Fig 2.

SET domain-dependent JAK2 downregulation by G9a. (A and B) K562 cells were transfected with pcDNA3 as a control and the indicated DNA constructs. Real-time PCR and Western blot analyses were performed using JAK2-specific primers and antibody. The expression of each construct is shown by Western blot analysis. (C and D) K562 cells were transfected with two different G9a shRNAs. JAK2 expression levels were confirmed by real-time PCR and Western blot analyses. (E and F) HL-60 cells were treated with 1 μM ATRA or DMSO and harvested 72 h later. G9a and Suv39h1 expression levels were confirmed in differentiated HL-60 cells via real-time PCR and Western blot analyses. (G and H) G9a and JAK2 expression patterns as a function of time after treatment with 30 μM hemin in K562 cells. (I and J) K562 cells were transfected with pEGFP-C1, pEGFP-G9a, and pEGFP-G9a-ΔSET. JAK2 expression levels were confirmed via real-time PCR and Western blot analyses. β-Actin was used as a loading control. All data are shown as means ± SD (n = 3). *, P < 0.05; **, P < 0.01; ***, P < 0.001. Results of the Western blot analysis were quantified and normalized to the level of β-actin.