TY - JOUR
T1 - Differential effects of C-terminal molecular tagged integrase on replication competent Moloney murine leukemia virus
AU - Seamon, J. A.
AU - Adams, M.
AU - Sengupta, S.
AU - Roth, M. J.
N1 - Funding Information:
This research is supported by National Institutes of Health (NIH) Grant R21-DK54374 and R01CA76545 to M.J.R. J.S. is supported in part by NIH Predoctoral Training Grants T32-GM8360 and T32-AI7403. We thank Christina Miller for her assistance.
PY - 2000/9/1
Y1 - 2000/9/1
N2 - Moloney murine leukemia virus (M-MuLV) proviruses carrying integrase (IN) protein tagged either with a simian virus 40 (SV40) nuclear localization signal (NLS) or various antigenic epitopes were generated. Hexahistidine (His6), hemagluttinin (HA), or two consecutive HA sequences (2XHA) were fused to the C-terminus of IN as antigenic markers. These epitope-tagged IN proteins were stably expressed through multiple rounds of infection. The IN-His6, IN-HA, and IN-2XHA proteins, purified from virus, could be immunoprecipitated with antibodies against His6 and HA, respectively. An M-MuLV provirus encoding the SV40 large T antigen NLS fused to IN at the same position as the epitope tags was also passaged through cells. In contrast to the stability of the epitope tags, the SV40 NLS sequence was rapidly mutated by a frameshift mutation that introduced negatively charged amino acids into the basic NLS. The instability of the NLS suggests that the strong nuclear localization of the IN-SV40 NLS may have detrimental effects on virus assembly. These observations have implications for studying nuclear transport properties of M-MuLV and for engineering a murine-based retroviral vector for gene therapy. (C) 2000 Academic Press.
AB - Moloney murine leukemia virus (M-MuLV) proviruses carrying integrase (IN) protein tagged either with a simian virus 40 (SV40) nuclear localization signal (NLS) or various antigenic epitopes were generated. Hexahistidine (His6), hemagluttinin (HA), or two consecutive HA sequences (2XHA) were fused to the C-terminus of IN as antigenic markers. These epitope-tagged IN proteins were stably expressed through multiple rounds of infection. The IN-His6, IN-HA, and IN-2XHA proteins, purified from virus, could be immunoprecipitated with antibodies against His6 and HA, respectively. An M-MuLV provirus encoding the SV40 large T antigen NLS fused to IN at the same position as the epitope tags was also passaged through cells. In contrast to the stability of the epitope tags, the SV40 NLS sequence was rapidly mutated by a frameshift mutation that introduced negatively charged amino acids into the basic NLS. The instability of the NLS suggests that the strong nuclear localization of the IN-SV40 NLS may have detrimental effects on virus assembly. These observations have implications for studying nuclear transport properties of M-MuLV and for engineering a murine-based retroviral vector for gene therapy. (C) 2000 Academic Press.
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U2 - 10.1006/viro.2000.0481
DO - 10.1006/viro.2000.0481
M3 - Article
C2 - 10964783
AN - SCOPUS:0034285069
SN - 0042-6822
VL - 274
SP - 412
EP - 419
JO - Virology
JF - Virology
IS - 2
ER -