The Histone H4 Lysine 20 Monomethyl Mark, Set by PR-Set7 and Stabilized by L(3)mbt, Is Necessary for Proper Interphase Chromatin Organization

Drosophila PR-Set7 or SET8 is a histone methyltransferase that specifically monomethylates histone H4 lysine 20 (H4K20). L(3)MBT has been identified as a reader of methylated H4K20. It contains several conserved domains including three MBT repeats binding mono- and dimethylated H4K20 peptides. We find that the depletion of PR-Set7 blocks de novo H4K20me1 resulting in the immediate activation of the DNA damage checkpoint, an increase in the size of interphase nuclei, and drastic reduction of cell viability. L(3)mbt on the other hand stabilizes the monomethyl mark, as L(3)mbt-depleted S2 cells show a reduction of more than 60% of bulk monomethylated H4K20 (H4K20me1) while viability is barely affected. Ploidy and basic chromatin structure show only small changes in PR-Set7-depleted cells, but higher order interphase chromatin organization is significantly affected presumably resulting in the activation of the DNA damage checkpoint. In the absence of any other known functions of PR-Set7, the setting of the de novo monomethyl mark appears essential for cell viability in the presence or absence of the DNA damage checkpoint, but once newly assembled chromatin is established the monomethyl mark, protected by L(3)mbt, is dispensable.

果蝇（Drosophila）PR-Set7（或SET8）是一种组蛋白甲基转移酶（histone methyltransferase），可特异性催化组蛋白H4赖氨酸20（histone H4 lysine 20，H4K20）的单甲基化修饰。L(3)MBT已被鉴定为甲基化H4K20的组蛋白阅读器（reader），其包含多个保守结构域，其中包括3个MBT重复序列（MBT repeats），可结合单甲基化与二甲基化的H4K20肽段。我们发现，PR-Set7的表达沉默会阻断从头建立的H4K20单甲基化修饰（H4K20me1），进而快速激活DNA损伤检验点（DNA damage checkpoint），导致间期细胞核体积增大，并使细胞活力大幅下降。而L(3)mbt则可稳定该单甲基化修饰标记：在L(3)mbt表达沉默的果蝇S2细胞中，整体单甲基化H4K20（H4K20me1）的水平下降超过60%，但细胞活力几乎未受影响。在PR-Set7表达沉默的细胞中，细胞倍性与基础染色质结构仅发生微小变化，但间期染色质的高级组织结构却受到显著影响，推测这会导致DNA损伤检验点的激活。鉴于目前尚未明确PR-Set7的其他已知功能，从头建立单甲基化修饰标记这一过程，无论DNA损伤检验点存在与否，似乎都对细胞活力至关重要；但一旦新组装的染色质形成，由L(3)mbt保护的单甲基化修饰标记便不再必需。