The Role Of DNA Demethylation In AdSCs Differentiation And Erythrocyte Development

Methylation and demethylation of DNA are the complementary processes of epigenetic regulation. Methylation and demethylation influence a diverse array of cellular activities, including X chromosome inactivation, genomic imprinting, chromatin modification, and the silencing of endogenous genes. It was generally believed that DNA demethylation occurs passively over several cycles of DNA replication, and that active DNA demethylation is rare. Recently, evidence for active DNA demethylation has been obtain in several cancer, neuronal, and embryonic stem cell lines. However, the mechanisms of active DNA demethylation are still unknown and its role in stem cell differentiation and hematogenesis are poorly understood. Here we investigated the DNA demethylation in mice and zebrafish model and found it is essential for adipose-derived stem cell (ADSCs) differentiation and zebrafish erythrocyte development.Studies in embryonic stem cell models, however, suggested that active DNA demethylation might be restricted to the early development of progenitor cells. Whether active demethylation is involved in terminal differentiation of adult stem cells is unknown. Now we provide evidence that active DNA demethylation does occur during terminal specification of stem cells in an ADSC-derived osteogenic differentiation model. The promoters of the Dlx5, Runx2, BGP, and Osterix osteogenic genes were demethylated during the increase in gene expression associated with osteogenic differentiation. The growth arrest and DNA-damage-inducible protein Gadd45a was up-regulated in these processes. Knockdown of Gadd45a led to hypermethylation of Dlx5, Runx2, BGP, and Osterix promoters, followed by suppression of Dlx5, Runx2, BGP, and Osterix expression and interruption of osteogenic differentiation. These results reveal that Gadd45a plays an essential role in gene-specific active DNA demethylation during adult stem cell differentiation. These results enhance current knowledge of osteogenic specification and may also lead to a better understanding of the common mechanisms underlying epigenetic regulation in adult stem cell differentiation.Recently the ten-eleven translocation (TET) proteins were found to play a crucial role in DNA demethylation by hydrolyzing to5-hydroxymethylcytosine, which can be converted to cytosine. We cloned three TET genes in zebrafish firstly, TET1, TET2and TET3. Knockdown of TET2and TET3genes by Morpholinos inhibit zebrafish hematopoietic gene expressions as well as erythrocyte development. These results shows that the convers of5-methylcytosine to5-hydroxymethylcytosine is important for zebrafish hematogenesis.In a word, we studied the role of DNA demethylation in cell differentiation and development in mice and zebrafish model, and found Gadd45a is essential for gene-specific active DNA demethylation during ADSCs osteogenic differentiation, and TET proteins participate in erythrocyte development in zebrafish.