Publications by authors named "Akatsuki Saito"

45 Publications

Bovine respiratory coronavirus enhances bacterial adherence by upregulating expression of cellular receptors on bovine respiratory epithelial cells.

Vet Microbiol 2021 Apr 17;255:109017. Epub 2021 Feb 17.

Department of Veterinary Science, Faculty of Agriculture, University of Miyazaki, Miyazaki, 889-2192, Japan; Graduate School of Medicine and Veterinary Medicine, University of Miyazaki, Miyazaki, 889-2192, Japan; Center for Animal Disease Control, University of Miyazaki, Miyazaki, 889-2192, Japan. Electronic address:

Bovine coronavirus (BCoV) is one of the agents causing bovine respiratory disease complex (BRDC), with single infection tending to be mild to moderate; the probability of developing pneumonia in BRDC may be affected by viral and bacterial combinations. Previously, we reported that bovine respiratory syncytial virus (BRSV) infection enhances adherence of Pasteurella multocida (PM) to cells derived from the bovine lower respiratory tract but that BRSV infection in cells derived from the upper respiratory tract reduces PM adherence. In this study, we sought to clarify whether the modulation of bacterial adherence to cells derived from the bovine upper and lower respiratory tract is shared by other BRDC-related viruses by infecting bovine epithelial cells from the trachea, bronchus and lung with BCoV and/or PM. The results showed that cells derived from both the upper and lower respiratory tract were susceptible to BCoV infection. Furthermore, all cells infected with BCoV exhibited increased PM adherence via upregulation of two major bacterial adhesion molecules, intercellular adhesion molecule-1 (ICAM-1) and platelet-activating factor receptor (PAF-R), suggesting that compared with BRSV infection, BCoV infection differentially modulates bacterial adherence. In summary, we identified distinct interaction between bovine respiratory viruses and bacterial infections.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.vetmic.2021.109017DOI Listing
April 2021

Seroprevalence of Severe Fever with Thrombocytopenia Syndrome Virus in Small-Animal Veterinarians and Nurses in the Japanese Prefecture with the Highest Case Load.

Viruses 2021 02 2;13(2). Epub 2021 Feb 2.

Center for Animal Disease Control, University of Miyazaki, Miyazaki 889-2192, Japan.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is the causative agent of SFTS, an emerging tick-borne disease in East Asia, and is maintained in enzootic cycles involving ticks and a range of wild animal hosts. Direct transmission of SFTSV from cats and dogs to humans has been identified in Japan, suggesting that veterinarians and veterinary nurses involved in small-animal practice are at occupational risk of SFTSV infection. To characterize this risk, we performed a sero-epidemiological survey in small-animal-practice workers and healthy blood donors in Miyazaki prefecture, which is the prefecture with the highest per capita number of recorded cases of SFTS in Japan. Three small-animal-practice workers were identified as seropositive by ELISA, but one had a negative neutralization-test result and so was finally determined to be seronegative, giving a seropositive rate of 2.2% (2 of 90), which was significantly higher than that in healthy blood donors (0%, 0 of 1000; < 0.05). The seroprevalence identified here in small-animal-practice workers was slightly higher than that previously reported in other high-risk workers engaged in agriculture and forestry in Japan. Thus, enhancement of small-animal-practice workers' awareness of biosafety at animal hospitals is necessary for control of SFTSV.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/v13020229DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7912989PMC
February 2021

A Potent anti-Simian Immunodeficiency Virus Neutralizing Antibody Induction Associated with a Germline Immunoglobulin Gene Polymorphism in Rhesus Macaques.

J Virol 2021 Jan 13. Epub 2021 Jan 13.

AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan

Virus infection induces B cells with a wide variety of B cell receptor (BCR) repertoires. Patterns of induced BCR repertoires are different in individuals, while the underlying mechanism causing this difference remains largely unclear. In particular, the impact of germ line BCR immunoglobulin (Ig) gene polymorphism on B cell/antibody induction has not fully been determined. In the present study, we found a potent antibody induction associated with a germ line BCR Ig gene polymorphism. B404-class antibodies, which were previously reported as potent anti-simian immunodeficiency virus (SIV) neutralizing antibodies using the germ line VH3.33 gene-derived Ig heavy chain, were induced in five of 10 rhesus macaques after SIVsmH635FC infection. Investigation of VH3.33 genes in B404-class antibody inducers ( = 5) and non-inducers ( = 5) revealed association of B404-class antibody induction with a germ line VH3.33 polymorphism. Analysis of reconstructed antibodies indicated that the VH3.33 residue 38 is the determinant for B404-class antibody induction. B404-class antibodies were induced in all the macaques possessing the B404-associated VH3.33 allele, even under undetectable viremia. Our results show that a single nucleotide polymorphism in germ line VH genes could be a determinant for induction of potent antibodies against virus infection, implying that germ line VH-gene polymorphisms can be a factor restricting effective antibody induction or responsiveness to vaccination. Vaccines against a wide variety of infectious diseases have been developed mostly to induce antibodies targeting pathogens. However, small but significant percentage of people fail to mount potent antibody responses after vaccination, while the underlying mechanism of host failure in antibody induction remains largely unclear. In particular, the impact of germ line B cell receptor (BCR)/antibody immunoglobulin (Ig) gene polymorphism on B cell/antibody induction has not fully been determined. In the present study, we found a potent anti-simian immunodeficiency virus neutralizing antibody induction associated with a germ line BCR/antibody Ig gene polymorphism in rhesus macaques. Our results demonstrate that a single nucleotide polymorphism in germ line Ig genes could be a determinant for induction of potent antibodies against virus infection, implying that germ line BCR/antibody Ig gene polymorphisms can be a factor restricting effective antibody induction or responsiveness to vaccination.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.02455-20DOI Listing
January 2021

Bovine Respiratory Syncytial Virus Enhances the Adherence of to Bovine Lower Respiratory Tract Epithelial Cells by Upregulating the Platelet-Activating Factor Receptor.

Front Microbiol 2020 31;11:1676. Epub 2020 Jul 31.

Graduate School of Medicine and Veterinary Medicine, University of Miyazaki, Miyazaki, Japan.

Coinfection by bovine respiratory syncytial virus (BRSV) and (PM) frequently has been observed in cattle that develop severe pneumonia. We recently reported that BRSV infection significantly increased PM adherence to bovine lower respiratory tract epithelial cells. However, the molecular mechanisms of enhanced PM adherence are not completely understood. To investigate whether BRSV infection regulates any cellular adherence receptors on bovine bronchus- and lung-epithelial cells, we performed proteomic and functional analyses. The proteomic analysis showed that BRSV infection increased the accumulation of the platelet-activating factor receptor (PAFR) in both cell types. Molecular experiments, including specific blockade, knockdown, and overexpression of PAFR, indicated that PM adherence to these cell types depended on PAFR expression. These findings highlight the role, in cattle with severe pneumonia, of the synergistic effect of coinfection by BRSV and PM in the lower respiratory tract.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fmicb.2020.01676DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7411089PMC
July 2020

Rapid inactivation of SARS-CoV-2 with deep-UV LED irradiation.

Emerg Microbes Infect 2020 Dec;9(1):1744-1747

Department of Hemovascular Medicine and Artificial Organs, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.

The spread of novel coronavirus disease 2019 (COVID-19) infections worldwide has raised concerns about the prevention and control of SARS-CoV-2. Devices that rapidly inactivate viruses can reduce the chance of infection through aerosols and contact transmission. This study demonstrated that irradiation with a deep ultraviolet light-emitting diode (DUV-LED) of 280 ± 5 nm wavelength rapidly inactivates SARS-CoV-2 obtained from a COVID-19 patient. Development of devices equipped with DUV-LED is expected to prevent virus invasion through the air and after touching contaminated objects.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/22221751.2020.1796529DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7473214PMC
December 2020

Bovine Respiratory Syncytial Virus Decreased Pasteurella multocida Adherence by Downregulating the Expression of Intercellular Adhesion Molecule-1 on the Surface of Upper Respiratory Epithelial Cells.

Vet Microbiol 2020 Jul 2;246:108748. Epub 2020 Jun 2.

Graduate School of Medicine and Veterinary Medicine, University of Miyazaki, Miyazaki, Japan; Department of Veterinary Science, Faculty of Agriculture, University of Miyazaki, Miyazaki, Japan; Center for Animal Disease Control, University of Miyazaki, Miyazaki, Japan. Electronic address:

The synergistic infection of bovine respiratory syncytial virus (BRSV) and Pasteurella multocida (PM) may predispose cattle to develop severe pneumonia. Previously, we reported that BRSV infection significantly decreased PM adherence to the upper respiratory epithelial cells. It may allow bacteria to invade into the lower respiratory tract and lead to severe pneumonia. To investigate whether BRSV infection regulates the cell surface adherence receptor on bovine trachea epithelial cells (bTECs), we performed proteomic and functional analyses. BRSV infection decreased the expression of intercellular adhesion molecule-1 (ICAM1) on bTECs. Inhibition and knockdown experiments using anti-ICAM1 antibody and siRNAs targeting ICAM1 indicated that PM adherence to bTECs was dependent on ICAM1 expression. These data suggest that under normal conditions bTECs may capture PM in the upper respiratory tract, while BRSV infection reverses this mechanism. The proposed gateway function of bTECs is disrupted by BRSV infection that may facilitate bacterial invasion into the lower respiratory tract and lead to secondary or more severe respiratory infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.vetmic.2020.108748DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7265823PMC
July 2020

The 4th and 112th Residues of Viral Capsid Cooperatively Modulate Capsid-CPSF6 Interactions of HIV-1.

