title: Basic residues in the nucleocapsid domain of Gag are required for interaction of HIV-1 gag with ABCE1 (HP68), a cellular protein important for HIV-1 capsid assembly.
abstract: During human immunodeficiency virus, type 1 (HIV-1) assembly, Gag polypeptides multimerize into immature HIV-1 capsids. The cellular ATP-binding protein ABCE1 (also called HP68 or RNase L inhibitor) appears to be critical for proper assembly of the HIV-1 capsid. In primate cells, ABCE1 associates with Gag polypeptides present in immature capsid assembly intermediates. Here we demonstrate that the NC domain of Gag is critical for interaction with endogenous primate ABCE1, whereas other domains in Gag can be deleted without eliminating the association of Gag with ABCE1. NC contains two Cys-His boxes that form zinc finger motifs and are responsible for encapsidation of HIV-1 genomic RNA. In addition, NC contains basic residues known to play a critical role in nonspecific RNA binding, Gag-Gag interactions, and particle formation. We demonstrate that basic residues in NC are needed for the Gag-ABCE1 interaction, whereas the cysteine and histidine residues in the zinc fingers are dispensable. Constructs that fail to interact with primate ABCE1 or interact poorly also fail to form capsids and are arrested at an early point in the immature capsid assembly pathway. Whereas others have shown that basic residues in NC bind nonspecifically to RNA, which in turn scaffolds or nucleates assembly, our data demonstrate that the same basic residues in NC act either directly or indirectly to recruit a cellular protein that also promotes capsid formation. Thus, in cells, basic residues in NC appear to act by two mechanisms, recruiting both RNA and a cellular ATPase in order to facilitate efficient assembly of HIV-1 capsids.
authors.affiliation: Department of Pathobiology, University of Washington, Seattle, 98195, USA. jais@u.washington.edu
authors.forename: Jaisri R
authors.initials: JR
authors.lastname: Lingappa
authors.forename: Julia E
authors.initials: JE
authors.lastname: Dooher
authors.forename: Michael A
authors.initials: MA
authors.lastname: Newman
authors.forename: Patti K
authors.initials: PK
authors.lastname: Kiser
authors.forename: Kevin C
authors.initials: KC
authors.lastname: Klein
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry*; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D018833:Chaperonins / Q000737:chemistry*; D002522:Chlorocebus aethiops; D015683:Gene Products, gag / Q000737:chemistry*; D015497:HIV-1 / Q000737:chemistry*; D006801:Humans; D008969:Molecular Sequence Data; D019251:Nucleocapsid / Q000737:chemistry*; D012259:Ribonuclease, Pancreatic / Q000494:pharmacology; D019065:Virus Assembly
chemical_list: C501917:ABCE1 protein, human; D018528:ATP-Binding Cassette Transporters; D015683:Gene Products, gag; D012259:Ribonuclease, Pancreatic; D018833:Chaperonins