Matthew Stremlau

The cytoplasmic body component TRIM5α restricts HIV-1 infection in Old World monkeys

Host cell barriers to the early phase of immunodeficiency virus replication explain the current distribution of these viruses among human and non-human primate species. Human immunodeficiency virus type 1 (HIV-1), the cause of acquired immunodeficiency syndrome (AIDS) in humans, efficiently enters the cells of Old World monkeys but encounters a block before reverse transcription. This species-specific restriction acts on the incoming HIV-1 capsid and is mediated by a dominant repressive factor. Here we identify TRIM5α, a component of cytoplasmic bodies, as the blocking factor. HIV-1 infection is restricted more efficiently by rhesus monkey TRIM5α than by human TRIM5α. The simian immunodeficiency virus, which naturally infects Old World monkeys, is less susceptible to the TRIM5α-mediated block than is HIV-1, and this difference in susceptibility is due to the viral capsid. The early block to HIV-1 infection in monkey cells is relieved by interference with TRIM5α expression. Our studies identify TRIM5α as a species-specific mediator of innate cellular resistance to HIV-1 and reveal host cell components that modulate the uncoating of a retroviral capsid.

Species-Specific Variation in the B30.2(SPRY) Domain of TRIM5α Determines the Potency of Human Immunodeficiency Virus Restriction

ABSTRACT Retroviruses encounter dominant postentry restrictions in cells of particular species. Human immunodeficiency virus type 1 (HIV-1) is blocked in the cells of Old World monkeys by TRIM5α, a tripartite motif (TRIM) protein composed of RING, B-box 2, coiled-coil, and B30.2(SPRY) domains. Rhesus monkey TRIM5α (TRIM5αrh) more potently blocks HIV-1 infection than human TRIM5α (TRIM5αhu). Here, by studying chimeric TRIM5α proteins, we demonstrate that the major determinant of anti-HIV-1 potency is the B30.2(SPRY) domain. Analysis of species-specific variation in TRIM5α has identified three variable regions (v1, v2, and v3) within the B30.2 domain. The TRIM5α proteins of Old World primates exhibit expansion, duplication, and residue variation specifically in the v1 region. Replacement of three amino acids in the N terminus of the TRIM5αhu B30.2 v1 region with the corresponding TRIM5αrh residues resulted in a TRIM5α molecule that restricted HIV-1 nearly as efficiently as wild-type TRIM5αrh. Surprisingly, a single-amino-acid change in this region of TRIM5αhu allowed potent restriction of simian immunodeficiency virus, a phenotype not observed for either wild-type TRIM5αhu or TRIM5αrh. Some of the chimeric TRIM5α proteins that are >98% identical to the human protein yet mediate a strong restriction of HIV-1 infection may have therapeutic utility. These observations implicate the v1 variable region of the B30.2(SPRY) domain in TRIM5αrh antiviral potency.

Retrovirus Restriction by TRIM5α Variants from Old World and New World Primates

ABSTRACT The TRIM5α proteins of humans and some Old World monkeys have been shown to block infection of particular retroviruses following virus entry into the host cell. Infection of most New World monkey cells by the simian immunodeficiency virus of macaques (SIVmac) is restricted at a similar point. Here we examine the antiretroviral activity of TRIM5α orthologs from humans, apes, Old World monkeys, and New World monkeys. Chimpanzee and orangutan TRIM5α proteins functionally resembled human TRIM5α, potently restricting infection by N-tropic murine leukemia virus (N-MLV) and moderately restricting human immunodeficiency virus type 1 (HIV-1) infection. Notably, TRIM5α proteins from several New World monkey species restricted infection by SIVmac and the SIV of African green monkeys, SIVagm. Spider monkey TRIM5α, which has an expanded B30.2 domain v3 region due to a tandem triplication, potently blocked infection by a range of retroviruses, including SIVmac, SIVagm, HIV-1, and N-MLV. Tandem duplications in the TRIM5α B30.2 domain v1 region of African green monkeys are also associated with broader antiretroviral activity. Thus, variation in TRIM5α proteins among primate species accounts for the observed patterns of postentry restrictions in cells from these animals. The TRIM5α proteins of some monkey species exhibit dramatic lengthening of particular B30.2 variable regions and an expanded range of susceptible retroviruses.

