Figure 8. Phosphorylation of Miz1 at Ser178 is required for its suppression of Cebpd transcription.
(a) MLE12–Miz1(WT) cells were treated with or without TNF for 1 h. Xpress-Miz1 was immunoprecipitated with anti-Xpress antibody and resolved by SDS-PAGE, followed by Coomassie Brilliant Blue staining. Xpress-Miz1 band was excised and subjected to MS/MS analysis by the Mass Spectrometry Core Facility at Yale University. (b) Cebpd luciferase reporter gene assays in MLE-12–Miz1(KD) cells transfected with WT Miz1 or the Miz1(S178A) mutant and then treated with or without TNF. (c,d,e) C/EBP-δ transcription and IL-6 production (c) as well as ChIP analysis of the recruitment of Miz1 (d) and HDAC1 (e) to the Cebpd promoter were determined in MLE12–Miz1(WT) and MLE-12–Miz1(S178A) cells treated with or without TNF. Data are representative of three independent experiments (b–e). *, P < 0.05 by Student’s t-test.