Research Article: Global Rescue of Defects in HIV-1 Envelope Glycoprotein Incorporation: Implications for Matrix Structure

Date Published: November 14, 2013

Publisher: Public Library of Science

Author(s): Philip R. Tedbury, Sherimay D. Ablan, Eric O. Freed, Jeremy Luban.

http://doi.org/10.1371/journal.ppat.1003739

Abstract

The matrix (MA) domain of HIV-1 Gag plays key roles in membrane targeting of Gag, and envelope (Env) glycoprotein incorporation into virions. Although a trimeric MA structure has been available since 1996, evidence for functional MA trimers has been elusive. The mechanism of HIV-1 Env recruitment into virions likewise remains unclear. Here, we identify a point mutation in MA that rescues the Env incorporation defects imposed by an extensive panel of MA and Env mutations. Mapping the mutations onto the putative MA trimer reveals that the incorporation-defective mutations cluster at the tips of the trimer, around the perimeter of a putative gap in the MA lattice into which the cytoplasmic tail of gp41 could insert. By contrast, the rescue mutation is located at the trimer interface, suggesting that it may confer rescue of Env incorporation via modification of MA trimer interactions, a hypothesis consistent with additional mutational analysis. These data strongly support the existence of MA trimers in the immature Gag lattice and demonstrate that rescue of Env incorporation defects is mediated by modified interactions at the MA trimer interface. The data support the hypothesis that mutations in MA that block Env incorporation do so by imposing a steric clash with the gp41 cytoplasmic tail, rather than by disrupting a specific MA-gp41 interaction. The importance of the trimer interface in rescuing Env incorporation suggests that the trimeric arrangement of MA may be a critical factor in permitting incorporation of Env into the Gag lattice.

Partial Text

Human immunodeficiency virus type 1 (HIV-1), like all replication-competent orthoretroviruses, encodes three main polyproteins – Gag, Pol and Env – which contain determinants necessary for particle assembly, enzymatic functions, and virus entry, respectively. HIV-1 assembly occurs in a series of steps, driven by the Gag precursor protein Pr55Gag (for review, see [1]). HIV-1 Gag is comprised of four domains – matrix (MA), capsid (CA), nucleocapsid (NC) and p6 – and two spacer peptides located between CA and NC, and NC and p6. Pr55Gag is able to form virus-like particles (VLPs) when expressed in cells in the absence of any other viral protein. The MA domain at the N-terminus of Pr55Gag directs cytoplasmic Gag to bind raft-like domains of the plasma membrane (PM) via specific recognition of phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] [2] (for review, [3]). MA binding to PI(4,5)P2, as well as Gag oligomerization, triggers a myristyl switch, exposing the myristic acid moiety covalently linked to the amino-terminus of MA [4], [5]. The exposed myristic acid then inserts into the phospholipid bilayer, anchoring Gag to the PM.

Various models can be invoked to explain the incorporation of Env into HIV-1 particles, the principle unresolved issues being whether or not Env is actively recruited into virions and if so, whether Env interacts directly with Gag or indirectly via a bridging cellular factor [6]. Evidence to support active recruitment is provided by five observations. First, the existence of mutations in MA and Env that prevent Env incorporation is consistent with a direct recruitment and suggests the presence of interacting motifs, although it does not address the question of whether the interaction is direct or indirect. Secondly, it has been reported that HIV-1 Env is retained on immature particles even after removal of the viral membrane with detergent [43]. This retention is dependent on the long gp41 CT, again consistent with an interaction between the CT and Gag. Third, there have been reports of interaction between Gag and Env in cells and with recombinant proteins in vitro[37]–[39]; however, this interaction has been difficult to demonstrate consistently. Fourth, Env has been reported to influence the site of virus budding in polarized epithelial cells and T cells [44], [45]. Finally, the observation that Gag processing (virus maturation) affects Env fusogenicity is consistent with cross-talk between Gag and the CT of gp41 [46]–[48].

 

Source:

http://doi.org/10.1371/journal.ppat.1003739

 

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