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. 2016 Apr 13;11(4):e0153767. doi: 10.1371/journal.pone.0153767

Fig 6. Overexpression of GATA2 induces HBG and suppresses HBB in erythroid progenitors.

Fig 6

(A) Experimental design. BM cells were cultured for 3 days in CS1 expansion media, and then infected with lentivirus containing GFP (oeCtrl) or full length GATA2 transcript (oeG2). Infected cells were selected for by addition of puromycin to the culture for 2 days. Cells were then shifted to CS1 differentiation media for the remainder of the experiment. (B) GATA2 mRNA levels in oeG2 cells expressed relative to oeCtrl (mean ± SD, n = 2 QPCR replicates for each of n = 2 infection replicates). (C) GATA2 protein levels at day 5 of differentiation. Values represent GATA2 protein levels relative to ACTB and then normalized to oeCtrl. (D) HBG mRNA levels during differentiation of oeCtrl and oeG2 cells (mean ± SD, n = 2 QPCR replicates for each of n = 2 infection replicates). HBG mRNA (%) = [HBG/(HBB+HBD+HBG+HBE)]*100. (E) Samples from ‘D’ with each β-like globin transcript expressed relative to ACTB mRNA. (F) TFRC and GYPA cell surface expression at day 5 of differentiation. (G) Total β-like globin mRNA (sum of HBB, HBD, HBG, and HBE) in samples from ‘D’. For panels ‘D’, ‘E’, and ‘G’, P-values were calculated on day 5 using a two-tailed t test. *P<0.05, **P<0.005, ***P<0.0005 for oeG2 compared to oeCtrl. Data is representative of experiments performed using cells from three independent donors.