Inserting a nuclear targeting signal into a replication-competent Moloney murine leukemia virus affects viral export and is not sufficient for cell cycle-independent infection.
The effects of inserting reported nuclear localization signals (NLSs) into the Moloney murine leukemia virus (Mo-MuLV) integrase (IN) protein, within a replication-competent viral construct, were studied. In contrast to the virus harboring IN fused to the simian virus 40 (SV40) large T antigen NLS (SV40 NLS) (J. A. Seamon, M. Adams, S. Sengupta, and M. J. Roth, Virology 274:412-419, 2000), a codon-modified SV40 NLS was stably expressed during viral propagation. Incorporation of the codon-modified SV40 NLS into IN, however, altered the packaging of the Gag-Pol precursor in the virus; viral particles contained decreased levels of reverse transcriptase (RT) and IN. In addition, the virus showed delayed kinetics of viral DNA synthesis upon infection. A panel of infectious MuLVs containing alternative IN-NLS fusions was generated and assayed for cell cycle-independent infection. Viral infection with the NLS-tagged proteins, however, remained dependent on passage of the cells through mitosis. This finding has direct implications for engineering murine-based retroviral vectors for gene therapy.
Department of Biochemistry, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Piscataway 08854, USA.
Jennifer A
JA
Seamon
Kathryn S
KS
Jones
Christina
C
Miller
Monica J
MJ
Roth
D000595:Amino Acid Sequence; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics; D001483:Base Sequence; D002453:Cell Cycle; D002460:Cell Line; D004261:DNA Replication / Q000235:genetics; D004279:DNA, Viral / Q000096:biosynthesis / Q000235:genetics; D004285:Dogs; D015316:Genetic Therapy; D005822:Genetic Vectors; D019426:Integrases / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D008979:Moloney murine leukemia virus / Q000235:genetics* / Q000472:pathogenicity / Q000502:physiology*; D019913:Nuclear Localization Signals / Q000235:genetics*; D014162:Transfection; D016662:Virus Integration / Q000235:genetics; D014779:Virus Replication / Q000235:genetics
D000952:Antigens, Polyomavirus Transforming; D004279:DNA, Viral; D019913:Nuclear Localization Signals; D019426:Integrases