PARENT SESSION

(90) REGULATION OF NANOG GENE EXPRESSION BY DNA METHYLATION AND HISTONE MODIFICATION.

Hattori, Naoko¹, Imao, Yuko¹, Nishino, Koichiro², Hattori, Naka¹, Tanaka, Satoshi¹, Shiota, Kunio¹, ¹ The University of Tokyo, Tokyo, Japan² Samuel Lunenfeld Research Institute, Toronto, ON, Canada

ABSTRACT- The expression of Nanog gene is found in inner cell mass at the blastocyst stage, but is not detected in trophectoderm and general somatic cells. Nanog gene is also expressed in embryonic stem (ES) cells, while severely repressed in somatic cell lines and trophoblast stem (TS) cells. Thus, Nanog may be a master regulator responsible for stemness in early embryos. DNA methylation is a main epigenetic mechanism for gene regulation, which is a heritable change without sequence alteration in the genomic DNA. Here we found that there is a tissue-dependent, differentially methylated region (T-DMR) in Nanog upstream region by analyzing DNA methylation status. The T-DMR is hypomethylated in ES cells, but hypermethylated in TS cells, NIH/3T3 cells and adult liver. T-DMR at Nanog upstream region has a promoter activity. In the reporter assay using T-DMR-luciferase construct, in vitro DNA methylation of T-DMR severely suppressed the promoter activity in ES cells. Therefore, the regulatory region of Nanog gene is a target of DNA methylation. Furthermore, chromatin immunoprecipitation assay revealed that histone H3 proteins are hyperacetylated, histone H3 lysine (K) 4 is hypermethylated and histone H3-K9 and K27 are hypomethylated at Nanog gene locus in ES cells. In contrast, in TS cells, histone H3 proteins are hypoacetylated, histone H3-K4 is hypomethylated and histone H3-K9 and K27 are hypermethylated at Nanog gene locus. Therefore, chromatin structure at Nanog upstream region is relaxed in ES cells, and chromatin condensation occurs over Nanog upstream region in TS cells. This study clearly revealed that DNA methylation is associated with chromatin structure at the Nanog T-DMR. We conclude that an epigenetic mechanism consisting of DNA methylation and histone modifications regulates the expression of Nanog gene.

KEY WORDS: Nanog, DNA methylation, histone modification, embryonic stem cell