The B30.2(SPRY) Domain of the Retroviral Restriction Factor TRIM5α Exhibits Lineage-Specific Length and Sequence Variation in Primates

ABSTRACT Tripartite motif (TRIM) proteins are composed of RING, B-box 2, and coiled coil domains. Some TRIM proteins, such as TRIM5α, also possess a carboxy-terminal B30.2(SPRY) domain and localize to cytoplasmic bodies. TRIM5α has recently been shown to mediate innate intracellular resistance to retroviruses, an activity dependent on the integrity of the B30.2 domain, in particular primate species. An examination of the sequences of several TRIM proteins related to TRIM5 revealed the existence of four variable regions (v1, v2, v3, and v4) in the B30.2 domain. Species-specific variation in TRIM5α was analyzed by amplifying, cloning, and sequencing nonhuman primate TRIM5 orthologs. Lineage-specific expansion and sequential duplication occurred in the TRIM5α B30.2 v1 region in Old World primates and in v3 in New World monkeys. We observed substitution patterns indicative of selection bordering these particular B30.2 domain variable elements. These results suggest that occasional, complex changes were incorporated into the TRIM5α B30.2 domain at discrete time points during the evolution of primates. Some of these time points correspond to periods during which primates were exposed to retroviral infections, based on the appearance of particular endogenous retroviruses in primate genomes. The results are consistent with a role for TRIM5α in innate immunity against retroviruses.

Retroviral Restriction Factor TRIM5α Is a Trimer

ABSTRACT The retrovirus restriction factor TRIM5α targets the viral capsid soon after entry. Here we show that the TRIM5α protein oligomerizes into trimers. The TRIM5α coiled-coil and B30.2(SPRY) domains make important contributions to the formation and/or stability of the trimers. A functionally defective TRIM5α mutant with the RING and B-box 2 domains deleted can form heterotrimers with wild-type TRIM5α, accounting for the observed dominant-negative activity of the mutant protein. Trimerization potentially allows TRIM5α to interact with threefold pseudosymmetrical structures on retroviral capsids.

Removal of Arginine 332 Allows Human TRIM5α To Bind Human Immunodeficiency Virus Capsids and To Restrict Infection

ABSTRACT Human TRIM5α (TRIM5αhu) only modestly inhibits human immunodeficiency virus type 1 (HIV-1) and does not inhibit simian immunodeficiency virus (SIVmac). Alteration of arginine 332 in the TRIM5αhu B30.2 domain to proline, the residue found in rhesus monkey TRIM5α, has been shown to create a potent restricting factor for both HIV-1 and SIVmac. Here we demonstrate that the potentiation of HIV-1 inhibition results from the removal of a positively charged residue at position 332 of TRIM5αhu. The increase in restricting activity correlated with an increase in the ability of TRIM5αhu mutants lacking arginine 332 to bind HIV-1 capsid complexes. A change in the cyclophilin A-binding loop of the HIV-1 capsid decreased TRIM5αhu R332P binding and allowed escape from restriction. The ability of TRIM5αhu to restrict SIVmac could be disrupted by the presence of any charged residue at position 332. Thus, charged residues in the v1 region of the TRIM5αhu B30.2 domain can modulate capsid binding and restriction potency. Therapeutic strategies designed to neutralize arginine 332 of TRIM5αhu might potentiate the innate resistance of human cells to HIV-1 infection.

The Human TRIM5α Restriction Factor Mediates Accelerated Uncoating of the N-Tropic Murine Leukemia Virus Capsid

ABSTRACT The host cell factors TRIM5αhu and Fv-1 restrict N-tropic murine leukemia virus (N-MLV) infection at an early postentry step before or after reverse transcription, respectively. Interestingly, the identity of residue 110 of the MLV capsid determines susceptibility to both TRIM5αhu and Fv-1. In this study, we investigate the fate of the MLV capsid in cells expressing either the TRIM5αhu or Fv-1 restriction factor. The expression of TRIM5αhu, but not Fv-1, specifically promoted the premature conversion of particulate N-MLV capsids within infected cells to soluble capsid proteins. The TRIM5αhu-mediated disassembly of particulate N-MLV capsids was dependent upon residue 110 of the viral capsid. Furthermore, the deletion or disruption of TRIM5αhu domains necessary for potent N-MLV restriction completely abrogated the disappearance of particulate N-MLV capsids observed with wild-type TRIM5αhu. These results suggest that premature disassembly of the viral capsid contributes to the restriction of N-MLV infection by TRIM5αhu, but not by Fv-1.

