Stability of XIST repression in relation to genomic imprinting following global genome demethylation in a human cell line

• Published in: Brazilian journal of medical and biological research 2014-12, Vol.47 (12), p.1029-1035
• Main Authors: Araújo, E S S de, Vasques, L R, Stabellini, R, Krepischi, A C V, Pereira, L V
• Format: Article
• Language: English
• Subjects: 5-aza-2′-deoxycytidine; Apoptosis Regulatory Proteins; Azacitidine - administration & dosage; Azacitidine - analogs & derivatives; BIOLOGY; Biomedical Sciences; Decitabine; DNA (Cytosine-5-)-Methyltransferase 1; DNA (Cytosine-5-)-Methyltransferases - genetics; DNA methylation; DNA Methylation - drug effects; DNA Methylation - genetics; DNA Methyltransferase 3B; DNA-Binding Proteins; Epigenetic Repression - genetics; Gene expression; Gene Knockout Techniques; Genome - genetics; Genome, Human - drug effects; Genomic Imprinting - genetics; HCT116 Cells; Health aspects; Human cell line; Humans; Imprinted genes; In Situ Hybridization, Fluorescence - methods; Insulin-Like Growth Factor II - genetics; Medical research; MEDICINE, RESEARCH & EXPERIMENTAL; Microarray Analysis; Polymorphism, Single Nucleotide; Proteins - metabolism; Real-Time Polymerase Chain Reaction - methods; Reverse Transcriptase Polymerase Chain Reaction - methods; RNA, Long Noncoding - genetics; RNA, Long Noncoding - metabolism; RNA-Binding Proteins; X chromosome; XIST
• ISSN: 0100-879X; 1414-431X; 1414-431X
• DOI: 10.1590/1414-431X20144058

DNA methylation is essential in X chromosome inactivation and genomic imprinting, maintaining repression of XIST in the active X chromosome and monoallelic repression of imprinted genes. Disruption of the DNA methyltransferase genes DNMT1 and DNMT3B in the HCT116 cell line (DKO cells) leads to global DNA hypomethylation and biallelic expression of the imprinted gene IGF2 but does not lead to reactivation of XIST expression, suggesting that XIST repression is due to a more stable epigenetic mark than imprinting. To test this hypothesis, we induced acute hypomethylation in HCT116 cells by 5-aza-2'-deoxycytidine (5-aza-CdR) treatment (HCT116-5-aza-CdR) and compared that to DKO cells, evaluating DNA methylation by microarray and monitoring the expression of XIST and imprinted genes IGF2, H19, and PEG10. Whereas imprinted genes showed biallelic expression in HCT116-5-aza-CdR and DKO cells, the XIST locus was hypomethylated and weakly expressed only under acute hypomethylation conditions, indicating the importance of XIST repression in the active X to cell survival. Given that DNMT3A is the only active DNMT in DKO cells, it may be responsible for ensuring the repression of XIST in those cells. Taken together, our data suggest that XIST repression is more tightly controlled than genomic imprinting and, at least in part, is due to DNMT3A.