Genetic and physiological regulation of non-homologous end-joining in mammalian cells

被引:22
作者
Tachibana, A [1 ]
机构
[1] Kyoto Univ, Ctr Radiat Biol, Sakyo Ku, Kyoto 6068501, Japan
来源
NOVEL DEVELOPMENTS ON GENETIC RECOMBINATION: DNA DOUBLE STRAND BREAK AND DNA END-JOINING | 2004年 / 38卷
关键词
D O I
10.1016/S0065-227X(04)80046-7
中图分类号
Q6 [生物物理学];
学科分类号
071011 ;
摘要
Repair of DSBs is important to prevent chromosomal fragmentation, translocations and deletions. To investigate the process in NHEJ, we have established an in vitro system to clarify the measurement and analysis of the efficiency and the fidelity of rejoining of DSBs, and applied the method to investigate NHEJ in human cells derived from patients suffering from cancer-prone hereditary diseases. A DSB was introduced in plasmid pZErO-2 at a specific site within the ccdB gene that is lethal to E. coli cells, and treated with nuclear extracts from human cells. The efficiency of rejoining in the nuclear extract from an A-T cell line was comparable to that from a control cell line. However, the accuracy of rejoining was much lower for the A-T cell extract than for the control cell extract. All mutations were deletions, most of which contained short direct repeats at the breakpoint junctions. The deletion spectrum caused by the A-T nuclear extract was distinct from that by the control extract. These results indicate that A-T cells have certain deficiencies in end-joining of double-strand breaks in DNA. The extract from BS cells also showed the similar activity and the lower fidelity of rejoing compared to that from normal cells. From the sequencing analysis of the junction of DSBs, it is speculated that the defect in the BLM helicase might cause irregular rejoining of DSBs. Radioadaptive response is the acquirement of cellular resistance to ionizing radiation by prior exposure to low dose. We investigated the in vitro end-joining activity of DNA ends in radioadaptive cells. Both the efficiency and the fidelity of rejoining in the cells pre-exposed to low dose are increased comparing to those without pre-exposure. We also investigated the joining activity of DNA ends in p53-deficient cells. Pre-irradiation caused no apparent alteration in both the efficiency and fidelity of end-joining. These results suggest that the exposure to low dose activates a cellular function to repair DSBs efficiently, which is dependent on p53. These results indicate that NHEJ pathway is regulated by many factors; genetic regulation by ATM and BLM, and physiological conditions such as irradiation with ionizing radiation. The observations also suggest that in some occasions p53 might play a key role in NHEJ.
引用
收藏
页码:21 / 44
页数:24
相关论文
共 104 条
[1]   Phosphorylation and rapid relocalization of 53BP1 to nuclear foci upon DNA damage [J].
Anderson, L ;
Henderson, C ;
Adachi, Y .
MOLECULAR AND CELLULAR BIOLOGY, 2001, 21 (05) :1719-1729
[2]   HIGH-FREQUENCIES OF INVERSIONS AND TRANSLOCATIONS OF CHROMOSOMES 7 AND 14 IN ATAXIA TELANGIECTASIA [J].
AURIAS, A ;
DUTRILLAUX, B ;
BURIOT, D ;
LEJEUNE, J .
MUTATION RESEARCH, 1980, 69 (02) :369-374
[3]   Enhanced phosphorylation of p53 by ATN in response to DNA damage [J].
Banin, S ;
Moyal, L ;
Shieh, SY ;
Taya, Y ;
Anderson, CW ;
Chessa, L ;
Smorodinsky, NI ;
Prives, C ;
Reiss, Y ;
Shiloh, Y ;
Ziv, Y .
SCIENCE, 1998, 281 (5383) :1674-1677
[4]   DNA end-joining catalyzed by human cell-free extracts [J].
Baumann, P ;
West, SC .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1998, 95 (24) :14066-14070
[5]   Identification of a Saccharomyces cerevisiae Ku80 homologue: Roles in DNA double strand break rejoining and in telomeric maintenance [J].
Boulton, SJ ;
Jackson, SP .
NUCLEIC ACIDS RESEARCH, 1996, 24 (23) :4639-4648
[6]   Saccharomyces cerevisiae Ku70 potentiates illegitimate DNA double-strand break repair and serves as a barrier to error-prone DNA repair pathways [J].
Boulton, SJ ;
Jackson, SP .
EMBO JOURNAL, 1996, 15 (18) :5093-5103
[7]   ERKS - A FAMILY OF PROTEIN-SERINE THREONINE KINASES THAT ARE ACTIVATED AND TYROSINE PHOSPHORYLATED IN RESPONSE TO INSULIN AND NGF [J].
BOULTON, TG ;
NYE, SH ;
ROBBINS, DJ ;
IP, NY ;
RADZIEJEWSKA, E ;
MORGENBESSER, SD ;
DEPINHO, RA ;
PANAYOTATOS, N ;
COBB, MH ;
YANCOPOULOS, GD .
CELL, 1991, 65 (04) :663-675
[8]   Interaction of p53 with the human Rad51 protein [J].
Buchhop, S ;
Gibson, MK ;
Wang, XW ;
Wagner, P ;
Sturzbecher, HW ;
Harris, CC .
NUCLEIC ACIDS RESEARCH, 1997, 25 (19) :3868-3874
[9]   Activation of the ATM kinase by ionizing radiation and phosphorylation of p53 [J].
Canman, CE ;
Lim, DS ;
Cimprich, KA ;
Taya, Y ;
Tamai, K ;
Sakaguchi, K ;
Appella, E ;
Kastan, MB ;
Siliciano, JD .
SCIENCE, 1998, 281 (5383) :1677-1679
[10]   The hMre11/hRad50 protein complex and Nijmegen breakage syndrome: Linkage of double-strand break repair to the cellular DNA damage response [J].
Carney, JP ;
Maser, RS ;
Olivares, H ;
Davis, EM ;
Le Beau, M ;
Yates, JR ;
Hays, L ;
Morgan, WF ;
Petrini, JHJ .
CELL, 1998, 93 (03) :477-486