Integrin-linked kinase, a novel component of the cardiac mechanical stretch sensor, controls contractility in the zebrafish heart

Bendig, Garnet; Grimmler, Matthias; Huttner, Inken G.; Wessels, G; Dahme, Tillman; Just, Steffen; Trano, Nicole; Katus, Hugo A.; Fishman, Mark C.; Rottbauer, Wolfgang

doi:10.1101/gad.1448306

Cited by 189 publications

(178 citation statements)

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“…renal agenesis during kidney development (21). One can similarly explain two other phenotypic single amino acid substitution mutations identified in ILK KD, L308P that causes a lethal heart failure in zebrafish (8) and A262V that causes a severe dilated cardiomyopathy in humans (9). As shown in supplemental Fig.…”

Section: Discussionmentioning

confidence: 78%

Biochemical, Proteomic, Structural, and Thermodynamic Characterizations of Integrin-linked Kinase (ILK)

Fukuda

Knight

Piszczek

et al. 2011

Journal of Biological Chemistry

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Integrin-linked kinase (ILK) is one of the few evolutionarily conserved focal adhesion proteins involved in diverse cell adhesion-dependent physiological and pathological responses. Despite more than a decade of studies and extensive literature, the kinase function of ILK is controversial. ILK contains a highly degraded kinase active site but it has been argued that ILK may be an unusual manganese (Mn)-dependent serine-threonine kinase that targets specific substrates such as glycogen synthase kinase-3␤ (GSK-3␤). In this study, we have tackled this issue by a systematic bottom-up biochemical, proteomic, structural, and thermodynamic analysis of ILK. We show that recombinant ILK from either bacteria or mammalian cells exhibits no kinase activity on GSK-3␤ in the presence of either Mn 2؉ or the conventional kinase co-factor Mg 2؉ . A comprehensive and unbiased whole cell-based kinase assay using entire mammalian CG-4 and C2C12 cell lysate did not identify any specific ILK substrates. High resolution crystallographic structure analysis further confirmed that the Mn-bound ILK adopts the same pseudo active site conformation as that of the Mg-bound ILK. More importantly, thermodynamic analysis revealed that the K220M mutation, previously thought to inactivate ILK by disrupting ATP binding, significantly impairs the structural integrity and stability of ILK, which provides a new basis for understanding how this mutation caused renal agenesis, a failure of fetal kidney development. Collectively, our data provide strong evidence that ILK lacks intrinsic kinase function. It is a bona fide pseudokinase that likely evolved from an ancestral catalytic counterpart to act as a distinct scaffold to mediate protein-protein interactions during focal adhesion assembly and many other cellular events.The adhesion of cells to the extracellular matrix (ECM) 3 is primarily mediated by heterodimeric integrin transmembrane receptors. Upon activation, the integrin extracellular domain adheres to the ECM by binding to numerous ligands such as fibronectin and fibrinogen. However, for cells to firmly adhere to the ECM, integrins must connect to the backbone of the cell, the actin cytoskeleton, via its cytoplasmic tails (CTs) (1-2). Such connection is mediated by the supramolecular focal adhesion (FAs) complex. The reassembly/disassembly of FAs can lead to dynamic cell adhesion processes such as cell migration and spreading, which are fundamental for a variety of physiological responses. Over the past decades, major effort has been made to search for proteins involved in FA assembly and regulation. Integrin-linked kinase (ILK) was discovered in 1996 as a binding partner of integrin ␤ CTs (3) and was found to critically regulate FAs (4). Genetic and genomic analyses have established that ILK is one of the few essential and evolutionarily conserved proteins coupling integrins to actin (5-7), and its dysregulation has been linked to major human diseases such as heart failure (8 -10) and cancer (11). ILK has a molecular mass of ϳ51 kDa containi...

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Section: Discussionmentioning

confidence: 78%

Biochemical, Proteomic, Structural, and Thermodynamic Characterizations of Integrin-linked Kinase (ILK)

Fukuda

Knight

Piszczek

et al. 2011

Journal of Biological Chemistry

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“…8 Cardioprotective protein kinase B is an important target of ILK phosphorylation: the cardiomyopathy resulting from loss of ILK phosphorylation in PINCH-inactivated zebrafish is reversed by overexpression of constitutively activated protein kinase B. 16 ILK is upstream to a wide range of protein kinases involved in cell growth and cardiac hypertrophy, including protein kinase B, glycogen synthase kinase-3-β, extracellular signal-related kinase, and mammalian target of rapamycin 2 and ILK deletion causes severe cardiomyocyte apoptosis. 17 ILK is upregulated in human cardiac hypertrophy caused by left ventricular outflow obstruction, along with a wide range of hypertrophic genes.…”

