HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 279, Issue 25, 26797-26801, June 18, 2004

This Article Full Text Full Text (PDF) All Versions of this Article: 279/25/26797    most recent M402832200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Blanco, M. G. Articles by Gómez-Márquez, J. Search for Related Content PubMed PubMed Citation Articles by Blanco, M. G. Articles by Gómez-Márquez, J. Social Bookmarking                      What's this?

A Paradox in the in Vitro End-joining Assays^*

Miguel G. Blanco, Francisco Boán, and Jaime Gómez-Márquez¶

From the Departamento de Bioquímica y Biología Molecular, Facultad de Biología, Universidad de Santiago de Compostela, 15782 Santiago de Compostela, A Coruña, Spain

Much work has been focused on the pathways that restore the integrity of the genome after different kinds of lesions, especially double-strand breaks. A classical method to investigate double-strand break repair is the incubation of a DNA substrate with cell-free extracts. In these end-joining assays, the DNA is efficiently ligated by the proteins present in the extract, generating circular molecules and/or multimers. In contrast, using a similar in vitro system, we detected DNA cleavage rather than end ligation. When comparing our results with previous works, a paradox emerges: lower amounts of DNA become multimerized instead of degraded and higher amounts of DNA are degraded rather than multimerized. Here, we have demonstrated that when the DNA/protein ratio is low enough, the DNA-binding proteins of the nuclear extract protect the DNA substrate, avoiding DNA degradation and vice versa. Therefore, the variation of the DNA/protein ratio is enough to switch the outcome of the experiment from a DNA cleavage assay to a typical end-joining assay.

Received for publication, March 12, 2004 , and in revised form, April 6, 2004.

^* This work was supported by Grant BMC2001-3242 from the Spanish Ministerio de Ciencia y Tecnología. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Both authors contributed equally to this work.

Recipient of a Formación de Profesorado Universitario grant from the Spanish Ministerio de Educación, Cultura y Deporte.

^¶ To whom correspondence should be addressed. Tel.: 34-981563100; Fax: 34-981596904; E-mail: bnjgm{at}usc.es.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				B. Rosidi, M. Wang, W. Wu, A. Sharma, H. Wang, and G. Iliakis Histone H1 functions as a stimulatory factor in backup pathways of NHEJ Nucleic Acids Res., March 1, 2008; 36(5): 1610 - 1623. [Abstract] [Full Text] [PDF]

				P. Burton, D. J. McBride, J. M. Wilkes, J. D. Barry, and R. McCulloch Ku Heterodimer-Independent End Joining in Trypanosoma brucei Cell Extracts Relies upon Sequence Microhomology Eukaryot. Cell, October 1, 2007; 6(10): 1773 - 1781. [Abstract] [Full Text] [PDF]

				S. Kuhfittig-Kulle, E. Feldmann, A. Odersky, A. Kuliczkowska, W. Goedecke, A. Eggert, and P. Pfeiffer The mutagenic potential of non-homologous end joining in the absence of the NHEJ core factors Ku70/80, DNA-PKcs and XRCC4-LigIV Mutagenesis, May 1, 2007; 22(3): 217 - 233. [Abstract] [Full Text] [PDF]

				L. Liang, L. Deng, Y. Chen, G. C. Li, C. Shao, and J. A. Tischfield Modulation of DNA End Joining by Nuclear Proteins J. Biol. Chem., September 9, 2005; 280(36): 31442 - 31449. [Abstract] [Full Text] [PDF]