Figure 5.

Histone demethylase Jhdm2a facilitates Oct4 reactivation during ESC-induced reprogramming. (A): EST profiles of histone demethylases. EST counts (transcripts per million) were collected from National Center for Biotechnology Information Unigene expression resources (http://www.ncbi.nlm.nih.gov/sites/entrez) and plotted against a panel of currently identified histone demethylases. Distributions of EST for individual demethylases across different developmental stages are shown. (B): Expression of Jhdm2a in ESCs and adult neural stem cells/progenitors (NSCs). Shown are quantitative real-time polymerase chain reaction analyses of the expression levels of Jhdm2a in Z-Red ESCs, CIPOE NSCs, and CIPOE NSCs with virally transduced Jhdm2a WT or H1120Y enzymatically inactive mutant. mRNA abundance is normalized to the levels of Z-Red cells, and values represent mean ± SEM (n = 3; *, p < .01, Student’s t test). (C): Ectopic expression of Jhdm2a, but not the enzyme-inactive mutant, induces global loss of H3K9 dimethylation in CIPOE NSCs, as indicated by arrows. Scale bar = 20 μm. (D): Summary of reprogramming frequencies. Shown are results from cell fusion experiments between Z-Red and CIPOE, CIPOE-J2WT, or CIPOE-J2HY cells. Values represent mean ± SEM (n = 3; *, p < .01, Student’s t test). (E): Summary of reprogramming efficacy. Cumulative distribution plots of DsRed⁺ population with graded EGFP fluorescence intensities are shown for comparison of the reprogramming efficacy of Z-Red ESCs with CIPOE-J2 or CIPOE-J2HY cells at day 4 after cell fusion. Values represent mean ± SEM (n = 3; *, p < .01, Kolmogorov-Smirnov test). Abbreviations: DAPI, 4′,6-diamidino-2-phenylindole; EGFP, enhanced green fluorescent protein; EST, expressed sequence tag; H3K9diM, histone H3 lysine 9 dimethylation; TPM, transcripts per million; WT, wild-type.