Abstract
HIV-1 buds form infected cells in an immature, non-infectious form. Maturation into an infectious virion requires proteolytic cleavage of the Gag polyprotein at five positions, leading to a dramatic change in virus morphology. Immature virions contain an incomplete spherical shell where Gag is arranged with the N-terminal MA domain adjacent to the membrane, the CA domain adopting a hexameric lattice below the membrane, and beneath this, the NC domain and viral RNA forming a disordered layer. After maturation, NC and RNA are condensed within the particle surrounded by a conical CA core. Little is known about the sequence of structural changes that take place during maturation, however. Here we have used cryo-electron tomography and subtomogram averaging to resolve the structure of the Gag lattice in a panel of viruses containing point mutations abolishing cleavage at individual or multiple Gag cleavage sites. These studies describe the structural intermediates correlating with the ordered processing events that occur during the HIV-1 maturation process. After the first cleavage between SP1 and NC, the condensed NC-RNA may retain a link to the remaining Gag lattice. Initiation of disassembly of the immature Gag lattice requires cleavage to occur on both sides of CA-SP1, while assembly of the mature core also requires cleavage of SP1 from CA.
Highlights
The assembly of HIV-1 occurs at the plasma membrane of infected cells
We used cryo-electron tomography and sub-tomogram averaging to visualise the arrangement of Gag in 3D
We show that the fastest cleavage event leads to condensation of the RNA genome complexed with viral proteins. This inner RNA/protein structure appears to maintain a link with the remaining Gag lattice
Summary
The assembly of HIV-1 occurs at the plasma membrane of infected cells. The primary structural component of the assembling virus is the 55-kDa polyprotein Gag. Multiple copies of Gag assemble to form an incomplete sphere underneath the plasma membrane, which recruits components of the cellular ESCRT machinery to mediate membrane scission and release of the budding virus from the cell [1,2,3]. Gag consists of three major structural components: MA, the membrane binding domain, CA, the capsid domain, and NC, the nucleocapsid domain, which interacts with the viral RNA. The viral genome is recruited to the assembling particle from the pool of cellular mRNA via interactions between a packaging signal (Y) present at the 59 end of the genome [7] and zinc-fingers present in NC [4]
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