Betaretroviral Envelope Subunits Are Noncovalently Associated and Restricted to the Mammalian Class

Abstract

The structure of the transmembrane subunit (TM) of the retroviral envelope glycoprotein (Env) is highly conserved among most retrovirus genera and includes a pair of cysteines that forms an intramolecular disulfide loop within the ectodomain. Alpha-, gamma-, and deltaretroviruses have a third cysteine, adjacent to the loop, which forms a disulfide bond between TM and the surface subunit (SU) of Env, while lentiviruses, which have noncovalently associated subunits, lack this third cysteine. The Betaretrovirus genus includes Jaagsiekte sheep retrovirus (JSRV) and mouse mammary tumor virus (MMTV), as well as many endogenous retroviruses. Envelope subunit association had not been characterized in the betaretroviruses, but lack of a third cysteine in the TM ectodomain suggested noncovalently associated subunits. We tested the Env proteins of JSRV and MMTV, as well as human endogenous retrovirus K (HERV-K)108 -a betaretrovirus-like human endogenous retrovirus-for intersubunit bonding and found that, as in the lentiviruses, the Env subunits lack an intersubunit disulfide bond. Since these results suggest that the number of cysteines in the TM loop region readily distinguishes between covalent and noncovalent structure, we surveyed endogenous retroviral TM sequences in the genomes of vertebrates represented in public databases and found that (i) retroviruses with noncovalently associated subunits have been present during all of anthropoid evolution and (ii) the noncovalent env motif is limited to mammals, while the covalent type is found among five vertebrate classes. We discuss implications of these findings for retroviral evolution, cross-species transmissions, and recombination events involving the env gene. R etrovirus infection is mediated by the envelope (Env) glycoprotein, which is a class I viral fusion protein (VFP) in all of the orthoretroviral genera except the epsilonretroviruses. Class I VFPs are synthesized as precursors that trimerize in the endoplasmic reticulum and are cleaved by cellular furin in the late Golgi apparatus to generate two subunits: a surface subunit (SU), which interacts with the receptor on target cells, and a transmembrane subunit (TM), which mediates fusion and is anchored in the viral envelope near its C terminus. After cleavage, SU and TM remain associated either noncovalently or via an intersubunit disulfide bond (1-3, 82, 83), resulting in trimers of heterodimers that traffic to the cell's plasma membrane to become part of the viral envelope when newly synthesized particles bud from the cell (4).During infection, the fusion subunits of class I VFPs go through a series of conformational changes that lead from a metastable to a stable state. The first conformational change, which occurs upon binding of SU to the receptor, exposes a hydrophobic region at the N terminus of TM-the fusion peptide-which inserts into the plasma membrane of the target cell. This step then triggers the formation by TM of a highly stable six-helix bundle, a trimeric hairpin structure built around alpha-helic...

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“…1). There is a conserved immunosuppressive domain (ISD) and a CX 6 CC motif, typical of gammaretroviral and alpharetroviral, but not betaretroviral, TM proteins (4,5).…”

“…While SU is the most variable region of the genome, TM is highly conserved and can be aligned across a wide variety of retroviruses (4). The majority of retroviral envelope glycoproteins can be divided into those with Env subunits that are covalently associated (gamma-type) and those with noncovalently associated subunits (beta-type), and the two groups can be readily distinguished by sequence motifs in TM (5,6) (Fig. 1).…”

“…1). Sequences representing the noncovalent Env type (typical of betaretroviruses and lentiviruses) were found only in mammals, while covalent type sequences (typical of gammaretroviruses) were found among species representing five vertebrate classes (5).…”

Crystal Structures of Beta- and Gammaretrovirus Fusion Proteins Reveal a Role for Electrostatic Stapling in Viral Entry

“…Retroviral envelope glycoproteins may be divided into two types: ones that contain a disulfide bond between the SU and TM subunits and ones that are noncovalently associated (27). Covalent-type retroviral Env contains a conserved immunosuppressive domain (ISD) and a CX 6 CC motif in the chain reversal region of its fusion subunit.…”

“…5). The betaretrovirus genus is unique from the others, as examples of both covalent-type and non-covalent-type viral glycoproteins are observed (27). For instance, MPMV Env, which contains a CX 6 CC motif, is thought to be derived from a gammaretrovirus through a recombination event.…”