Chemical Inhibition of Histone Deacetylases 1 and 2 Induces Fetal Hemoglobin through Activation of GATA2
Fig 7
GATA2 knockdown attenuates HBG induction by ACY-957.
(A) Experimental design. BM cells were cultured for 3 days in CS1 expansion media, and then infected with lentivirus containing a control hairpin that does not target any human gene (shCtrl) or with two different GATA2 targeting hairpins (shG2-1, shG2-2). Infected cells were selected by addition of puromycin to the culture for 2 days. Following removal of puromycin at day 0, cells expressing each hairpin were cultured for 4 days in CS1 expansion media with 1 μM ACY-957 or vehicle control. (B) GATA2 mRNA levels were measured by QPCR and expressed relative to ACTB mRNA (mean ± SD, n = 2 QPCR replicates for each of n = 2 infection replicates). (C) GATA2 protein levels at day 4. Values represent GATA2 protein levels normalized to ACTB and relative to vehicle treated shCtrl. (D) HBG mRNA levels were measured by QPCR and expressed relative to ACTB mRNA (mean ± SD, n = 2 QPCR replicates for each of n = 2 infection replicates). For panel ‘D’ P-values were calculated on day 4 using a two-tailed t test, ***P<0.0005.