Hiwi expression correlates with DNA methylation.

<p>(A) IAP and Line1 transposon expression is decreased in Hiwi-MSCs. IAP and Line1 semi-quantitative PCRs were run for 25 cycles. Actin is a loading control. (B) Global DNA methylation is increased in Hiwi-MSCs. Error bars represent standard error. * = p<0.05 by Student's T Test (C) Global DNA methylation is decreased to non-detectable levels (ND) after 18 h treatment with 10 uM 5-azacytidine in Hiwi-MSCs. Lower doses of 5-azacytidine (including 1 uM) did not change DNA methylation levels. Error bars represent standard error * = p<0.05 by Student's T Test (D) Western blots of indicated proteins reveal increased expression of DNMT1, DNMT3a and MBP2 in Hiwi-MSCs. Actin is a loading control. (E) Treatment of Hiwi expressing MSCs with 0 uM, 5 uM, 10 uM, or 50 uM 5-azacytidine for 18 h restores IAP and Line1 transposon expression in a dose-dependent manner. IAP and Line1 quantitative RT-PCRs were run and parental MSC of each 5-azacytidine concentration were used to normalize the rest of the samples at that concentration. Error bars represent standard error. All experiments were performed in triplicate.