Trib1 modulates super-enhancer activity and gene expression via C/EBPα degradation. (A) Trib1 hi cells were treated with the MEK1 inhibitor U0126 (10 μM) for 24 hours. Inhibition of ERK1/2 phosphorylation is evidenced by immunoblotting (left). Quantitative ChIP-PCR showing relative signals of H3K27ac to input DNA (right). The Chek1 locus was used as a negative control. (B) Quantitative RT-PCR showing mild downregulation of Erg and Spns2 expression by U0126 treatment (right). Expression of the Erg protein (arrow) was also diminished (top). (C) Cebpa was silenced by shRNA treatment using 2 different sequences in Trib1 null cells. The effect of shRNA was confirmed by immunoblotting (left). Quantitative ChIP-PCR showing relative signals of H3K27ac to input DNA (right). The Chek1 locus was used as a negative control. Three distinct loci for each super-enhancer were examined and the average accumulation in these 3 loci is indicated. (D) Quantitative RT-PCR showing upregulation of Erg, Spns2, Rgl1, and Pik3cd expression by Cebpa silencing (right). Expression of the Erg protein (arrow) is significantly upregulated (left). (E) Quantitative RT-PCR showing partial reduction of the Cebpa silencing effect on Erg, Spns2, Rgl1, and Pik3cd expression by human CEBPA p42, but not p30, expression (left). Western blot showing moderate decrease of the Erg protein by p42, but not p30, expression (center, top). Myc-tagged p42 or p30 expression (center, middle). Quantitative ChIP-PCR showing relative signals of H3K27ac to input DNA (right). The Chek1 locus was used as a negative control. *P < .05, **P < .01, ***P < .001. DMSO, dimethyl sulfoxide.