Epigenetic Regulation Of E-cadherin Expression By The Histone Demethylase UTX In Colon Cancer Cells

BackgroundDecreased epithelial cadherin(E-cadherin)gene expression,a hallmark of epithelial-mesenchymal transition(EMT),is essential for triggering metastatic advantage of colon cancer.Most studies investigating the mechanisms underlying EMT focus on genetic mechanism;however,the epigenetic mechanism by which E-cadherin is regulated in EMT process is less well understood.In this paper,we have identified ubiquitously transcribed tetratricopeptide repeat on chromosome X(UTX),a histone demethylase responsible for dior tri-methylated histone 3 lysine 27(H3K27me2/3),played a potential role in regulating E-cadherin expression in human colon cancer cells HCT-116.We demonstrated that inactivation of UTX down-regulated E-cadherin gene expression,while overexpression of UTX did the opposite.Notably,overexpression of UTX resulted in inhibition of migration and invasion of HCT-116 cells.We found that UTX demethylated H3K27me3,a histone transcriptional repressive mark,leading to decreased H3K27me3 at the E-cadherin promoter.We showed that UTX interacted with the histone acetyltransferase(HAT)protein CBP and recruited it to the E-cadherin promoter resulting in increased H3K27 acetylation(H3K27ac),a histone transcriptional active mark.UTX positively regulates E-cadherin expression through coordinated regulation of H3K27 demethylation and acetylation,switching the transcriptional repressive state to the transcriptional active state at the E-cadherin promoter.In conclusion,these results conclude that UTX may play a potential role in regulation of E-cadherin gene expression in HCT-116 cells and may pave a way for developing new treatment strategies against colon cancer metastasis.ObjectiveTo investigate the regulatory effect of UTX on E-cadherin expression in human colon cancer cells,and reveal the underlying mechanism.Materials and methods1.Human colon cancer cell line HCT-116 was used for phenotypic study.2.UTX plasmid and siRNA were used for functional study.3.Chromatin immunoprecipitation(Ch IP)and co-immunoprecipitation(Co IP)were used for molecular mechanism study.Results1.UTX positively regulates E-cadherin expression.Eight of fourteen histone methyltransferases and demethylases had higher expressions compared to the rest of enzymes in HCT-116 cells.Inactivation of UTX significantly suppressed E-cadherin mRNA expression.Both UTX and E-cadherin protein levels were significantly repressed by UTX-siRNA as assessed by immunoblotting.And overexpression of UTX increased E-cadherin m RNA and protein expressions.2.UTX inhibits the migration and invasion capacity of human colon cancer cells.Inactivation of UTX significantly increased the migratory potential and invasiveness capacity of HCT-116 cells.Whereas,overexpression of UTX did the opposite.3.H3K27me3 is the downstream target of UTX in HCT-116 cells.Loss of UTX expression induced to a significant increase in H3K27me3.On the contrary,overexpression of UTX received the function of demethylation on H3K27me3.Downregulation of UTX enhanced the binding of H3K27me3 at the E-cadherin promoter.In contrast,overexpression of UTX reduced the binding of H3K27me3 at the E-cadherin promoter.4.UTX inhibits EZH2/SUZ12 binding at the E-cadherin promoter.UTX deficiency by siRNA knockdown enhanced the bindings of histone methyltransferase EZH2 and SUZ12 at the E-cadherin promoter.Conversely,UTX overexpression suppressed the recruitment levels of EZH2 and SUZ12 at the E-cadherin promoter.5.UTX recruits acetylated H3K27 to the E-cadherin promoter by physically interacting with CBP.Inactivation of UTX inhibited H3K27 ac binding at the E-cadherin promoter.In contrast,overexpression of UTX increased H3K27 ac at the E-cadherin promoter.UTX knockdown decreased histone acetyltransferase CBP recruitment to the E-cadherin promoter,while UTX overexpression did the opposite,which resembles the pattern of H3K27 ac.After immunoprecipitating CBP in HCT-116 cells,we detected the associated UTX.Immunoprecipitation of UTX with specific anti-UTX antibody also pulled down the CBP protein.Conclusions1.UTX is a key determinant of E-cadherin expression in human colon cancer cells.2.UTX may inhibit the migration and invasion capacity of HCT-116 cells mediated by E-cadherin.3.UTX may serve as a novel therapeutic target in the treatment of colon cancer metastasis.