Complementary roles for histone deacetylases 1, 2, and 3 in differentiation of pluripotent stem cells

Abstract In eukaryotic cells, covalent modifications to core histones contribute to the establishment and maintenance of cellular phenotype via regulation of gene expression. Histone acetyltransferases (HATs) cooperate with histone deacetylases (HDACs) to establish and maintain specific patterns of histone acetylation. HDAC inhibitors can cause pluripotent stem cells to cease proliferating and enter terminal differentiation pathways in culture. To better define the roles of individual HDACs in stem cell differentiation, we have constructed "dominant-negative" stem cell lines expressing mutant, Flag-tagged HDACs with reduced enzymatic activity. Replacement of a single residue (His→Ala) in the catalytic center reduced the activity of HDACs 1 and 2 by 80%, and abolished HDAC3 activity; the mutant HDACs were expressed at similar levels and in the same multiprotein complexes as wild-type HDACs. Hexamethylene bisacetamide-induced MEL cell differentiation was potentiated by the individual mutant HDACs, but only to 2%, versus 60% for an HDAC inhibitor, sodium butyrate, suggesting that inhibition of multiple HDACs is required for full potentiation. Cultured E14.5 cortical stem cells differentiate to neurons, astrocytes, and oligodendrocytes upon withdrawal of basic fibroblast growth factor. Transduction of stem cells with mutant HDACs 1, 2, or 3 shifted cell fate choice toward oligodendrocytes. Mutant HDAC2 also increased differentiation to astrocytes, while mutant HDAC1 reduced differentiation to neurons by 50%. These results indicate that HDAC activity inhibits differentiation to oligodendrocytes, and that HDAC2 activity specifically inhibits differentiation to astrocytes, while HDAC1 activity is required for differentiation to neurons.     

Coordinated expression of p300 and HDAC3 upregulates histone acetylation during dentinogenesis

Cellular differentiation is caused by highly controlled modifications in the gene expression but rarely involves a change in the DNA sequence itself. Histone acetylation is a major epigenetic factor that adds an acetyl group to histone proteins, thus altering their interaction with DNA and nuclear proteins. Illumination of the histone acetylation during dentinogenesis is important for odontoblast differentiation and dentinogenesis. In the current study, we aimed to discover the roles and regulation of acetylation at histone 3 lysine 9 (H3K9ac) and H3K27ac during dentinogenesis. We first found that both of these modifications were enhanced during odontoblast differentiation and dentinogenesis. These modifications are dynamically catalyzed by histone acetyltransferases (HATs) and deacetylases (HDACs), among which HDAC3 was decreased while p300 increased during odontoblast differentiation. Moreover, overexpression of HDAC3 or knockdown p300 inhibited odontoblast differentiation in vitro, and inhibition of HDAC3 and p300 with trichostatin A or C646 regulated odontoblast differentiation. Taken together, the results of our present study suggest that histone acetylation is involved in dentinogenesis and coordinated expression of p300- and HDAC3-regulated odontoblast differentiation through upregulating histone acetylation.