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Journal of Virology, December 2000, p. 11278-11285, Vol. 74, No. 23
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

DNA-Dependent Protein Kinase Is Not Required for Efficient Lentivirus Integration

Veerle Baekelandt,1 Anje Claeys,2 Peter Cherepanov,2 Erik De Clercq,2 Bart De Strooper,3,4 Bart Nuttin,1 and Zeger Debyser2,*

Rega Institute for Medical Research,2 Laboratory for Experimental Neurosurgery and Neuroanatomy,1 Center for Human Genetics,3 and Flanders Interuniversitary Institute for Biotechnology,4 Katholieke Universiteit Leuven, Leuven, Belgium

Received 26 April 2000/Accepted 26 July 2000

How DNA is repaired after retrovirus integration is not well understood. DNA-dependent protein kinase (DNA-PK) is known to play a central role in the repair of double-stranded DNA breaks. Recently, a role for DNA-PK in retroviral DNA integration has been proposed (R. Daniel, R. A. Katz, and A. M. Skalka, Science 284:644-647, 1999). Reduced transduction efficiency and increased cell death by apoptosis were observed upon retrovirus infection of cultured scid cells. We have used a human immunodeficiency virus (HIV) type 1 (HIV-1)-derived lentivirus vector system to further investigate the role of DNA-PK during integration. We measured lentivirus transduction of scid mouse embryonic fibroblasts (MEF) and xrs-5 or xrs-6 cells. These cells are deficient in the catalytic subunit of DNA-PK and in Ku, the DNA-binding subunit of DNA-PK, respectively. At low vector titers, efficient and stable lentivirus transduction was obtained, excluding an essential role for DNA-PK in lentivirus integration. Likewise, the efficiency of transduction of HIV-derived vectors in scid mouse brain was as efficient as that in control mice, without evidence of apoptosis. We observed increased cell death in scid MEF and xrs-5 or xrs-6 cells, but only after transduction with high vector titers (multiplicity of infection [MOI], >1 transducing unit [TU]/cell) and subsequent passage of the transduced cells. At an MOI of <1 TU/cell, however, transduction efficiency was even higher in DNA-PK-deficient cells than in control cells. Taken together, the data suggest a protective role of DNA-PK against cellular toxicity induced by high levels of retrovirus integrase or integration. Another candidate cellular enzyme that has been claimed to play an important role during retrovirus integration is poly(ADP-ribose) polymerase (PARP). However, no inhibition of lentivirus vector-mediated transduction or HIV-1 replication by 3-methoxybenzamide, a known PARP inhibitor, was observed. In conclusion, DNA-PK and PARP are not essential for lentivirus integration.

* Corresponding author. Mailing address: Rega Institute for Medical Research, Minderbroedersstr. 10, B-3000 Leuven, Belgium. Phone: 32 16 33 21 82. Fax: 32 16 33 21 31. E-mail: zeger.debyser{at}uz.kuleuven.ac.be.

Journal of Virology, December 2000, p. 11278-11285, Vol. 74, No. 23
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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