H4K20 methylation regulates quiescence and chromatin compaction

Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA. Massachusetts General Hospital Cancer Center, Harvard Medical School, Charlestown, MA, 02129, USA. Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, PA 19104, USA. Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles and Department of Biological Chemistry, David Geffen School of Medicine, Los Angeles, CA 90095, USA.
Molecular biology of the cell (Impact Factor: 4.47). 08/2013; 24(19). DOI: 10.1091/mbc.E12-07-0529
Source: PubMed


The transition between proliferation and quiescence is frequently associated with changes in gene expression, in the extent of chromatin compaction and in histone modifications, but whether changes in chromatin state actually regulate cell cycle exit with quiescence is unclear. We discovered that primary human fibroblasts induced into quiescence exhibited tighter chromatin compaction. Mass spectrometry analysis of histone modifications revealed that H4K20me2 and -me3 are increased in quiescence and that other histone modifications are present at similar levels in proliferating and quiescent cells. Analysis of cells in S, G2/M, and G1 phases shows that H4K20me1 increases after S phase and is converted to -me2 and -me3 in quiescence. Knockdown of the enzymes that create H4K20me2 and -me3 resulted in an increased fraction of cells in S phase, a defect in exiting the cell cycle, and decreased chromatin compaction. Overexpression of Suv4-20h1, the enzyme that creates H4K20me2 from H4K20me1, resulted in G2 arrest, consistent with a role for H4K20me1 in mitosis. The results suggest that the same lysine on H4K20 may, in its different methylation states, facilitate mitotic functions in M phase and promote chromatin compaction and cell cycle exit in quiescent cells.

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    • "For measurement of proliferation by Click-iT analysis, MEFs (lines WT 3 and KO 3) were incubated for 1 h in 10 µM EdU, and labeled as previously described (Evertts et al., 2013). Briefly, 1 to 2×105 cells were pelleted, fixed with 4% paraformaldehyde, permeabilized and treated with Alexa 488 azide (Invitrogen). "
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    • "Increased chromatin compaction was confirmed by probing chromatin accessibility by micrococcal nuclease (MNase) digestion . In accord with a recent study showing global chromatin compaction and upregulation of H4K20me3 in contact-inhibited human fibroblasts (Evertts et al., 2013), quiescent NIH 3T3 cells and wild-type MEFs exhibited a chromatin structure that is less accessible to MNase digestion (Figures 3C, 3D, and S3A). In contrast, chromatin from Suv4-20h À/À MEFs was readily digested , regardless of whether cells were cultured in 10% or 0.1% serum, indicating that H4K20me3-mediated chromatin compaction alters global chromatin structure in response to growth arrest. "
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    • "It has a distinctive transcriptional profile, with overall changes in the level of transcription and chromatin organisation [13]. Entry into quiescence is generally associated with increased chromatin compaction [14] and it is therefore perhaps surprising that this was accompanied by reduced trypsin-sensitive masking of 5meC. However, this observation seems consistent with results of staining in the zygote where the level of 5meC epitope masking was greatest in the late zygote stage where chromatin structure is generally considered least compacted. "
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