Recombinant replication-restricted VSV as an expression vector for murine cytokines.
Vesicular stomatitis virus (VSV) is a prototypic non-segmented, negative-strand RNA virus that rapidly and efficiently shuts down the production of host cell-encoded proteins and utilizes the cell's protein production machinery to express high levels of virally encoded proteins. In an effort to take advantage of this characteristic of VSV, we have employed a reverse genetics system to create recombinant forms of VSV encoding a variety of murine cytokines. Previous studies have revealed that cells infected with recombinant VSV that lack expression of the surface glycoprotein (G protein), designated deltaG-VSV, more efficiently express and secrete recombinant proteins than do recombinant "wild-type" VSV. Therefore, murine cytokine-expressing recombinants were produced as deltaG viruses. Propagation of these deltaG viruses in cells that transiently express G protein in vitro results in G-complemented virions that can infect cells, shut down host protein synthesis, and express at high levels each virally encoded protein (including the designated cytokine). We assessed the ability of each deltaG-VSV construct to express recombinant cytokine by infecting BHK cells and then monitoring/measuring the production of the desired cytokine. When possible, the bioactivity of the cytokine products was also measured. The results presented here reveal that large quantities of bioactive cytokines can be produced rapidly and inexpensively using deltaG-VSV as a protein expression system.
Miller
Mark A
MA
Department of Molecular Sciences, University of Tennessee Health Science Center, 858 Madison Avenue, Memphis, TN 38163, USA. mamiller@utmem.edu
Lavine
Christy L
CL
Klas
Sheri D
SD
Pfeffer
Lawrence M
LM
Whitt
Michael A
MA
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D016207:Cytokines / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D016679:Genome, Viral; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D011995:Recombination, Genetic; D014721:Vesicular stomatitis Indiana virus / Q000235:genetics* / Q000378:metabolism* / Q000472:pathogenicity / Q000502:physiology; D014779:Virus Replication
D016207:Cytokines; D011993:Recombinant Fusion Proteins