Optimization of epigenomic modification with regulatory RNAs considering the presence of endogenous non-coding RNAs

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Recent accumulating evidences have indicated that various non-coding RNAs(ncRNAs) are involved in region-specific epigenetic regulations. Since many endogenous ncRNAs show tissue/cell-type-dependent expression patterns, it is important to reveal the relationships between tissue/cell-type-dependent expression and epigenetic regulations. Our data indicated that many genes had ncRNAs within promoter regions of tissu/cell-type-specific genes whose DNA methylation status changed by deficiency of ncRNA-related enzymes. The Sall4 gene, which is crucial for stemness of embryonic stem(ES) cells, had an endogenous antisense ncRNA(ASncRNA) overlapping with the promoter region of the protein-coding Sall4 gene. The Sall4 ASncRNA regulated DNA methylation status of the Sall4 promoter independent of double-strand RNA synthesis by Dicer. An ectopic overexpression of the ASncRNA in fibroblast cells resulted DNA demethylation of the Sall4 promoter region. The Sall4 ASncRNA was proven indispensable for maintenance of unmethylated and activated status in ES cells by shRNA knockdown of the endogenous ASncRNA. Our data also indicated that oocyte-specific histone and environmental chemicals were involved in epigenetic regulations. In conclusion, we could identify many ASncRNAs that regulate DNA methylation status in a target-specific manner, and these ASncRNAs are thought as useful for epigenome modification to establish epigenome-related disease model animals and cells.