End‐joining of reconstituted histone H2AX‐containing chromatin in vitro by soluble nuclear proteins from human cells

FEBS Letters

Bradbury, E.Morton¹  Zalenskaya, Irina A¹  Tomilin, Nikolai V¹  Yau, Peter M¹  Siino, Joseph S¹  Nazarov, Igor B¹ 

[1] Department of Biological Chemistry, UC Davis School of Medicine, Davis, CA 95616, USA

关键词: Reconstituted chromatin;    DNA end-joining;    Histone H2AX;    Phosphorylation;    NHEJ;    non-homologous end-joining;    DSB;    double-strand DNA break;    DTT;    dithiothreitol;    AFM;    atomic force microscopy;    rhH2AX;    recombinant human H2AX;    NCP;    nucleosome core particle;   

DOI  :  10.1016/S0014-5793(02)03176-9

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Non-homologous end-joining is an important pathway for the repair of DNA double-strand breaks. This type of DNA break is followed by the rapid phosphorylation of Ser-139 in the histone variant H2AX to form γ-H2AX. Here we report efficient in vitro end-joining of reconstituted chromatin containing nucleosomes made with either H2A or H2AX. This reaction is catalyzed by nuclear extracts from human cells and this end-joining is not suppressed by the PI-3 kinase inhibitor wortmannin. During the end-joining reaction H2AX is phosphorylated at Ser-139 as detected by immunoblot with specific antibodies and this phosphorylation is inhibited by wortmannin. Therefore, in vitro the DNA end-joining reaction appears to be independent of H2AX phosphorylation.

【 授权许可】

Unknown