Interaction of Syntenin-1 and the NG2 Proteoglycan in Migratory Oligodendrocyte Precursor Cells

Abstract

Migration of oligodendrocyte precursors along axons is a necessary prerequisite for myelination, but little is known about underlying mechanisms. NG2 is a large membrane proteoglycan implicated in oligodendrocyte migration. Here we show that a PDZ domain protein termed syntenin-1 interacts with NG2 and that syntenin-1 is necessary for normal rates of migration. The association of syntenin-1 with NG2, identified in a yeast twohybrid screen, was confirmed by colocalization of both proteins within processes of oligodendroglial precursor cells and by coimmunoprecipitation from cell extracts. Syntenin-1 also colocalizes with NG2 in "co-capping" assays, demonstrating a lateral association of both proteins in live oligodendrocytes. RNA interference-mediated down-regulation of syntenin-1 in glial cells results in a significant reduction of migration in vitro, as does the presence of polyclonal antibody against NG2. Thus syntenin plays a role in the migration of oligodendroglial precursors, and we suggest that NG2-syntenin-1 interactions contribute to this.The NG2 proteoglycan is a type I membrane protein that is expressed by a variety of immature cells of several embryonic tissue origins including glia, muscle progenitor cells, and pericytes (1). In the central nervous system, expression of NG2 was originally thought to specify oligodendroglial progenitor cells, but more recent data suggest that NG2-expressing cells encompass a wider range of immature glial cells in white and gray matter. These include glia that make synaptic-like contacts with neurons in the hippocampus and cerebellum (2) and glial cells specifically associated with the nodes of Ranvier (3). Interestingly, many NG2-positive cells are both proliferative and motile or exhibit local process motility (4, 5). Antibodies to the NG2 extracellular domain inhibit migration of oligodendroglial progenitor cells and immature Schwann cells in in vitro migration assays (5, 6), and NG2 also plays a role in cell spreading in melanoma tumors, which express melanoma chondroitin sulfate proteoglycan (MCSP), the human ortholog of NG2 (7). Identifying the intracellular NG2-interacting proteins should aid in elucidating the function of this multidomain protein in migratory cells of the oligodendrocyte lineage.The intracellular domain of NG2 consists of the C-terminal 76 amino acids and has the PDZ (postsynaptic density-95/discs large/zona occludens-1) binding motif QYWV, which can interact with PDZ domain-containing proteins (8). To define relevant intracellular partners of the glycoprotein, we carried out a yeast two-hybrid screen using the complete intracellular domain of NG2 as bait. One of the proteins that we identified is syntenin-1 (also termed MDA-9). Syntenin-1 is a widely expressed PDZ protein that is often overexpressed in highly migratory metastatic tumors including melanoma (9).Here we demonstrate that syntenin is expressed by primary oligodendrocytes and show functional studies using the oligodendroglial precursor cell line Oli-neu. We provide biochemical, mor...

NG2 Regulates Directional Migration of Oligodendrocyte Precursor Cells via Rho GTPases and Polarity Complex Proteins

“…Homozygous NG2-EYFP knock-in mice (Karram et al, 2008) and wild-type littermates at the age of 3 months of either sex underwent a stab wound as described previously (Dimou et al, 2008). Briefly, a 1-1.5 mm long/0.7 mm deep stab wound was made using a V-lancet scalpel in the right sensorimotor cortex.…”

“…siRNA oligos were purchased from Qiagen and transfected into Oli-neu cells and primary oligodendrocytes in suspension with AMAXA electroporation system using protocol for primary neural cells. The targeted sequences of mouse cDNA were as followed: (Chatterjee et al, 2008). The nonsilencing siRNA siControl [5Ј-AATTCT CCGAACGTGTCACGT-3Ј] was used as a control.…”

A role for the NG2 proteoglycan in glioma progression

“…In this respect, syntenin-1 has been implicated in NG2-dependent oligodendrocyte progenitor migration. 23 …”

“…This implies that the site of cytoplasmic phosphorylation of NG2 can determine its ability to interact with cytoplasmic scaffolding proteins that anchor the proteoglycan (and its integrin binding partner) within specific membrane microdomains. MUPP1, 21 GRIP1, 22 syntenin-1 23 and ezrin 24 are possible candidates for such anchoring functions. In this respect, syntenin-1 has been implicated in NG2-dependent oligodendrocyte progenitor migration.…”

“…At the extreme C-terminus, the PDZ-binding motif QYWV mediates the interaction of NG2 with the multi-PDZ scaffolding proteins MUPP1, 21 GRIP1 22 and syntenin-1. 23 There are several cytoplasmic threonine residues, at least two of which are sites of functionally important phosphorylation. Thr-2256 is phosphorylated by PKCα, 24 while Thr-2314 is phosphorylated by ERK.…”