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Fig. 4 | Epigenetics & Chromatin

Fig. 4

From: The pioneer factor activity of c-Myb involves recruitment of p300 and induction of histone acetylation followed by acetylation-induced chromatin dissociation

Fig. 4

Interaction with both H3 and p300 is essential for c-Myb-dependent activation of a chromatin-embedded gene. a HD-11 cells were transfected with plasmids encoding full-length HA-tagged c-Myb, c-Myb D152V and c-Myb M303V. Total RNA was isolated 24 h after transfection, and mim-1 expression was measured by qRT-PCR relative to the reference gene hprt. The western blots were analysed using anti-c-Myb and anti-GAPDH primary antibodies. b Co-immunoprecipitation was performed with COS-1 cells transfected with plasmids encoding full-length HA-tagged c-Myb, c-Myb D152V and p300-Myc and lysed 24 h after transfection. Immunoprecipitation was performed using anti-p300 antibody (SC-585, Santa Cruz Biotechnology), and the proteins were detected by western blotting using anti-HA primary antibody (H6908, Sigma-Aldrich). 5% of total transfected cell lysate was loaded as reference. c HD-11 cells were transfected with plasmids encoding full-length HA-tagged c-Myb, c-Myb D152V, c-Myb M303V, c-Myb D152V-M303V and p300-Myc. Total RNA was isolated 24 h after transfection, and mim-1 expression was measured by qRT-PCR relative to the reference gene hprt. The western blots were analysed using anti-HA and anti-GAPDH primary antibodies. The qRT-PCR results in a and c are presented as mean ± SEM of three independent biological replicates. Significance was evaluated by unpaired, two-tailed t tests on selected pairs and indicated with p values (*p < 0.05; **p < 0.01; ns p > 0.05)

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