Spleen necrosis virus-derived C-type retroviral vectors for gene transfer to quiescent cells

Abstract

Gene therapy applications of retroviral vectors derived from C-type retroviruses have been limited to introducing genes into dividing target cells. Here, we report genetically engineered C-type retroviral vectors derived from spleen necrosis virus (SNV), which are capable of infecting nondividing cells. This has been achieved by introducing a nuclear localization signal (NLS) sequence into the matrix protein (MA) of SNV by site-directed mutagenesis. This increased the efficiency of infecting nondividing cells and was sufficient to endow the virus with the capability to efficiently infect growth-arrested human T lymphocytes and quiescent primary monocyte-derived macrophages. We demonstrate that this vector actively penetrates the nucleus of a target cell, and has potential use as a gene therapy vector to transfer genes into nondividing cells.

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Figure 1: Plasmid constructs.
Figure 2: Nuclear localization sequence and SNV MA mutants.
Figure 3: Experimental system.
Figure 4: FACS analysis of the DNA content of D17, C8166, or H9 cells.
Figure 5: Experiments to test for the presence of viral DNA in the nuclei of infected quiescent cells.
Figure 6: Infection of human blood monocyte-derived macrophages.

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Acknowledgements

We would like to thank Dr. M. Schnell at Thomas Jefferson University for helpful comments on this manuscript. This research was supported by funds obtained from the National Institutes of Health (grant number 1RO1 AI 41899–01).

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Correspondence to Ralph Dornburg.

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Parveen, Z., Krupetsky, A., Engelstädter, M. et al. Spleen necrosis virus-derived C-type retroviral vectors for gene transfer to quiescent cells. Nat Biotechnol 18, 623–629 (2000). https://doi.org/10.1038/76458

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