Fig. 2: EGF increases CHD6 stability.
a Ten cancer-related pathways that were significantly associated with genes affected by CHD6 in CRC (GSE2109 and GSE14333). Hallmark pathways emerged following IPA ‘core analysis’. Enrichment scores were displayed as −log10 (P value) by Fisher’s exact test. b GSEA plot of AKT signaling pathway signature correlated with CHD6 highly related genes. Normalized enrichment score (NES) and nominal P value of correlation were shown. c Representative immunoblot analysis of the indicated proteins in HCT116 and DLD-1 cells treated with EGF (100 ng/mL) at the indicated time points. d CHD6 mRNA levels in the indicated cells treated with EGF (100 ng/mL) for the indicated hours. ns, not significant. e Representative immunoblot analysis of the indicated proteins in HCT116 cells treated with MK2206 (5 μM) at different time points. f Representative immunoblot analysis of the indicated proteins in HCT116 cells treated with or without MK2206, followed by EGF treatment. g Representative immunoblots showing CHD6 protein turnover rate in HCT116 cells treated with cycloheximide (CHX, 60 μg/mL), in the presence or absence of EGF (100 ng/mL) treatment (top). Quantification of g (bottom). IOD, integrated optical density. The relative density of CHD6 was normalized to Vinculin and then normalized to the t = 0 control. h Representative immunoblots showing ubiquitination of Flag-tagged CHD6, under EGF (100 ng/mL) treatment in 293T cells. Cells were treated with MG132 (10 μM) 6 h before harvest. The cell lysates were pulled down (PD) with nickel beads (Ni-NTA) and immunoblotted with the indicated antibodies. WCL whole cell lysate.
