In vitro cell-free conversion of noninfectious Moloney retrovirus particles to an infectious form by the addition of the vesicular stomatitis virus surrogate envelope G protein.
In the absence of envelope gene expression, retrovirus packaging cell lines expressing Moloney murine leukemia virus (MLV) gag and pol genes produce large amounts of noninfectious virus-like particles that contain reverse transcriptase, processed Gag protein, and viral RNA (gag-pol RNA particles). We demonstrate that these particles can be made infectious in an in vitro, cell-free system by the addition of a surrogate envelope protein, the G spike glycoprotein of vesicular stomatitis virus (VSV-G). The appearance of infectivity is accompanied by physical association of the G protein with the immature, noninfectious virus particles. Similarly, exposure in vitro of wild-type VSV-G to a fusion-defective pseudotyped virus containing a mutant VSV-G markedly increases the infectivity of the virus to titers similar to those of conventional VSV-G pseudotyped viruses. Furthermore, similar treatment of an amphotropic murine leukemia virus significantly allows infection of BHK cells not otherwise susceptible to infection with native amphotropic virus. The partially cell-free virus maturation system reported here should be useful for studies aimed at the preparation of tissue-targeted retrovirus vectors and will also aid in studies of nucleocapsid-envelope interactions during budding and of virus assembly and virus-receptor interactions during virus uptake into infected cells. It may also represent a potentially useful step toward the eventual development of a completely cell-free retrovirus assembly system.
Abe
A
A
Department of Pediatrics, Center for Molecular Genetics, University of California, San Diego, La Jolla, California 92093-0634, USA.
Chen
S T
ST
Miyanohara
A
A
Friedmann
T
T
D000818:Animals; D002460:Cell Line; D002461:Cell Line, Transformed; D002474:Cell-Free System; D006224:Cricetinae; D003470:Culture Media; D015685:Fusion Proteins, gag-pol / Q000235:genetics / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D008562:Membrane Glycoproteins; D008979:Moloney murine leukemia virus / Q000145:classification / Q000235:genetics / Q000472:pathogenicity*; D012367:RNA, Viral / Q000502:physiology; D014721:Vesicular stomatitis Indiana virus / Q000378:metabolism*; D014759:Viral Envelope Proteins / Q000378:metabolism*; D014771:Virion
D003470:Culture Media; D015685:Fusion Proteins, gag-pol; C028391:G protein, vesicular stomatitis virus; D008562:Membrane Glycoproteins; D012367:RNA, Viral; D014759:Viral Envelope Proteins