A microarray to measure repair of damaged plasmids by cell lysates

被引：36

作者：

Millau, J. -F. ^{[1
]}

Raffin, A. -L. ^{[1
]}

Caillat, S. ^{[1
]}

Claudet, C. ^{[1
]}

Arras, G. ^{[2
]}

Ugolin, N. ^{[2
]}

Douki, T. ^{[1
]}

Ravanat, J. -L. ^{[1
]}

Breton, J. ^{[1
]}

Oddos, T. ^{[3
]}

Dumontet, C. ^{[6
,7
]}

Sarasin, A. ^{[4
,5
]}

Chevillard, S. ^{[2
]}

Favier, A. ^{[1
]}

Sauvaigo, S. ^{[1
]}

机构：

[1] CEA Grenoble, INAC, LCIB,UMR E 3,CEA,UJF, Lab Les Acides Nuclei, F-38054 Grenoble 9, France

[2] CEA, DSV, DRR, Expt Cancerol Lab, F-92265 Fontenay Aux Roses, France

[3] Johnson & Johnson Consumer Europe, F-27100 Val De Reuil, France

[4] Inst Gustave Roussy, CNRS, FRE 2939, F-94805 Villejuif, France

[5] Univ Paris 11, F-94805 Villejuif, France

[6] INSERM, U590, F-69008 Lyon, France

[7] Univ Lyon, LCMT, F-69003 Lyon, France

来源：

LAB ON A CHIP | 2008年 / 8卷 / 10期

关键词：

D O I：

10.1039/b806634e

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

DNA repair mechanisms constitute major defences against agents that cause cancer, degenerative disease and aging. Different repair systems cooperate to maintain the integrity of genetic information. Investigations of DNA repair involvement in human pathology require an efficient tool that takes into account the variety and complexity of repair systems. We have developed a highly sensitive damaged plasmid microarray to quantify cell lysate excision/synthesis (ES) capacities using small amounts of proteins. This microsystem is based on efficient immobilization and conservation on hydrogel coated glass slides of plasmid DNA damaged with a panel of genotoxic agents. Fluorescent signals are generated from incorporation of labelled dNTPs by DNA excision-repair synthesis mechanisms at plasmid sites. Highly precise DNA repair phenotypes i.e. simultaneous quantitative measures of ES capacities toward seven lesions repaired by distinct repair pathways, are obtained. Applied to the characterization of xeroderma pigmentosum (XP) cells at basal level and in response to a low dose of UVB irradiation, the assay showed the multifunctional role of different XP proteins in cell protection against all types of damage. On the other hand, measurement of the ES of peripheral blood mononuclear cells from six donors revealed significant diversity between individuals. Our results illustrate the power of such a parallelized approach with high potential for several applications including the discovery of new cancer biomarkers and the screening of chemical agents modulating DNA repair systems.

引用

页码：1713 / 1722

页数：10

共 44 条

[11] ACCURATE TRANSCRIPTION INITIATION BY RNA POLYMERASE-II IN A SOLUBLE EXTRACT FROM ISOLATED MAMMALIAN NUCLEI [J].

DIGNAM, JD ;

LEBOVITZ, RM ;

ROEDER, RG .

NUCLEIC ACIDS RESEARCH, 1983, 11 (05) :1475-1489

[12] Emerging cancer therapeutic opportunities target DNA-repair systems [J].

Ding, Jian ;

Miao, Ze-Hong ;

Meng, Ling-Hua ;

Geng, Mei-Yu .

TRENDS IN PHARMACOLOGICAL SCIENCES, 2006, 27 (06) :338-344

[13] Possible involvement of XPA in repair of oxidative DNA damage deduced from analysis of damage, repair and genotype in a human population study [J].

Dusinska, Maria ;

Dzupinkova, Zuzana ;

Wsolova, Ladislava ;

Harrington, Vikki ;

Collins, Andrew R. .

MUTAGENESIS, 2006, 21 (03) :205-211

[14]

GAIVAO I, 2007, CELL BIOL TOXICOL

[15] Repair of oxidative damage of thymine by HeLa whole-cell extracts: simultaneous analysis using a microsupport and comparison with traditional PAGE analysis [J].

Guerniou, V ;

Rapin, D ;

Millau, JF ;

Bufflier, E ;

Favier, A ;

Cadet, J ;

Sauvaigo, S .

BIOCHIMIE, 2005, 87 (02) :151-159

[16] Recognition of DNA adducts by human nucleotide excision repair - Evidence for a thermodynamic probing mechanism [J].

Gunz, D ;

Hess, MT ;

Naegeli, H .

JOURNAL OF BIOLOGICAL CHEMISTRY, 1996, 271 (41) :25089-25098

[17] Functional characterization of global genomic DNA repair and its implications for cancer [J].

Hanawalt, PC ;

Ford, JA ;

Lloyd, DR .

MUTATION RESEARCH-REVIEWS IN MUTATION RESEARCH, 2003, 544 (2-3) :107-114

[18] Age-dependent DNA repair and cell cycle distribution of human skin fibroblasts in response to UVA irradiation [J].

Hazane, F ;

Sauvaigo, S ;

Douki, T ;

Favier, A ;

Beani, JC .

JOURNAL OF PHOTOCHEMISTRY AND PHOTOBIOLOGY B-BIOLOGY, 2006, 82 (03) :214-223

[19] Recognition of cisplatin adducts by cellular proteins [J].

Kartalou, M ;

Essigmann, JM .

MUTATION RESEARCH-FUNDAMENTAL AND MOLECULAR MECHANISMS OF MUTAGENESIS, 2001, 478 (1-2) :1-21

[20]

Kraemer K H, 1985, Clin Dermatol, V3, P33, DOI 10.1016/0738-081X(85)90096-3

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