title: A PP4-phosphatase complex dephosphorylates gamma-H2AX generated during DNA replication.
abstract: The histone H2A variant H2AX is rapidly phosphorylated in response to DNA double-stranded breaks to produce gamma-H2AX. gamma-H2AX stabilizes cell-cycle checkpoint proteins and DNA repair factors at the break site. We previously found that the protein phosphatase PP2A is required to resolve gamma-H2AX foci and complete DNA repair after exogenous DNA damage. Here we describe a three-protein PP4 phosphatase complex in mammalian cells, containing PP4C, PP4R2, and PP4R3beta, that specifically dephosphorylates ATR-mediated gamma-H2AX generated during DNA replication. PP4 efficiently dephosphorylates gamma-H2AX within mononucleosomes in vitro and does not directly alter ATR or checkpoint kinase activity, suggesting that PP4 acts directly on gamma-H2AX in cells. When the PP4 complex is silenced, repair of DNA replication-mediated breaks is inefficient, and cells are hypersensitive to DNA replication inhibitors, but not radiomimetic drugs. Therefore, gamma-H2AX elimination at DNA damage foci is required for DNA damage repair, but accomplishing this task involves distinct phosphatases with potentially overlapping roles.
authors.affiliation: Immune Disease Institute, Harvard Medical School, Boston, MA 02115, USA. dipanjan_chowdhury@dfci.harvard.edu
authors.forename: Dipanjan
authors.initials: D
authors.lastname: Chowdhury
authors.forename: Xingzhi
authors.initials: X
authors.lastname: Xu
authors.forename: Xueyan
authors.initials: X
authors.lastname: Zhong
authors.forename: Fariyal
authors.initials: F
authors.lastname: Ahmed
authors.forename: Jianing
authors.initials: J
authors.lastname: Zhong
authors.forename: Ji
authors.initials: J
authors.lastname: Liao
authors.forename: Derek M
authors.initials: DM
authors.lastname: Dykxhoorn
authors.forename: David M
authors.initials: DM
authors.lastname: Weinstock
authors.forename: Gerd P
authors.initials: GP
authors.lastname: Pfeifer
authors.forename: Judy
authors.initials: J
authors.lastname: Lieberman
mesh_terms: D000595:Amino Acid Sequence; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000378:metabolism; D002455:Cell Division; D002460:Cell Line; D002843:Chromatin / Q000378:metabolism; D004249:DNA Damage; D004261:DNA Replication; D020868:Gene Silencing; D006657:Histones / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D054648:Protein Phosphatase 2 / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D021122:Protein Subunits / Q000378:metabolism; D021381:Protein Transport; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D002843:Chromatin; C468783:H2AX protein, human; D006657:Histones; D046912:Multiprotein Complexes; D021122:Protein Subunits; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; C080406:protein phosphatase 4