HIV-2 Immature Particle Morphology Provides Insights into Gag Lattice Stability and Virus Maturation.

Retrovirus immature particle morphology consists of a membrane enclosed, pleomorphic, spherical and incomplete lattice of Gag hexamers. Previously, we demonstrated that human immunodeficiency virus type 2 (HIV-2) immature particles possess a distinct and extensive Gag lattice morphology. To better understand the nature of the continuously curved hexagonal Gag lattice, we have used the single particle cryo-electron microscopy method to determine the HIV-2 Gag lattice structure for immature virions.

Human Immunodeficiency Virus Type 2 Capsid Protein Mutagenesis Reveals Amino Acid Residues Important for Virus Particle Assembly.

Human immunodeficiency virus (HIV) Gag drives virus particle assembly. The capsid (CA) domain is critical for Gag multimerization mediated by protein-protein interactions. The Gag protein interaction network defines critical aspects of the retroviral lifecycle at steps such as particle assembly and maturation.

Molecular Biology and Diversification of Human Retroviruses.

Studies of retroviruses have led to many extraordinary discoveries that have advanced our understanding of not only human diseases, but also molecular biology as a whole. The most recognizable human retrovirus, human immunodeficiency virus type 1 (HIV-1), is the causative agent of the global AIDS epidemic and has been extensively studied. Other human retroviruses, such as human immunodeficiency virus type 2 (HIV-2) and human T-cell leukemia virus type 1 (HTLV-1), have received less attention, and many of the assumptions about the replication and biology of these viruses are based on knowledge of HIV-1.

Distinct Antiretroviral Mechanisms Elicited by a Viral Mutagen.

5-aza-cytidine (5-aza-C) has been shown to be a potent human immunodeficiency virus type 1 (HIV-1) mutagen that induces G-to-C hypermutagenesis by incorporation of the reduced form (i.e., 5-aza-dC, 5-aza-dCTP).

Inhibiting HTLV-1 Protease: A Viable Antiviral Target.

Human T-cell lymphotropic virus type 1 (HTLV-1) is a retrovirus that can cause severe paralytic neurologic disease and immune disorders as well as cancer. An estimated 20 million people worldwide are infected with HTLV-1, with prevalence reaching 30% in some parts of the world. In stark contrast to HIV-1, no direct acting antivirals (DAAs) exist against HTLV-1.

Slow Receptor Binding of the Noncytopathic HIV-2 Envs Is Balanced by Long-Lived Activation State and Efficient Fusion Activity.

Many HIV strains downregulate the levels of CD4 receptor on the surface of infected cells to prevent superinfection. In contrast, the rare HIV-2 strain is noncytopathic and has no effect on CD4 expression in infected cells but still replicates as efficiently as more cytopathic strains in peripheral blood mononuclear cells (PBMCs). Here, we show that HIV-2 Env interactions with the CD4 receptor exhibit slow association kinetics, whereas the dissociation kinetics is within the range of cytopathic strains.

Anisotropic ESCRT-III architecture governs helical membrane tube formation.

ESCRT-III proteins assemble into ubiquitous membrane-remodeling polymers during many cellular processes. Here we describe the structure of helical membrane tubes that are scaffolded by bundled ESCRT-III filaments. Cryo-ET reveals how the shape of the helical membrane tube arises from the assembly of two distinct bundles of helical filaments that have the same helical path but bind the membrane with different interfaces.

Membrane constriction and thinning by sequential ESCRT-III polymerization.

The endosomal sorting complexes required for transport (ESCRTs) mediate diverse membrane remodeling events. These typically require ESCRT-III proteins to stabilize negatively curved membranes; however, recent work has indicated that certain ESCRT-IIIs also participate in positive-curvature membrane-shaping reactions. ESCRT-IIIs polymerize into membrane-binding filaments, but the structural basis for negative versus positive membrane remodeling by these proteins remains poorly understood.

Structural inhibition of dynamin-mediated membrane fission by endophilin.

Dynamin, which mediates membrane fission during endocytosis, binds endophilin and other members of the in-mphiphysin-vs (BAR) protein family. How endophilin influences endocytic membrane fission is still unclear. Here, we show that dynamin-mediated membrane fission is potently inhibited in vitro when an excess of endophilin co-assembles with dynamin around membrane tubules.

Structure and membrane remodeling activity of ESCRT-III helical polymers.

The endosomal sorting complexes required for transport (ESCRT) proteins mediate fundamental membrane remodeling events that require stabilizing negative membrane curvature. These include endosomal intralumenal vesicle formation, HIV budding, nuclear envelope closure, and cytokinetic abscission. ESCRT-III subunits perform key roles in these processes by changing conformation and polymerizing into membrane-remodeling filaments.

Structural and functional studies of membrane remodeling machines.

Building cells from their component parts will hinge upon our ability to reconstitute biochemical compartmentalization and exchange between membrane-delimited organelles. By contrast with our understanding of other cellular events, the mechanisms that govern membrane trafficking has lagged because the presence of phospholipid bilayers complicates the use of standard methods. This chapter describes in vitro methods for purifying, reconstituting, and visualizing membrane remodeling activities directly by electron cryomicroscopy.