Restriction of HIV-1 by rhesus TRIM5alpha is governed by alpha-helices in the Linker2 region Journal Article

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Authors: Sastri, J.; Johnsen, L.; Smolin, N; Imam, S; Mukherjee, S; Lukic, Z.; Brandariz-Nunez, A.; Robia, S. L.; Diaz-Griffero, F; Wiethoff, C.; Campbell, E. M.
Article Title: Restriction of HIV-1 by rhesus TRIM5alpha is governed by alpha-helices in the Linker2 region
Abstract: TRIM5alpha proteins are a potent barrier to the cross-species transmission of retroviruses. TRIM5alpha proteins exhibit an ability to self-associate at many levels, ultimately leading to the formation of protein assemblies with hexagonal symmetry in vitro and cytoplasmic assemblies when expressed in cells. However, the role of these assemblies in restriction, the determinants that mediate their formation and the organization of TRIM5alpha molecules within these assemblies have remained unclear. Here we show that alpha-helical elements within the Linker2 region of rhesus macaque TRIM5alpha govern the ability to form cytoplasmic assemblies in cells and restrict HIV-1 infection. Mutations which reduce alpha-helix formation by the Linker2 region disrupt assembly and restriction. More importantly, mutations that enhance the alpha-helical content of the Linker2 region, relative to wild type protein, also exhibit an increased ability to form cytoplasmic assemblies and restrict HIV-1 infection. Molecular modeling of the TRIM5alpha dimer suggests a model in which alpha-helical elements within the Linker2 region dock to alpha-helices of the coiled-coil domain, likely establishing proper orientation and spacing of protein domains necessary for assembly and restriction. Collectively, these studies provide critical insight into the determinants governing TRIM5alpha assembly and restriction and demonstrate that the antiviral potency of TRIM5alpha proteins can be significantly increased without altering the affinity of SPRY/capsid binding. IMPORTANCE: Many members of the TRIpartite Motif (TRIM) family of proteins act as restriction factors which directly inhibit viral infection and activate innate immune signaling pathways. Another common feature of TRIM proteins is the ability to form protein assemblies in the nucleus or the cytoplasm. However, the determinants in TRIM proteins required for assembly and the degree to which assembly affects TRIM protein function has been poorly understood. Here, we show that alpha helices in the Linker2 region of rhesus TRIM5alpha govern assembly and restriction of HIV-1 infection. Helix disrupting mutations disrupt assembly and restriction of HIV-1, while helix stabilizing mutations enhance assembly and restriction relative to wild type protein. Circular dichroism analysis suggests that that the formation of this helical structure is supported by intermolecular interactions with the coiled-coil domain in the CC-L2 dimer. These studies reveal a novel mechanism by which the antiviral activity of TRIM5alpha proteins can be regulated and provides detailed insight into the assembly determinants of TRIM family proteins.
Journal Title: Journal of virology
Volume: 88
Issue: 16
ISSN: 1098-5514; 0022-538X
Publisher: American Society for Microbiology. All Rights Reserved  
Date Published: 2014
Start Page: 8911
End Page: 8923
Language: ENG
DOI/URL:

JVI.01134-14
Notes: LR: 20150408; CI: Copyright (c) 2014; GR: 1G20RR030939/RR/NCRR NIH HHS/United States; GR: R01 AI087390/AI/NIAID NIH HHS/United States; GR: R01 AI093258/AI/NIAID NIH HHS/United States; GR: R01 AI093258/AI/NIAID NIH HHS/United States; GR: R01 HL106189/HL/NHLBI NIH HHS/United States; JID: 0113724; 0 (Carrier Proteins); 0 (Chemokine CCL2); OID: NLM: PMC4136267; 2014/05/28 [aheadofprint]; ppublish