Related Experiment Videos
DNA-Dependent protein kinase is not required for efficient lentivirus integration
V Baekelandt1, A Claeys, P Cherepanov
1Laboratory for Experimental Neurosurgery and Neuroanatomy, Katholieke Universiteit Leuven, Leuven, Belgium.
Journal of Virology
|November 9, 2000
Summary
DNA-dependent protein kinase (DNA-PK) and poly(ADP-ribose) polymerase (PARP) are not essential for lentivirus integration. DNA-PK may protect cells from toxicity at high retroviral vector titers.
Area of Science:
- Molecular Biology
- Virology
- Cellular Biology
Background:
- Retroviral DNA integration is a critical step in viral replication, but the host cell factors involved are not fully understood.
- DNA-dependent protein kinase (DNA-PK) has been implicated in repairing DNA double-strand breaks and potentially in retroviral integration.
- Previous studies suggested a role for DNA-PK in retroviral integration, with reduced efficiency and increased cell death observed in DNA-PK-deficient cells.
Purpose of the Study:
- To investigate the role of DNA-dependent protein kinase (DNA-PK) in lentivirus integration using a human immunodeficiency virus type 1 (HIV-1)-derived lentivirus vector.
- To determine if poly(ADP-ribose) polymerase (PARP) is essential for lentivirus transduction or HIV-1 replication.
Main Methods:
- Lentivirus transduction assays were performed using scid mouse embryonic fibroblasts (MEF) and DNA-PK-deficient cells (xrs-5 and xrs-6).
- Transduction efficiency was measured at various vector titers (multiplicity of infection [MOI]).
- The effect of a PARP inhibitor (3-methoxybenzamide) on lentivirus transduction and HIV-1 replication was assessed.
Main Results:
- Efficient and stable lentivirus transduction was achieved in DNA-PK-deficient cells at low vector titers, indicating DNA-PK is not essential for integration.
- Transduction efficiency was even higher in DNA-PK-deficient cells than in control cells at MOI <1 TU/cell.
- Increased cell death was observed in DNA-PK-deficient cells only at high vector titers (>1 TU/cell), suggesting a protective role for DNA-PK against retroviral toxicity.
- PARP inhibition did not affect lentivirus transduction or HIV-1 replication.
Conclusions:
- DNA-dependent protein kinase (DNA-PK) and poly(ADP-ribose) polymerase (PARP) are not essential for the integration of lentivirus vectors.
- DNA-PK may play a protective role against cellular toxicity induced by high levels of retroviral integrase or integration.