Abstract:The restriction factor TRIM5α binds to the capsid protein of the retroviral core and blocks retroviral replication. The affinity of TRIM5α for the capsid is a major host tropism determinant of HIV and other primate immunodeficiency viruses, but the molecular interface involved in this host–pathogen interaction remains poorly characterized. Here we use NMR spectroscopy to investigate binding of the rhesus TRIM5α SPRY domain to a selection of HIV capsid constructs. The data are consistent with a model in which o… Show more
“…For example, oligomerization is essential for the ability of rhesus monkey TRIM5␣ proteins to bind to the HIV-1 core. In the case of TRIM5␣ proteins, oligomerization provides the necessary avidity for the SPRY domain to interact with capsid (32)(33)(34)(35)(36). In agreement with this, oligomerization of the restriction factor MxA is important for its ability to block viral infection (37).…”
The alpha interferon (IFN-␣)-inducible restriction factor myxovirus B (MxB) blocks HIV-1 infection after reverse transcription but prior to integration. MxB binds to the HIV-1 core, which is composed of capsid protein, and this interaction leads to inhibition of the uncoating process of HIV-1. Previous studies suggested that HIV-1 restriction by MxB requires binding to capsid. This work tests the hypothesis that MxB oligomerization is important for the ability of MxB to bind to the HIV-1 core. For this purpose, we modeled the structure of MxB using the published tertiary structure of MxA. The modeled structure of MxB guided our mutagenic studies and led to the discovery of several MxB variants that lose the capacity to oligomerize. In agreement with our hypothesis, MxB variants that lost the oligomerization capacity also lost the ability to bind to the HIV-1 core. MxB variants deficient for oligomerization were not able to block HIV-1 infection. Overall, our work showed that oligomerization is required for the ability of MxB to bind to the HIV-1 core and block HIV-1 infection.
IMPORTANCEMxB is a novel restriction factor that blocks infection of HIV-1. MxB is inducible by IFN-␣, particularly in T cells. The current work studies the oligomerization determinants of MxB and carefully explores the contribution of oligomerization to capsid binding and restriction. This work takes advantage of the current structure of MxA and models the structure of MxB, which is used to guide structure-function studies. This work leads to the conclusion that MxB oligomerization is important for HIV-1 capsid binding and restriction.
“…For example, oligomerization is essential for the ability of rhesus monkey TRIM5␣ proteins to bind to the HIV-1 core. In the case of TRIM5␣ proteins, oligomerization provides the necessary avidity for the SPRY domain to interact with capsid (32)(33)(34)(35)(36). In agreement with this, oligomerization of the restriction factor MxA is important for its ability to block viral infection (37).…”
The alpha interferon (IFN-␣)-inducible restriction factor myxovirus B (MxB) blocks HIV-1 infection after reverse transcription but prior to integration. MxB binds to the HIV-1 core, which is composed of capsid protein, and this interaction leads to inhibition of the uncoating process of HIV-1. Previous studies suggested that HIV-1 restriction by MxB requires binding to capsid. This work tests the hypothesis that MxB oligomerization is important for the ability of MxB to bind to the HIV-1 core. For this purpose, we modeled the structure of MxB using the published tertiary structure of MxA. The modeled structure of MxB guided our mutagenic studies and led to the discovery of several MxB variants that lose the capacity to oligomerize. In agreement with our hypothesis, MxB variants that lost the oligomerization capacity also lost the ability to bind to the HIV-1 core. MxB variants deficient for oligomerization were not able to block HIV-1 infection. Overall, our work showed that oligomerization is required for the ability of MxB to bind to the HIV-1 core and block HIV-1 infection.
IMPORTANCEMxB is a novel restriction factor that blocks infection of HIV-1. MxB is inducible by IFN-␣, particularly in T cells. The current work studies the oligomerization determinants of MxB and carefully explores the contribution of oligomerization to capsid binding and restriction. This work takes advantage of the current structure of MxA and models the structure of MxB, which is used to guide structure-function studies. This work leads to the conclusion that MxB oligomerization is important for HIV-1 capsid binding and restriction.
