Molecular basis of microhomology-mediated end-joining by purified full-length Polθ

被引：78

作者：

Black, Samuel J. ^{[1
]}

Ozdemir, Ahmet Y. ^{[1
]}

Kashkina, Ekaterina ^{[1
]}

Kent, Tatiana ^{[1
]}

Rusanov, Timur ^{[1
]}

Ristic, Dejan ^{[2
,3
]}

Shin, Yeonoh ^{[4
]}

Suma, Antonio ^{[5
]}

Hoang, Trung ^{[1
]}

Chandramouly, Gurushankar ^{[1
]}

Siddique, Labiba A. ^{[1
]}

Borisonnik, Nikita ^{[1
]}

Sullivan-Reed, Katherine ^{[6
]}

Mallon, Joseph S. ^{[1
]}

Skorski, Tomasz ^{[6
]}

Carnevale, Vincenzo ^{[5
]}

Murakami, Katsuhiko S. ^{[4
]}

Wyman, Claire ^{[2
,3
]}

Pomerantz, Richard T. ^{[1
]}

机构：

[1] Temple Univ, Lewis Katz Sch Med, Dept Med Genet & Mol Biochem, Fels Inst Canc Res, Philadelphia, PA 19140 USA

[2] Erasmus MC, Dept Mol Genet, NL-3000 CA Rotterdam, Netherlands

[3] Erasmus MC, Dept Radiat Oncol, NL-3000 CA Rotterdam, Netherlands

[4] Penn State Univ, Ctr RNA Mol Biol, University Pk, PA 16802 USA

[5] Temple Univ, Inst Computat Mol Sci, Philadelphia, PA 19122 USA

[6] Temple Univ, Lewis Katz Sch Med, Dept Microbiol & Immunol, Fels Inst Canc Res, Philadelphia, PA 19140 USA

来源：

NATURE COMMUNICATIONS | 2019年 / 10卷 / 1期

基金：

美国国家科学基金会; 美国国家卫生研究院;

关键词：

DNA-POLYMERASE-THETA; HELICASE DOMAIN; REPAIR; NHEJ; REPLICATION; LESION;

D O I：

10.1038/s41467-019-12272-9

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

DNA polymerase theta (Pol theta) is a unique polymerase-helicase fusion protein that promotes microhomology-mediated end-joining (MMEJ) of DNA double-strand breaks (DSBs). How full-length human Pol theta performs MMEJ at the molecular level remains unknown. Using a biochemical approach, we find that the helicase is essential for Pol theta MMEJ of long ssDNA overhangs which model resected DSBs. Remarkably, Pol theta MMEJ of ssDNA overhangs requires polymerase-helicase attachment, but not the disordered central domain, and occurs independently of helicase ATPase activity. Using single-particle microscopy and biophysical methods, we find that polymerase-helicase attachment promotes multimeric gel-like Pol theta complexes that facilitate DNA accumulation, DNA synapsis, and MMEJ. We further find that the central domain regulates Pol theta multimerization and governs its DNA substrate requirements for MMEJ. These studies identify unexpected functions for the helicase and central domain and demonstrate the importance of polymerase-helicase tethering in MMEJ and the structural organization of Pol theta.

引用

页数：16

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