Stable 5‐Hydroxymethylcytosine (5hmC) Acquisition Marks Gene Activation During Chondrogenic Differentiation

• Published in: Journal of bone and mineral research Vol. 31; no. 3; pp. 524 - 534
• Main Authors: Taylor, Sarah EB, Li, Ye Henry, Smeriglio, Piera, Rath, Madhusikta, Wong, Wing H, Bhutani, Nidhi
• Format: Journal Article
• Language: English
• Published: England Wiley Subscription Services, Inc 01-03-2016; American Society for Bone and Mineral Research
• Subjects: 5-Methylcytosine - analogs & derivatives; 5‐HYDROXYMETHYLCYTOSINE; Animals; Biochemistry, Molecular Biology; Cartilage - embryology; Cell Differentiation - genetics; Cellular Biology; Chondrocytes - cytology; Chondrocytes - metabolism; CHONDROGENESIS; Chondrogenesis - genetics; Cytosine - analogs & derivatives; Cytosine - metabolism; DIFFERENTIATION; DNA DEMETHYLATION; DNA, Intergenic - genetics; DNA-Binding Proteins - metabolism; Embryonic Development - genetics; EPIGENETICS; Extremities - embryology; Gene Expression Regulation, Developmental; Human health and pathology; Life Sciences; Mice; Molecular biology; Proto-Oncogene Proteins - metabolism; Rhumatology and musculoskeletal system; Stem Cells - cytology; Transcriptional Activation - genetics; 5-HYDROXYMETHYLCYTOSINE; EPIGENETICS; DIFFERENTIATION; CHONDROGENESIS; DNA DEMETHYLATION
• ISSN: 0884-0431; 1523-4681
• DOI: 10.1002/jbmr.2711

ABSTRACT
Regulation of gene expression changes during chondrogenic differentiation by DNA methylation and demethylation is little understood. Methylated cytosines (5mC) are oxidized by the ten‐eleven‐translocation (TET) proteins to 5‐hydroxymethylcytosines (5hmC), 5‐formylcytosines (5fC), and 5‐carboxylcytosines (5caC), eventually leading to a replacement by unmethylated cytosines (C), ie, DNA demethylation. Additionally, 5hmC is stable and acts as an epigenetic mark by itself. Here, we report that global changes in 5hmC mark chondrogenic differentiation in vivo and in vitro. Tibia anlagen and growth plate analyses during limb development at mouse embryonic days E 11.5, 13.5, and 17.5 showed dynamic changes in 5hmC levels in the differentiating chondrocytes. A similar increase in 5hmC levels was observed in the ATDC5 chondroprogenitor cell line accompanied by increased expression of the TET proteins during in vitro differentiation. Loss of TET1 in ATDC5 decreased 5hmC levels and impaired differentiation, demonstrating a functional role for TET1‐mediated 5hmC dynamics in chondrogenic differentiation. Global analyses of the 5hmC‐enriched sequences during early and late chondrogenic differentiation identified 5hmC distribution to be enriched in the regulatory regions of genes preceding the transcription start site (TSS), as well as in the gene bodies. Stable gains in 5hmC were observed in specific subsets of genes, including genes associated with cartilage development and in chondrogenic lineage‐specific genes. 5hmC gains in regulatory promoter and enhancer regions as well as in gene bodies were strongly associated with activated but not repressed genes, indicating a potential regulatory role for DNA hydroxymethylation in chondrogenic gene expression. © 2015 American Society for Bone and Mineral Research.