To determine whether the repressive role of OLIG2 is essential for myelination, a dominant-active form of OLIG2 (OLIG2 fused to 4X transactivation domain of VP16, referred to as OLIG2-VP64) (Supplementary Fig. 1b) was used to override endogenous OLIG2 in rat OPC in vitro. OLIG2-VP64, the dominant-active form of OLIG2, is reported in prior studies to abolish the transcriptional inhibition by transforming OLIG2 into transcriptional activation form15,21. Consistently, most of the differentially expressed genes (DEGs) were upregulated in cells transfected with OLIG2-VP64 (Supplementary Fig. 1c). Upon triiodothyronine (T3) stimulation for promoting differentiation, OPCs transfected with OLIG2-VP64 failed to exhibit morphological features of mOLs (Fig. 1c). Expression of Myelin marker proteins such as CNP or MBP was found in fewer cells expressing OLIG2-VP64 (Fig. 1d) and the expression levels of these marker genes were also decreased in these cells (Fig. 1e). Note that overexpressing VP64 alone had no appreciable effect on the OPC differentiation (Supplementary Fig. 1d). Transcriptomic analyses revealed that genes programmed to be inhibited by OLIG2 upon differentiation were upregulated in the OLIG2-VP64 overexpressed cells (Fig. 1f). To assess the