“…The strong tendency toward hydrophobic collapse observed in the C-terminal segment of v1 may be functionally important because mutation of five consecutive residues (NFNYC) in the C-terminus of v1 to alanine displays a very strong defect in HIV restriction and capsid binding. 8,14 …”
The binding of the TRIM5α restriction factor to the HIV capsid is mediated by the C-terminal SPRY domain of TRIM5α. Atomic-level details of this host-pathogen interaction, which involves mobile variable loops of the SPRY domain, remain unclear. Some of the key determinants of restriction are encompassed by the long and disordered v1 loop of the SPRY domain. We applied molecular modeling to elucidate the conformational repertoire of the v1 loop and its role in the interaction with the capsid. All-atom replica exchange molecular dynamics revealed multiple transient, interconverting states of the v1 loop consistent with the intrinsic disorder observed experimentally. The docking of the SPRY conformations representing 10 most populated states onto the high-resolution model of the assembled HIV-1 capsid revealed that a subset of v1 conformations produced plausible binding poses, in which the SPRY domain binds close to the pseudo-2-fold symmetry axis and the v1 loop spans the interhexamer gap. Such binding mode is well supported by the NMR binding data and known escape mutants. We speculate that the binding mode that involves interaction of the capsid with a subset of preexisting SPRY conformations arising from the intrinsic disorder of the v1 loop may explain the remarkable ability of TRIM5α to resist viral evasion by mutagenesis and to restrict divergent retroviruses.
“…The B-box 2 and coiled-coil domains make an integrated antiparallel dimer fold (Goldstone et al, 2014; Sanchez et al, 2014; Weinert et al, 2015), which acts as a scaffold that organizes the upstream and downstream domains (Figure 1B). In TRIM5α, the C-terminal domain is a β-sandwich fold called SPRY (or PRYSPRY/B30.2), which mediates direct binding to retroviral capsids (Biris et al, 2012; 2013; Diaz-Griffero et al, 2006b; Kovalskyy and Ivanov, 2014; Sawyer et al, 2005; Sayah et al, 2004; Sebastian and Luban, 2005; Stremlau et al, 2006; Yang et al, 2012). The L2 linker that connects the SPRY domain to the coiled-coil packs against the coiled-coil scaffold, and so in the TRIM5α dimer, two SPRY domains are oriented to bind the capsid simultaneously (Figure 1B) (Goldstone et al, 2014; Li et al, 2014; Sanchez et al, 2014; Weinert et al, 2015).…”
Section: Introductionmentioning
confidence: 99%
“…Higher-order assembly of TRIM5α requires the B-box 2 domain, which is thought to mediate three-fold symmetric interactions that connect coiled-coil mediated TRIM5α dimers into a hexagonal net (Diaz-Griffero et al, 2009; Ganser-Pornillos et al, 2011; Javanbakht et al, 2005; Li and Sodroski, 2008; Li et al, 2016). This hexagonal scaffold is proposed to position the SPRY domains to match the orientations – both translational and rotational – of their corresponding binding epitopes on retroviral capsids, and thereby generate powerful avidity effects that amplify very weak (millimolar level (Biris et al, 2013)) intrinsic affinities between the SPRY and capsid subunits.10.7554/eLife.16309.003Figure 1.Design and oligomeric behavior of miniTRIM proteins.( A ) Schematic of the TRIM5α primary sequence. ( B ) Schematic of the antiparallel full-length dimer.…”
Restriction factors and pattern recognition receptors are important components of intrinsic cellular defenses against viral infection. Mammalian TRIM5α proteins are restriction factors and receptors that target the capsid cores of retroviruses and activate ubiquitin-dependent antiviral responses upon capsid recognition. Here, we report crystallographic and functional studies of the TRIM5α B-box 2 domain, which mediates higher-order assembly of TRIM5 proteins. The B-box can form both dimers and trimers, and the trimers can link multiple TRIM5α proteins into a hexagonal net that matches the lattice arrangement of capsid subunits and enables avid capsid binding. Two modes of conformational flexibility allow TRIM5α to accommodate the variable curvature of retroviral capsids. B-box mediated interactions also modulate TRIM5α’s E3 ubiquitin ligase activity, by stereochemically restricting how the N-terminal RING domain can dimerize. Overall, these studies define important molecular details of cellular recognition of retroviruses, and how recognition links to downstream processes to disable the virus.DOI:
http://dx.doi.org/10.7554/eLife.16309.001
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