Permanent demyelination, the major pathology of multiple sclerosis, involves the disruption of axonal myelin and neuronal damage. Effective repair requires the repopulation of lesions with oligodendrocytes capable of differentiating and remyelinating damaged neurons. Mature oligodendrocytes originate from progenitor cells (OPCs), whose developmental pathways are poorly understood. Using DMA microarray analysis, we have established a developmental gene expression profile of EGFP+ oligodendrocyte lineage cells FACS-purified from postnatal day 2 (P2)-P30 CNP-EGFP mice. A gene sub-cluster identified by a peak of expression at P15 was found to contain the SoxF family members (Sox7/Sox17/Sox18), whose expression and function in oligodendrocytes are unexplored. Developmental expression of Sox17 displayed temporally-related coordinate clustering with myelin genes. In FACS-purified EGFP+ cells, Sox17 expression increased between P2 and P15, coincident with the onset of myelination and downregulation of cell cycle genes, and decreased thereafter. In white matter, Sox17 was expressed at higher levels in O4+ cells. In cultured OPCs, a transient increase of Sox17 expression correlated with i) increases in p21Cip1/Waf1, p27Kip1 and MBP expression, and ii) a subsequent decrease in proliferating cell nuclear antigen. siRNA-induced Sox17 downregulation increased OPC proliferation and decreased differentiation. Reporter assays showed that Sox17 can activate the MBP promoter. We plan first to characterize the expression and regulation of Sox17 in OPCs cultured under conditions that promote cell differentiation, to associate changes in Sox17 expression with specific stages of OPC development and the expression of cell cycle regulators. We will define a functional role for Sox17 in cell cycle regulation and OPC maturation by studying the effects of: i) Sox17 siRNA on OPC gene expression in culture by microarray analysis and ii) Sox17 overexpression in vivo in a CNP-Sox17-IRES-EGFP transgenic mouse strain. We will identify Sox17-responsive regions in the p21 and MBP gene promoters by reporter assays, and analyze DMA binding activity at these sites. Finally, we will characterize Sox17 expression in the oligodendrocyte lineage in vivo during normal development and during remyelination following chemical demyelination. Our proposed studies hope to shed light on crucial regulators of oligodendrocyte maturation that are potentially amenable to therapeutic intervention. ? ? ?
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