title: Conservation of a stepwise, energy-sensitive pathway involving HP68 for assembly of primate lentivirus capsids in cells.
abstract: Previously we have described a stepwise, energy-dependent pathway for human immunodeficiency virus type 1 (HIV-1) capsid assembly in a cell-free system. In this pathway, Gag polypeptides utilize the cellular factor HP68 and assemble into immature capsids by way of assembly intermediates that have defined biochemical characteristics. Here we address whether this pathway is universally conserved among primate lentiviruses and can be observed in mammalian cells. We demonstrate that HIV-2 Gag associates with human HP68 in a cell-free system and that Gag proteins of HIV-2, simian immunodeficiency virus SIVmac239, and SIVagm associate with endogenous HP68 in primate cells, as is seen for HIV-1. Analysis of primate cells expressing lentivirus Gag proteins revealed Gag-containing complexes with the same sedimentation values as seen for previously described HIV-1 assembly intermediates in the cell-free system (10S, 80-150S, and 500S). These complexes fit criteria for assembly intermediates as judged by energy sensitivity, pattern of HP68 association, and the failure of specific complexes to be formed by assembly-incompetent Gag mutants. We also demonstrate that virus-like particles released from cells do not appear to contain HP68, suggesting that HP68 is released from Gag upon completion of capsid assembly in cells, as was observed previously in the cell-free system. Together these findings support a model in which all primate lentivirus capsids assemble by a conserved pathway of HP68-containing, energy-dependent assembly intermediates that have specific biochemical features.
authors.affiliation: Department of Pathobiology, University of Washington, Seattle, Washington 98195, USA.
authors.forename: Julia E
authors.initials: JE
authors.lastname: Dooher
authors.forename: Jaisri R
authors.initials: JR
authors.lastname: Lingappa
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D019556:COS Cells; D002213:Capsid / Q000378:metabolism*; D002474:Cell-Free System; D018833:Chaperonins / Q000378:metabolism*; D002522:Chlorocebus aethiops; D015683:Gene Products, gag / Q000235:genetics / Q000378:metabolism*; D015498:HIV-2 / Q000378:metabolism; D006801:Humans; D008251:Macaca; D011499:Protein Processing, Post-Translational; D015302:Simian Immunodeficiency Virus / Q000378:metabolism*; D019065:Virus Assembly
chemical_list: C501917:ABCE1 protein, human; D018528:ATP-Binding Cassette Transporters; D015683:Gene Products, gag; D000255:Adenosine Triphosphate; D018833:Chaperonins