Effect of GATA1 knockdown on gene expression of TAL1 and recruitment of members of the TEC and Pol II. (A) GATA1 protein levels in K562 cells in a representative GATA1 siRNA knockdown (KD^GATA1) and luciferase siRNA control (LUC) at 48 and 96 hours after transfection. The immunoblot was stained in Bradford reagent (Bio-Rad) and imaged to use as a loading control (also shown). GATA1 protein is ∼47 kDa (black arrow). (B) TAL1 transcript levels (with SEs) in GATA1 knockdown samples at 48 and 96 hours after transfection are shown relative to levels in luciferase control samples. mRNA levels for controls (ACTB and TUBB), GATA1, LDB1, TCF3, and GATA2 levels are also shown. Occupancies of GATA1 (C) or LDB1 (D) at the TAL1 erythroid enhancer (+51), the TAL1 stem cell enhancer (+20), and TAL1 promoter 1a (P^TAL1) 48 or 96 hours after transfection with siRNA for GATA1 or luciferase. Occupancies of TCF3 (E47 isoform) (E) or Pol II (F) at the TAL1 erythroid enhancer (+51), the TAL1 stem cell enhancer (+20), and TAL1 promoter 1a (P^TAL1) 96 hours after transfection with siRNA for GATA1 or luciferase. Positive control for Pol II occupancy was the promoter of TBP (P^TBP). For the bar diagrams in panels C-F, ChIP enrichments (log₂) are shown with SEs. Annotation of test and negative control regions is denoted in black and gray text, respectively. Luciferase (LUC) and GATA1 knockdown treated samples (KD^GATA1) are shown as gray and black bars, respectively, as per the key in panel C. Vertical gray arrows highlight the occupancy data for the +51 enhancer, the +20 stem cell enhancer, and the TAL1 promoter 1a (P^TAL1). The percentage (%) occupancies of each protein in the GATA1 knockdown relative to the levels found in the luciferase control are also shown. *P < .05; **P < .01; ***P < .001; ****P < .0001.