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Nucleic Acids Research Advance Access published online on November 10, 2006
Nucleic Acids Research, doi:10.1093/nar/gkl890
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© 2006 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Molecular Biology

DAXX interacts with phage {Phi}C31 integrase and inhibits recombination

Jin-zhong Chen1, Chao-neng Ji1, Guan-lan Xu1, Rong-yan Pang1, Ji-hua Yao1, Huan-zhang Zhu1, Jing-lun Xue1,* and William Jia1,2,*

1 State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Sciences, Fudan University China 2 Department of Surgery, University of British Columbia Vancouver, Canada

*To whom correspondence should be addressed. Tel/Fax: +86 21 65649899; Email: Jlxue{at}fudan.ac.cn

Received July 10, 2006. Revised October 8, 2006. Accepted October 9, 2006.

Phage {Phi}C31 integrase has potential as a means of inserting therapeutic genes into specific sites in the human genome. However, the possible interactions between {Phi}C31 integrase and cellular proteins have never been investigated. Using pLexA-{Phi}C31 integrase as bait, we screened a pB42AD-human fetal brain cDNA library for potential interacting cellular proteins. Among 61 positives isolated from 106 independent clones, 51 contained DAXX C-terminal fragments. The strong interaction between DAXX and {Phi}C31 was further confirmed by co-immunoprecipitation. Deletion analysis revealed that the fas-binding domain of DAXX is also the region for {Phi}C31 binding. Hybridization between a {Phi}C31 integrase peptide array and an HEK293 cell extract revealed that a tetramer, 451RFGK454, in the C-terminus of {Phi}C31 is responsible for the interaction with DAXX. This tetramer is also necessary for {Phi}C31 integrase activity as removal of this tetramer resulted in a complete loss of integrase activity. Co-expression of DAXX with {Phi}C31 integrase in a HEK293-derived {Phi}C31 integrase activity reporter cell line significantly reduced the {Phi}C31-mediated recombination rate. Knocking down DAXX with a DAXX-specific duplex RNA resulted in increased recombination efficiency. Therefore, endogenous DAXX may interact with {Phi}C31 causing a mild inhibition in the integration efficiency. This is the first time that {Phi}C31 was shown to interact with an important cellular protein and the potential effect of this interaction should be further studied.

*Correspondence may also be addressed to William Jia. Tel: +604 822 0728; Fax: +604 322 0640; Email: wjia{at}interchange.ubc.ca


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[Abstract] [Full Text] [PDF]


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