Cell-Surface Glycoprotein of Oligodendrocyte Progenitors Involved in Migration

Abstract

Myelination by oligodendrocytes in the CNS involves the migration to and recognition and ensheathment of axons. These distinct developmental phases of myelination are assumed to involve the interplay of a precisely regulated set of cell adhesion molecules expressed by both neurons and glial cells. These molecules remain largely unelucidated. In this paper we have identified a large (330 kDa) glycoprotein expressed by murine oligodendrocyte progenitor cells in vitro and in vivo that is downregulated as oligodendrocytes mature. Antigen-positive oligodendrocyte progenitor cells purified by panning develop into myelin-associated glycoprotein-positive oligodendrocytes and also adhere to cultured neurons. Polyclonal antibodies directed against the protein reduce the migration of oligodendrocyte progenitor cells. The observations suggest that the AN2 antigen may play a role in early stages of myelination.

Origin of Oligodendrocytes in the Subventricular Zone of the Adult Brain

“…After blocking in PBS containing 10% of normal goat serum for 2 h at room temperature, cells or tissue sections were incubated with primary antibodies overnight at 4°C in the blocking solution. The following primary antibodies were used: mouse monoclonal anti-␤-tubulin (Tuj1) (1:500; Berkeley Antibody Company, Berkeley, CA), anti-O 4 (1:50; Chemicon, Temecula, CA), anti-GFAP (1:500; Chemicon), anti-polysialylated neural cell adhesion molecule (PSA-NCAM) (1:400; AbCys, Annecy, France), chicken monoclonal anti-GFP (1:1000; Aves Lab,Tigard, OR), rabbit polyclonal anti-oligodendrocyte lineage transcription factor 2 (Olig2) (1:5000) (Ligon et al, 2004), antiplatelet-derived growth factor receptor ␣ (PDGFR␣) (1:50; Santa Cruz Biotechnology, Santa Cruz, CA), for identification of NG2 we used AN2 (1:1000) (Niehaus et al, 1999) and anti-NG2 (Chemicon) anti-myelin basic protein (MBP) (1:100; Chemicon), and anti-CNPase (1:500; Chemicon). For fluorescent ICC, cells, tissue sections, or whole mounts were washed in PBS, incubated with the appropriate secondary antibodies [all cyan at 1:400 (Jackson ImmunoResearch, West Grove, PA); all Alexa at 1:1000 (Invitrogen, Carlsbad, CA)] in blocking solution for 2 h at room temperature [cyanine 2 (Cy2) anti-mouse IgG, Cy3 anti-mouse IgM, Cy3 anti-rabbit IgG, Alexa-488 anti-chicken IgG, or Alexa-564 antirabbit IgG], and washed in PBS.…”

Segregated Expression of AMPA-Type Glutamate Receptors and Glutamate Transporters Defines Distinct Astrocyte Populations in the Mouse Hippocampus

“…A second series of confocal imaging used double labeling against S100␤ and AN2 (Niehaus et al, 1999), the mouse homolog of NG2. Surprisingly, in approximately one-third of the green fluorescent cells, AN2/NG2 and S100␤ were coexpressed (n ϭ 41/109; 38%) (Fig.…”

Compartmentation of Fyn Kinase with Glycosylphosphatidylinositol-anchored Molecules in Oligodendrocytes Facilitates Kinase Activation during Myelination

“…As an additional control, monoclonal antibody to the 330-kDa cell surface glycoprotein AN2, which is expressed by these cells (31), was used in similar cross-linking experiments (Fig. 4B).…”