Effects of blocking individual maturation cleavages in murine leukemia virus gag.
A single protein, termed Gag, is responsible for retrovirus particle assembly. After the assembled virion is released from the cell, Gag is cleaved at several sites by the viral protease (PR). The cleavages catalyzed by PR bring about a wide variety of physical changes in the particle, collectively termed maturation, and convert the particle into an infectious virion. In murine leukemia virus (MLV) maturation, Gag is cleaved at three sites, resulting in formation of the matrix (MA), p12, capsid (CA), and nucleocapsid (NC) proteins. We introduced mutations into MLV that inhibited cleavage at individual sites in Gag. All mutants had lost the intensely staining ring characteristic of immature particles; thus, no single cleavage event is required for this feature of maturation. Mutant virions in which MA was not cleaved from p12 were still infectious, with a specific infectivity only approximately 10-fold below that of the wild type. Particles in which p12 and CA could not be separated from each other were noninfectious and lacked a well-delineated core despite the presence of dense material in their interiors. In both of these mutants, the dimeric viral RNA had undergone the stabilization normally associated with maturation, suggesting that this change may depend upon the separation of CA from NC. Alteration of the C-terminal end of CA blocked CA-NC cleavage but also reduced the efficiency of particle formation and, in some cases, severely disrupted the ability of Gag to assemble into regular structures. This observation highlights the critical role of this region of Gag in assembly.
Oshima
Masamichi
M
HIV Drug Resistance Program. Image Analysis Laboratory, SAIC Frederick, National Cancer Institute-Frederick, Frederick, Maryland 21702-1201, USA.
Muriaux
Delphine
D
Mirro
Jane
J
Nagashima
Kunio
K
Dryden
Kelly
K
Yeager
Mark
M
Rein
Alan
A
D000818:Animals; D002460:Cell Line; D003902:Detergents / Q000494:pharmacology; D010450:Endopeptidases / Q000378:metabolism; D015683:Gene Products, gag / Q000378:metabolism*; D006801:Humans; D009052:Leukemia Virus, Murine / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D051379:Mice; D008854:Microscopy, Electron; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D041681:NIH 3T3 Cells; D014771:Virion / Q000378:metabolism / Q000472:pathogenicity* / Q000648:ultrastructure; D019065:Virus Assembly
D003902:Detergents; D015683:Gene Products, gag; D010450:Endopeptidases