Figure 7. PRMT4 and Mi2 are important regulators of proliferation and differentiation function in human erythropoiesis.

A: PRMT4 and Mi2 have similar effects on cell cycle progression to c-Myb. K562 cells were transfected with siRNAs targeting c-Myb, PRMT4, Mi2α and Mi2β or with control siRNA (si ctrl). The DNA content of propidium iodide (PI)-stained cells was measured by flow cytometry (FACS). From each knockdown the percentage of cells in sub-G1, G1 and G2/M phase was determined. Shown are the changes in percentage (%) relative to the si ctrl condition. A representative data set is depicted. B, C: PRMT4 and Mi2 have similar pro-proliferative functions to those of c-Myb. K562 cells were transfected with siRNAs, as in A. After 3 days of transfection, cells were cultured in methylcellulose for 2 days and stained with INT (Iodonitrotetrazolium chloride) for 5 days. (B) Representative pictures of the colonies were taken for each knockdown condition at the same magnification. (C) For quantification, colonies larger than 0.1 mm were counted. The mean of colony numbers was calculated from three independent experiments and error bars are indicated accordingly. D: Depletion of PRMT4 and Mi2 recapitulates the pro-differentiating effects of c-Myb knockdown. K562 cells were transfected with the indicated plasmids for 3 days. Cells were then re-seeded and either left untreated or treated with 30 µM hemin for 3 days. Subsequently, benzidine staining of differentiated cells was performed. The mean percentage (%) of benzidine-positive K562 cells was determined from triplicate countings and error bars are indicated accordingly.