(A) RUNX1 phosphorylation by CBFβ and deletion variants of CBFβ-SMMHC. CBFβ, CBFβ-SMMHC, or deletion variants of CBFβ-SMMHC fusion proteins were co-expressed in 293T cells with RUNX1 and wild-type p300. Western blot analyses were performed using the indicated antibodies.
(B) p300 phosphorylation by CBFβ and deletion variants of CBFβ-SMMHC fusion proteins. Wild-type p300 or mutant p300 (p300 ΔSTP1,2,3) were transfected with RUNX1 and CBFβ, CBFβ-SMMHC, or the CBFβ-SMMHC deletion constructs. Immunoblotting with p300 antibody was performed with lysates from transiently transfected 293T cells.
(C) RUNX1 phosphorylation in human and mouse leukemia samples. ME1: a cell line derived from an inv(16)+ AML case (Yanagisawa et al., 1991), primary: primary leukemia cells from bone marrow of an inv(16)+ AML case (Liu et al., 1996). Both human cases contain the type A CBFB-MYH11 fusion. FL: full length CBFβ-SMMHC, D43: CBFβ-SMMHC d179-221, WT BM: wildtype whole bone marrow cells after erythrocyte lysis. Protein lysates were isolated from human and mouse leukemic cells and western blot analyses were performed using the indicated antibodies.