Abstract
TRIM proteins play important roles in the innate immune defense against retroviral infection, including human immunodeficiency virus type-1 (HIV-1). Rhesus macaque TRIM5α (TRIM5αrh) targets the HIV-1 capsid and blocks infection at an early post-entry stage, prior to reverse transcription. Studies have shown that binding of TRIM5α to the assembled capsid is essential for restriction and requires the coiled-coil and B30.2/SPRY domains, but the molecular mechanism of restriction is not fully understood. In this study, we investigated, by cryoEM combined with mutagenesis and chemical cross-linking, the direct interactions between HIV-1 capsid protein (CA) assemblies and purified TRIM5αrh containing coiled-coil and SPRY domains (CC-SPRYrh). Concentration-dependent binding of CC-SPRYrh to CA assemblies was observed, while under equivalent conditions the human protein did not bind. Importantly, CC-SPRYrh, but not its human counterpart, disrupted CA tubes in a non-random fashion, releasing fragments of protofilaments consisting of CA hexamers without dissociation into monomers. Furthermore, such structural destruction was prevented by inter-hexamer crosslinking using P207C/T216C mutant CA with disulfide bonds at the CTD-CTD trimer interface of capsid assemblies, but not by intra-hexamer crosslinking via A14C/E45C at the NTD-NTD interface. The same disruption effect by TRIM5αrh on the inter-hexamer interfaces also occurred with purified intact HIV-1 cores. These results provide insights concerning how TRIM5α disrupts the virion core and demonstrate that structural damage of the viral capsid by TRIM5α is likely one of the important components of the mechanism of TRIM5α-mediated HIV-1 restriction.
Highlights
TRIM5a is an important component of the innate immune defense against retroviral infection, including human immunodeficiency virus type -1 (HIV-1) [1,2], and numerous studies suggest that TRIM5a interacts with assembled capsids and induces premature capsid disassembly, before reverse transcription takes place [3,4,5,6]
We show that direct binding of a TRIM5a protein, consisting of the coiled-coil and B30.2/SPRY domains, to the viral capsid results in disruption of the surface lattice and fragmentation of the capsid, at inter-hexamer interfaces
Our results reinforce the notion that structural damage of the viral capsid by TRIM5a is central to the mechanism of TRIM5amediated HIV-1 restriction
Summary
TRIM5a is an important component of the innate immune defense against retroviral infection, including human immunodeficiency virus type -1 (HIV-1) [1,2], and numerous studies suggest that TRIM5a interacts with assembled capsids and induces premature capsid disassembly (uncoating), before reverse transcription takes place [3,4,5,6]. TRIM5a is a 56 kD protein comprising a tripartite motif (TRIM; with RING, B-box 2, and coiled-coil (CC) domains) followed by a C-terminal B30.2/SPRY domain [7,8,9]. Each of these domains plays distinct roles in the antiviral function of TRIM5a. Proteasome activity is necessary for the TRIM5a-mediated block to reverse transcription [27], and engagement of restrictionsensitive virus cores results in proteasome-dependent degradation of TRIM5a [28] Together, these data suggest that TRIM5a action in host restriction of retroviruses involves all of its domains
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