The Regulation and Function of Histone H4 Lysine-20 Methylation
Yang, Hongbo
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Permalink
https://hdl.handle.net/2142/72486
Description
Title
The Regulation and Function of Histone H4 Lysine-20 Methylation
Author(s)
Yang, Hongbo
Issue Date
2009
Doctoral Committee Chair(s)
Belmont, Andrew S.
Mizzen, Craig,
Department of Study
Cell and Developmental Biology
Discipline
Cell and Developmental Biology
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Cell
Chemistry, Biochemistry
Abstract
Posttranslational modifications (PTMs) of histone tails direct nuclear processes including transcription, DNA repair, and chromatin packaging. Lysine 20 (K20) of histone H4 is mono-, di-, or trimethylated in vivo, but the regulation and significance of these methylations are poorly understood. Our work revealed that ∼90% or ∼80% of K20 in histone H4 is dimethylated in asynchronous Drosophila S2 cells and human HeLa cells, respectively, as quantified from Top Down Mass Spectrometry. The SET domain proteins PR-Set7 and Suv4-20 have previously been implicated in mono- and trimethylation of K20, respectively, but enzymes that dimethylate K20 had not been identified when we began this work. Using an RNAi screen targeting all SET domain containing genes, we identified Drosophila Suv4-20 as a mixed product specificity methyltransferase that dimethylates ∼90% and trimethylates less than 5% of total H4 at K20 in Drosophila S2 cells. Trimethylation, but not dimethylation, is reduced in Drosophila larvae lacking heterochromatin protein 1 (HP1), suggesting HP1 is important for Suv4-20 mediated K20 trimethylation. Similarly, the human Suv4-20h1/h2 enzymes generate both di- and trimethyllysine 20. Overexpression and siRNA knockdown studies suggest that human Suv4-20h1 and h2 have complementary activity for K20 methylation but different product specificity preference. Our data suggest that Suv4-20h1 preferentially forms K20 dimethylation in vivo, whereas Suv4-20h2 appears to form trimethylation preferentially. Mass spectrometry in conjunction with cell synchronization, metabolic labeling, RNA interference, and other approaches reveal that PR-Set7 and Suv4-20 mediate progressive global mono-, di-, and trimethylation of K20 in newly synthesized histone H4 beginning approximately at the G2/M transition, well after new H4 is deposited in replicating chromatin during S phase. Functional analyses indicate that H4-K20 dimethylation is required for the formation of 53BP1 nuclear foci at DNA damage sites in human HeLa cells. Collectively, the data indicate that Suv4-20h1/h2 mediate ubiquitous dimethylation that facilitates DNA damage responses and selective trimethylation that is involved in heterochromatin formation.
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