Section: Functional Importance Of Ilkmentioning

confidence: 99%

Loss of Cardiomyocyte Integrin-Linked Kinase Produces an Arrhythmogenic Cardiomyopathy in Mice

Quang

Maguy

et al. 2015

Circ: Arrhythmia and Electrophysiology

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Background— Integrin-linked kinase (ILK), a serine/threonine protein kinase, has roles in cell signaling and molecular scaffolding. ILK mutation/deletion causes cardiomyopathic phenotypes, but the functional and electrophysiological features have not been characterized. This study investigated the structural, functional, ion channel, and electrophysiological changes associated with cardiomyocyte-directed ILK deletion in mice. Methods and Results— Adult mice with cardiomyocyte-directed ILK knockout were compared with littermate controls. Knockout mice showed markedly increased mortality, with sudden death beginning after 5 weeks and 100% mortality at 18 weeks. In 10-week-old knockout mice, spontaneous and inducible ventricular tachyarrhythmias were common, occurring in 60% and 86%, respectively, and absent in controls ( P <0.001, P <0.05 versus knockout mice). Ventricular refractoriness was prolonged, along with both QRS and QT interval. Action potentials were prolonged and displayed triggered activity. A wide range of ion currents were downregulated, including total, fast and slow components of transient outward K + current and inward rectifier K + current, along with corresponding ion channel subunit genes, providing a plausible explanation of action potential prolongation. At 5 weeks, only voltage-dependent K + currents were reduced, possibly related to direct ILK-Kv4.2 subunit interactions. Action potentials were prolonged, but no arrhythmias or cardiac dysfunction were noted. Structural remodeling was prominent at 10 weeks: connexin-43 was downregulated and redistributed to lateral cell margins, and left ventricular fibrosis occurred, with a strong regional distribution (predominating in the basal left ventricle). Conduction was slowed. High-throughput quantitative polymerase reaction gene-expression studies in 10-week-old ILK knockout showed upregulation of structural, remodeling and fibrosis-related genes, and downregulation of a wide range of ion channel and transporter subunits. Conclusions— Cardiomyocyte ILK deletion produces a lethal arrhythmogenic cardiomyopathy associated with important ion channel and structural remodeling.

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“…Integrin-linked kinase (ILK) is highly expressed in the sarcomeric Z-disk costamere (region of close intermyocyte linkage) and is a critical component of the protein scaffold required for integrated force transduction during normal cardiac function [10][11][12] . Indeed, ILK mutations have been reported in human DCM 1,13 and conditional gene deletion of Ilk leads to neonatal cardiomyopathy in mouse 14 .…”

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confidence: 99%

Integrin-linked kinase mediates force transduction in cardiomyocytes by modulating SERCA2a/PLN function

Traister

Aafaqi

et al. 2014

Nat Commun

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Human dilated cardiomyopathy (DCM) manifests as a profound reduction in biventricular cardiac function that typically progresses to death or cardiac transplantation. There is no effective mechanism-based therapy currently available for DCM, in part because the transduction of mechanical load into dynamic changes in cardiac contractility (termed mechanotransduction) remains an incompletely understood process during both normal cardiac function and in disease states. Here we show that the mechanoreceptor protein integrin-linked kinase (ILK) mediates cardiomyocyte force transduction through regulation of the key calcium regulatory protein sarcoplasmic/endoplasmic reticulum Ca 2 þ ATPase isoform 2a (SERCA-2a) and phosphorylation of phospholamban (PLN) in the human heart. A non-oncogenic ILK mutation with a synthetic point mutation in the pleckstrin homology-like domain (ILK R211A ) is shown to enhance global cardiac function through SERCA-2a/PLN. Thus, ILK serves to link mechanoreception to the dynamic modulation of cardiac contractility through a previously undiscovered interaction with the functional SERCA-2a/PLN module that can be exploited to rescue impaired mechanotransduction in DCM.

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Integrin-linked kinase, a novel component of the cardiac mechanical stretch sensor, controls contractility in the zebrafish heart

Cited by 189 publications

References 50 publications

Biochemical, Proteomic, Structural, and Thermodynamic Characterizations of Integrin-linked Kinase (ILK)

Biochemical, Proteomic, Structural, and Thermodynamic Characterizations of Integrin-linked Kinase (ILK)

Loss of Cardiomyocyte Integrin-Linked Kinase Produces an Arrhythmogenic Cardiomyopathy in Mice

Integrin-linked kinase mediates force transduction in cardiomyocytes by modulating SERCA2a/PLN function

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