Binding and Susceptibility to Postentry Restriction Factors in Monkey Cells Are Specified by Distinct Regions of the Human Immunodeficiency Virus Type 1 Capsid

ABSTRACT In cells of Old World and some New World monkeys, dominant factors restrict human immunodeficiency virus type 1 (HIV-1) infections after virus entry. The simian immunodeficiency virus SIVmac is less susceptible to these restrictions, a property that is determined largely by the viral capsid protein. For this study, we altered exposed amino acid residues on the surface of the HIV-1 capsid, changing them to the corresponding residues found on the SIVmac capsid. We identified two distinct pathways of escape from early, postentry restriction in monkey cells. One set of mutants that were altered near the base of the cyclophilin A-binding loop of the N-terminal capsid domain or in the interdomain linker exhibited a decreased ability to bind the restricting factor(s). Consistent with the location of this putative factor-binding site, cyclophilin A and the restricting factor(s) cooperated to achieve the postentry block. A second set of mutants that were altered in the ridge formed by helices 3 and 6 of the N-terminal capsid domain efficiently bound the restricting factor(s) but were resistant to the consequences of factor binding. These results imply that binding of the simian restricting factor(s) is not sufficient to mediate the postentry block to HIV-1 and that SIVmac capsids escape the block by decreases in both factor binding and susceptibility to the effects of the factor(s).

Clinical Sequencing Uncovers Origins and Evolution of Lassa Virus

The 2013-2015 West African epidemic of Ebola virus disease (EVD) reminds us of how little is known about biosafety level 4 viruses. Like Ebola virus, Lassa virus (LASV) can cause hemorrhagic fever with high case fatality rates. We generated a genomic catalog of almost 200 LASV sequences from clinical and rodent reservoir samples. We show that whereas the 2013-2015 EVD epidemic is fueled by human-to-human transmissions, LASV infections mainly result from reservoir-to-human infections. We elucidated the spread of LASV across West Africa and show that this migration was accompanied by changes in LASV genome abundance, fatality rates, codon adaptation, and translational efficiency. By investigating intrahost evolution, we found that mutations accumulate in epitopes of viral surface proteins, suggesting selection for immune escape. This catalog will serve as a foundation for the development of vaccines and diagnostics. VIDEO ABSTRACT.

Mode of Transmission Affects the Sensitivity of Human Immunodeficiency Virus Type 1 to Restriction by Rhesus TRIM5α

ABSTRACT Rhesus TRIM5α (rhTRIM5α), but not human TRIM5α (huTRIM5α), potently inhibits human immunodeficiency virus (HIV) infection and is thus a potentially valuable therapeutic tool. Primary human CD4 T cells engineered to express rhTRIM5α were highly resistant to cell-free HIV type 1 (HIV-1) infection. However, when cocultured with unmodified T cells, rhTRIM5α-expressing cells became highly permissive to HIV-1 infection. Physical separation of rhTRIM5α-expressing cells and unmodified cells revealed that rhTRIM5α efficiently restricts cell-free but not cell-associated HIV transmission. Furthermore, we observed that HIV-infected human cells could infect rhesus CD4 T cells by cell-to-cell contact, but the infection was self-limiting. Subsequently, we noted that a spreading infection ensued when HIV-1-infected rhTRIM5α-expressing human cells were cultured with huTRIM5α- but not rhTRIM5α-expressing cells. Our results suggest that cell-associated HIV transmission in humans is blocked only when both donor and recipient cells express rhTRIM5α. These studies further define the role of rhTRIM5α in cell-free and cell-associated HIV transmission and delineate the utility of rhTRIM5α in anti-HIV therapy.