Role and regulation of Juxtanodin in actin cytoskeleton of Oligodendrocyte

Abstract

Oligodendrocytes are the myelinating cells of the central nervous system. They are derived from oligodendrocyte precursor cells which undergo a complex and precisely regulated timed program of proliferation, migration and differentiation finally to form the myelin, a multilamellar lipid/protein insulating layer. Myelin wraps around axons and enables nerve fibers propagating electric impulses rapidly by saltatory conduction. Failure to form a tightly wrapped myelin sheath (dysmyelination), or to maintain it (degeneration), results in delayed or disrupted signal transduction, most clearly illustrated in the CNS by the autoimmune disease Multiple Sclerosis.

While the exact mechanisms underlying the specialized wrapping stage of myelination have been little elucidated, considerably more is known about how the reorganization of actin cytoskeleton and the regulation of actin-binding proteins, coupled with those of tubulin cytoskeleton, are mediated by extracellular and intracellular signals during oligodendrocyte migration and differentiation.

Juxtanodin was firstly identified as an oligodendroglial protein by screening cell type-specific CNS genes. In amino acid sequence, it shared some similarity with ERM (ezrin, radixin and moesin) proteins. Previous studies in our lab showed that Juxtanodin expression paralleled temporally and spatially the onset of myelination in vivo. Furthermore, overexpression of Juxtanodin promoted arborization of cultured OLN-93 cells and primary oligodendrocytes. To further clarify molecular interaction, possible biochemical effects and mechanisms of activity regulation of Juxtanodin, we carried out a series of in vitro and in culture experiments in the current study.

It was found that JN could directly interact with actin and this interaction was mediated by the C-terminal F-actin binding domain, which comprised the last 14 amino acid of JN. Studies on the role of JN in actin dynamics showed that it could prevent F-actin depolymerization in vivo and in vitro, suggesting that JN was possibly involved in the F-actin based structures and behaviors of oligodendrocyte. As expected, JN over-expression in the cultured oligodendrocyte cell line dramatically induced the formation of F-actin-based cellular structures, such as filopodia at the cell edge and stress-fiber in the cytoplasm. JN also promoted cell spreading and inhibited cellular migration.

As expected, the activity of Juxtanodin for actin cytoskeleton should be precisely regulated and reversible in vivo, which was supported by the fact that JN partially co-localized with F-actin in oligodendrocyte in central nervous system. In the current study, the phosphorylation at its serine 278 site was demonstrated to abolish JN¿s effect on actin dynamics and actin-based structures/behaviors at the biochemical and cellular levels. Further study suggested that RhoA GTPase was possibly involved in the phosphorylation of serine 278.

Taken together, our results point to Juxtanodin as an actin cytoskeleton-stabilizing protein that plays active roles in migration, differentiation of oligodendrocytes and maintenance of the myelin sheath. The results also suggest phosphorylation modification and RhoA GTPase pathway as important mechanisms in the regulation of Juxtanodin functions.