Role of Integrin-Linked Kinase in Nerve Growth Factor-Stimulated Neurite Outgrowth

Abstract

The role of integrin-linked kinase (ILK), a kinase that is involved in various cellular processes, including adhesion and migration, has not been studied in primary neurons. Using mRNA dot blot and Western blot analysis of ILK in rat and human brain tissue, we found that ILK is expressed in various regions of the CNS. Immunohistochemical and immunocytochemical techniques revealed granular ILK staining that is enriched in neurons and colocalizes with the beta1 integrin subunit. The role of ILK in neurite growth promotion by NGF was studied in rat pheochromocytoma cells and dorsal root ganglion neurons using a pharmacological inhibitor of ILK (KP-392) or after overexpression of dominant-negative ILK (ILK-DN). Both molecular and pharmacological inhibition of ILK activity significantly reduced NGF-induced neurite outgrowth. Survival assays indicate that KP-392-induced suppression of neurite outgrowth occurred in the absence of cell death. ILK kinase activity was stimulated by NGF. NGF-mediated stimulation of phosphorylation of both AKT and the Tau kinase glycogen synthase kinase-3 (GSK-3) was inhibited in the presence of KP-392 and after overexpression of ILK-DN. Consequently, ILK inhibition resulted in an increase in the hyperphosphorylation of Tau, a substrate of GSK-3. Together these findings indicate that ILK is an important effector in NGF-mediated neurite outgrowth.

Integrin-linked kinase activity regulates Rac- and Cdc42-mediated actin cytoskeleton reorganization via α-PIX

“…Additionally, our laboratory has demonstrated that treatment of cells with KP-392 results in decreased cell motility (Attwell et al, 2003;Tan et al, 2004), a process that is heavily influenced by the activation status of the Rho family of GTPases (Ridley et al, 2003). When the ILK 13-8 cells were pretreated with KP-392 (Figure 5a), there is a dose-dependent reduction in Rac activation (B65% inhibition) at concentrations previously described to inhibit ILK activity (Persad et al, 2001;Attwell et al, 2003;Mills et al, 2003;Tan et al, 2004). As expected, KP-392 treatment of these cells causes a substantial decrease (B90% inhibition) in the phosphorylation and activation of Akt (Figure 5a), suggesting that Rac, much like Akt, is activated in an ILK activity-dependent manner.…”

“…Our laboratory has previously characterized a small molecule inhibitor of ILK kinase activity (KP-392, formerly called KP-SD1). This highly selective ILK inhibitor is able to reduce the phosphorylation of ILK substrates such as GSK-3 and Akt (Persad et al, 2000(Persad et al, , 2001Mills et al, 2003;Tan et al, 2004). Additionally, our laboratory has demonstrated that treatment of cells with KP-392 results in decreased cell motility (Attwell et al, 2003;Tan et al, 2004), a process that is heavily influenced by the activation status of the Rho family of GTPases (Ridley et al, 2003).…”

An Essential Role for the Integrin-Linked Kinase–Glycogen Synthase Kinase-3β Pathway during Dendrite Initiation and Growth

“…We therefore asked whether ILK was required for the observed increase in GSK-3␤ phosphorylation, using the specific pharmacological ILK inhibitor QLT0254 (formerly KP-392; Mills et al, 2003;Yau et al, 2005). Neurons were established, pretreated for 1 h with or without the ILK inhibitor, and stimulated for 15 min with NGF plus KCl.…”

“…Integrin-linked kinase (ILK) is one signaling protein that has recently been implicated in axonal growth (Mills et al, 2003) but is not known to play a role in dendrite morphogenesis. ILK is a serine-threonine kinase that is activated in a phosphatidylinositol 3 (PI3)-kinase-dependent manner (Delcommenne et al, 1998) and that functions downstream of ␤-integrins and growth factor receptors to regulate cell growth, differentiation, and migration (Dedhar et al, 1999).…”

Genome-Wide Association and Exome Sequencing Study of Language Disorder in an Isolated Population

“…36 The protein encoded by ILK is 1 of the key regulators of neural stem cell astrocytic differentiation 37 and neurite outgrowth. 38 We also found that 7 (58%) of 12 individuals in the AZ population carried a known missense variant in CDH2 (18q12) that was found only at a 2% frequency in the 1000 Genomes data set. CDH2 codes for a major cadherin that is widely expressed prenatally in neural stem cells and supports their differentiation and migration, 39 regulating the laminar organization of the cortex.…”