NG2 cells in the central nervous system (CNS) produce mature myelinating oligodendrocytes (OL) and maintain their proliferating ability throughout life. NG2 is a chondroitin sulfate proteoglycan that is encoded by cspg4 gene, expressed in a various cell type including NG2 cells and pericytes in the CNS. Recent reports suggest that NG2 proteoglycan is required for NG2 cell proliferation or differentiation. Regardless of its importance in the CNS, the promoter / enhancer sequence of cspg4 in the genome and factors directly regulating NG2 expression remain unknown.
Here, we examined promoter / enhancer elements using mixed glial culture or chick embryos to analyze what factors are required for cspg4 regulation. We found that a promoter region of cspg4 gene, which is used in several previous reports, shows its non-specific activity in wide variety of cell type including neuron. We then identified a novel 1.4 kb intronic enhancer that specifically regulates cspg4 expression in a primary culture and in ovo electroporation assay. Further, we established transgenic mouse line that express GFP under the regulation of the intronic enhancer. In this mouse, GFP signals are specifically observed in oligodendrocyte lineage cells, while PDGFR-beta positive pericytes do not express GFP, suggesting that enhancer sequence of cspg4 expression is differentially regulated depending on cell type. In addition, we found that Sox9, Olig2, and Mash1 positively regulate the activity of cspg4 enhancer. These results elucidated the new regulatory sequence of cspg4 and its regulatory transcription factors, which provide significant insights into oligodendrocyte biology.