Integrin‐linked kinase regulates oligodendrocyte cytoskeleton, growth cone, and adhesion dynamics

Abstract

Integrin-linked kinase (ILK), a focal adhesion protein, brokers the link between cytoskeleton, cell membrane, and extracellular environment. Here, we demonstrate a role for ILK in laminin-2-mediated adhesion in primary murine oligodendrocytes (OLs) -with ILK loss leading to severe defects in process branching and outgrowth. These defects were partially recovered when the ILK-depleted OLs were instead grown on the non-integrin-activating substrate poly-L-lysine. Intriguingly, ILK loss on the neutral poly-L-lysine substrate led to swelling at the tips of OL processes, which we identified as enlarged growth cones. Employing the bloated ILK-depleted growth cones as template, we demonstrate the appearance of distinct cytoskeletal domains within OL growth cones bearing classic neuronal growth cone architecture. Further, microtubule organization was severely perturbed following ILK loss, with centripetal microtubule looping and failure to bundle occurring in a laminin-2-independent manner. Together, our work highlights differences in specific aspects of OL biology as driven by laminin-2-dependent or independent ILK governed mechanisms. We also reinforce the idea of OLs as growth cone bearing cells and describe the neuronal-like cytoskeleton therein. Finally, we demonstrate a role for ILK in OL growth cone maturation through microtubule regulation, the loss of which translates to decreased process length and myelin production capacity.

The oligodendrocyte growth cone and its actin cytoskeleton: A fundamental element for progenitor cell migration and CNS myelination

“…In further support of a role of the above pathway in specifically membrane spreading, OLGs deficient in ILK show a reduced capacity to enwrap neurites in a coculture system (Camara et al, ; O'Meara et al, ), and there is a developmentally transient reduction in the number of myelinated axons, again with a preferential effect on smaller diameter axons, in conditional Ilk knockout mice (Camara et al, ; O'Meara et al, ). Importantly, and consistent with a critical role of F‐actin disassembly during membrane spreading, aberrant F‐actin accumulation was seen when ILK‐deficient OLGs were differentiated on laminin2/merosin as a substrate (Michalski et al, ; O'Meara et al, ). It should be noted here, however, that ILK has additionally been shown to regulate OLG growth cones and their cytoskeleton by laminin2/merosin‐independent mechanisms (Elazar et al, ; Michalski et al, ).…”

“…Such individualized responses require specialized sensing features at the tips of outgrowing processes. In this regard, the tips of the leading processes of migratory OPCs have been shown to feature structural and functional similarities to neuronal growth cones (Jarjour & Kennedy, ; Schmidt et al, ; Simpson & Armstrong, ), and similar observations have been made for the tips of processes extending from differentiating OLGs (Asou, Hamada, & Sakota, ; Fox, Afshari, Alexander, Colello, & Fuss, ; Hardy & Friedrich Jr., ; Kachar, Behar, & Dubois‐Dalcq, ; Michalski, Cummings, O'Meara, & Kothary, ; Rumsby, Afsari, Stark, & Hughson, ; Song, Goetz, Baas, & Duncan, ; Zuchero et al, ). Thus, OLG/OPC growth cones emerge as a characteristic feature of OLG lineage cells.…”

“…Similar to OPCs, differentiating OLGs have been found to extend OLG growth cones at the distal tips of their multiple processes (Asou et al, ; Fox et al, ; Kachar et al, ; Michalski et al, ; Rumsby et al, ; Song et al, ; Zuchero et al, ). Given the postmigratory and premyelinating characteristics of these cells, the functional importance of their growth cones lies in surveying the local environment in search of axonal segments to be myelinated and/or inappropriate targets to be avoided.…”

“…Such permissive cues include extracellular matrix molecules, the best characterized of which is laminin‐2/merosin (Bechler et al, ). First, a number of in vitro studies identified laminin‐2/merosin as a permissive substrate for OLG growth cones on which filopodia formation, and process outgrowth and branching are augmented (Eyermann, Czaplinski, & Colognato, ; Fox et al, ; Lafrenaye & Fuss, ; Michalski et al, ). Collectively, these studies point toward a signaling pathway that is initiated by an interaction of laminin‐2/merosin with the nonintegrin extracellular matrix receptor dystroglycan, which is associated with point contact complexes of OLG filopodia and process branch points.…”

“…In the case of laminin2/merosin, however, it has additionally been proposed that for late‐stage differentiating OLGs, initial spreading is promoted at the OLG growth cone contact site with laminin2/merosin‐positive axons. The signaling pathway that has been implicated in this process involves the integrin receptor α6β1 and downstream activation of integrin‐linked kinase (ILK; Buttery & ffrench‐Constant, ; Chun et al, ; Colognato et al, ; Michalski et al, ). In vivo, reduced numbers of particularly smaller diameter myelinated axons have been reported during the developmental window of active myelination for mice in which a dominant‐negative form of integrin β1 is expressed in OLGs (Camara et al, ).…”