A block in virus-like particle maturation following assembly of murine leukaemia virus in insect cells.
Expression of the murine leukaemia virus (MLV) major Gag antigen p65(Gag) using the baculovirus expression system leads to efficient assembly and release of virus-like particles (VLP) representative of immature MLV. Expression of p180(Gag-Pol), facilitated normally in mammalian cells by readthrough of the p65(Gag) termination codon, also occurs efficiently in insect cells to provide a source of the MLV protease and a pattern of p65(Gag) processing similar to that observed in mammalian cells. VLP release from p180(Gag-Pol)-expressing cells however remains essentially immature with disproportionate levels of the uncleaved p65(Gag) precursor when compared to the intracellular Gag profile. Changing the p65(Gag) termination codon altered the level of p65(Gag) and p180(Gag-Pol) within expressing cells but did not alter the pattern of released VLP, which remained immature. Coexpression of p65(Gag) with a fixed readthrough p180(Gag-Pol) also led to only immature VLP release despite high intracellular protease levels. Our data suggest a mechanism that preferentially selects uncleaved p65(Gag) for the assembly of MLV in this heterologous expression system and implies that, in addition to their relative levels, active sorting of the correct p65(Gag) and p180(Gag-Pol) ratios may occur in producer cells.
Adamson
Catherine S
CS
School of Animal and Microbial Sciences, The University of Reading, Reading, RD6 6AJ, UK. c.s.adamson@rdg.ac.uk
Davies
Adam
A
Soneoka
Yuko
Y
Nermut
Milan
M
Mitrophanous
Kyriacos
K
Jones
Ian M
IM
D000818:Animals; D002478:Cells, Cultured; D015685:Fusion Proteins, gag-pol / Q000378:metabolism; D015683:Gene Products, gag / Q000378:metabolism; D009052:Leukemia Virus, Murine / Q000235:genetics / Q000378:metabolism*; D008854:Microscopy, Electron; D017924:Nucleopolyhedroviruses / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera / Q000821:virology; D014771:Virion / Q000378:metabolism* / Q000648:ultrastructure; D019065:Virus Assembly
D015685:Fusion Proteins, gag-pol; D015683:Gene Products, gag