Targeted DNA methylation in cell fate determination

4298 Locus-specific DNA methylation was developed to epigenomically modify the bivalent loci identified by ChIP-on-chip (trimethylated lysine 27 histone 3) and DNA methylation hybridization microarray to direct the cell fate of mesenchymal stem cell (MSC) and cancer cells. Trip10 was identified as bivalent locus, which methylation status was increased during MSC-to-liver/neuron differentiation but decreased during MSC-to-adipocyte differentiation. Trip10 promoter was mostly hypermethylated in breast (n=94) and brain tumor specimen (n=47), but hypomethylated in liver cancer samples (n=37). Overexpressed Trip10 in IMR32 (neuroblastoma) promoted tumor growth, as indicated by colony formation assay and subsequent tumor formation in subcutaneously injected immuno-deficient mice. However, outgrowth of tumor was suppressed when Trip10 was overexpressed in CP70 cells. Consistent with other reports, we found Trip10 interacted with Cdc42 and Huntingtin to regulate cell cycle/growth and apoptosis. If the bivalent nature of Trip10 were true, we hypothesized that methylation of Trip10 should be sufficient to change the cell fate. We evaluated this hypothesis by pre-depositing DNA methylation within the Trip10 promoter in MSC. Compared to the mock-treated MSC, those Trip10 methylated MSC exhibited accelerated MSC-to-neuron differentiation accompanied with reduced cell death after differentiation. Therefore, methylation of the bivalent loci is sufficient to determine the cell fate of somatic stem cells. Similar strategy was applied to specifically target the estrogen receptor non-expressing (ER-) breast cancer cell line, MDA-MB-231. While most of the ER target loci were methylated and silenced in ER- cells as compared with the methylation status in ER+ cells, there were 19 genes actually activated. We reasoned that these 19 loci were crucial for the survival or the oncogenic activity of MDA-MB-231. Tailored methylation of these 19 loci led to increased cell death of MDA-MB-231 but not in cells including MCF7, HepG2, CP70, IMR32, and MSC. It is thus concluded that we have developed a new method that can epigenetically tailor the cell fate of stem cells or kill the targeted tumor cells specifically. (Supported in part by NSC 95-3112-B-194-001, NSC 94-2320-B-194-006, NSC 94-2320-B-194-003)