].…”