Basic Residues in Human Immunodeficiency Virus Type 1 Nucleocapsid Promote Virion Assembly via Interaction with RNA

Abstract

Retroviral Gag polyproteins drive virion assembly by polymerizing to form a spherical shell that lines the inner membrane of nascent virions. Deletion of the nucleocapsid (NC) domain of the Gag polyprotein disrupts assembly, presumably because NC is required for polymerization. Human immunodeficiency virus type 1 NC possesses two zinc finger motifs that are required for specific recognition and packaging of viral genomic RNA. Though essential, zinc fingers and genomic RNA are not required for virion assembly. NC promiscuously associates with cellular RNAs, many of which are incorporated into virions. It has been hypothesized that Gag polymerization and virion assembly are promoted by nonspecific interaction of NC with RNA. Consistent with this model, we found an inverse relationship between the number of NC basic residues replaced with alanine and NC's nonspecific RNA-binding activity, Gag's ability to polymerize in vitro and in vivo, and Gag's capacity to assemble virions. In contrast, mutation of NC's zinc fingers had only minor effects on these properties.

The Prion Protein Has RNA Binding and Chaperoning Properties Characteristic of Nucleocapsid Protein NCp7 of HIV-1

“…1A). NCp7 with the two zinc fingers and an NCp7 mutant, designated NCp7-(12-53), without the basic N-and C-terminal domains necessary for RNA dimerization in vitro and virus assembly in vivo (16,24,25), were generated by peptide synthesis (Fig. 1B and data not shown) (23).…”

“…In cells infected by HIV-1, the retroviral Gag polyprotein encoding NC is membrane-anchored via its N terminus during virion formation and budding at the cell surface (16,25,43,50). Therefore, it is tempting to speculate that Gag and PrP interact at the plasma membrane in infected cells.…”

In Vitro Identification and Characterization of an Early Complex Linking HIV-1 Genomic RNA Recognition and Pr55Gag Multimerization

“…Our results suggest that the 11th ␣-helix of CA and the putative CA/SP1 ␣-helix might bind to HIV-1 genomic RNA (mainly SL6-ECP) in a helix-loop-helix (HLH) fashion. These elements are comprised of a short ␣-helix (11th ␣-helix of CA) and a long ␣-helix (putative CA/SP1), connected by a flexible linker (78), that is usually followed by a highly basic domain involved in the protein: nucleic acid interaction (38,63). In this model, a Gag dimer would interact with the 5Ј-LR as shown in Fig.…”

The structural biology of HIV assembly

“…1b) forms the innermost layer of the immature virion and also plays essential roles in particle formation [21,23,51,[78][79][80][81]. NC/RNA complex(es) do not follow the hexagonal symmetry of the CA and SP1 regions, and most experiments indicate that NC primarily "tethers" Gag molecules together, probably via RNA bridges, although additional NC-NC interactions may also occur [79,80,82,83]. NC/RNA tethers presumably increase the effective concentration of assembling Gag molecules, which could also play a more active role in orienting or otherwise facilitating CA-CA interactions.…”