AML1-FOG2 functions as a transcriptional repressor and recruits CtBP. (A) The reciprocal transcripts AML1-FOG2 (AF) and FOG2-AML1 (FA) can be detected in the bone marrow by RT-PCR. The AML1 transcript is also detected, whereas the FOG2 transcript is not detected. Control PCR reactions were performed in parallel using identical primer pairs and 50 ng plasmid DNA as template. (B) AML1-FOG2 represses the transcriptional activity of CBF. For each transfection, the graph depicts the mean and standard deviation for 6 independent replicates. The amount of each plasmid transfected is indicated in micrograms. CBFb refers to CBFβ. (C) AML1-FOG2 represses the transcriptional activity of GATA1. For each transfection, the graph depicts the mean and standard deviation for 3 independent replicates. The amount of each plasmid transfected is indicated in micrograms. (D) AML1-FOG2 and CtBP associate in a protein complex. 293 cells were transfected with either pCDNA3.1/V5-HisB (CTRL) or V5-epitope tagged AML1-FOG2 (AF). Molecular weights of markers are indicated in kilodaltons. (i) Immunoprecipitation of AML1-FOG2 with anti–V5 antibody specifically recovers CtBP. (ii) Immunoprecipitation of CtBP with anti–CtBP antibody specifically recovers AML1-FOG2. (iii) Control immunoprecipitation with anti-V5 shows that AML1-FOG2 is expressed after transfection and can be detected with anti–AML1 antibody. (iv) Control blot with anti–CtBP antibody shows equal amounts of endogenous CtBP in the protein lysates used for immunoprecipitation experiments.