AIDS Res Hum Retroviruses 2020 06 17;36(6):513-521. Epub 2020 Feb 17.

Research Institute for Microbial Diseases, Osaka University, Osaka, Japan.

Binding of HIV-1 capsid (CA) to cleavage and polyadenylation specificity factor 6 (CPSF6) is hypothesized to provide a significant fitness advantage to viral replication, explaining why CA-CPSF6 interactions are strictly conserved in primate lentiviruses. We recently identified a Q4R mutation in CA after propagation of an interferon (IFN)-β-hypersensitive CA mutant, RGDA/Q112D (H87R, A88G, P90D, P93A and Q112D) virus, in IFN-β-treated cells. The Q4R substitution conferred significant IFN-β resistance to the RGDA/Q112D virus by affecting several properties of the virus, including the sensitivity to myxovirus resistance protein B (MxB), the kinetics of reverse transcription, and the initiation of uncoating. Notably, the Q4R substitution restored the CPSF6 interaction of the RGDA/Q112D virus. To better understand how the Q4R substitution modulated the CA-CPSF6 interaction, we generated a series of CA mutants harboring substitutions at the 4th and 112th residues. In contrast to the effect in the RGDA/Q112D background, the Q4R substitution diminished CA-CPSF6 interaction in an otherwise wild-type virus. Our genetic and structural analyses revealed that while either the Q4R or Q112D substitution impaired CA-CPSF6 interaction, the combination of these substitutions restored this interaction. These results suggest that the 4th and 112th residues in HIV-1 CA cooperatively modulate CA-CPSF6 interactions, further highlighting the tremendous levels of plasticity in primate lentivirus CA, which is one of the barriers to antiretroviral therapy in HIV-1-infected individuals.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1089/AID.2019.0250DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7262650PMC
June 2020

Discovery of a small molecule inhibitor targeting dengue virus NS5 RNA-dependent RNA polymerase.

PLoS Negl Trop Dis 2019 11 18;13(11):e0007894. Epub 2019 Nov 18.

Research Institute for Microbial Diseases, Osaka University, Osaka, Japan.

Dengue is a mosquito-borne viral infection that has spread globally in recent years. Around half of the world's population, especially in the tropics and subtropics, is at risk of infection. Every year, 50-100 million clinical cases are reported, and more than 500,000 patients develop the symptoms of severe dengue infection: dengue haemorrhagic fever and dengue shock syndrome, which threaten life in Asia and Latin America. No antiviral drug for dengue is available. The dengue virus (DENV) non-structural protein 5 (NS5), which possesses the RNA-dependent RNA polymerase (RdRp) activity and is responsible for viral replication and transcription, is an attractive target for anti-dengue drug development. In the present study, 16,240 small-molecule compounds in a fragment library were screened for their capabilities to inhibit the DENV type 2 (DENV2) RdRp activities in vitro. Based on in cellulo antiviral and cytotoxity assays, we selected the compound RK-0404678 with the EC50 value of 6.0 μM for DENV2. Crystallographic analyses revealed two unique binding sites for RK-0404678 within the RdRp, which are conserved in flavivirus NS5 proteins. No resistant viruses emerged after nine rounds of serial passage of DENV2 in the presence of RK-0404678, suggesting the high genetic barrier of this compound to the emergence of a resistant virus. Collectively, RK-0404678 and its binding sites provide a new framework for antiviral drug development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pntd.0007894DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6886872PMC
November 2019

Multiple Pathways To Avoid Beta Interferon Sensitivity of HIV-1 by Mutations in Capsid.

J Virol 2019 12 13;93(23). Epub 2019 Nov 13.

Research Institute for Microbial Diseases, Osaka University, Osaka, Japan

Type I interferons (IFNs), including alpha IFN (IFN-α) and IFN-β, potently suppress HIV-1 replication by upregulating IFN-stimulated genes (ISGs). The viral capsid protein (CA) partly determines the sensitivity of HIV-1 to IFNs. However, it remains to be determined whether CA-related functions, including utilization of known host factors, reverse transcription, and uncoating, affect the sensitivity of HIV-1 to IFN-mediated restriction. Recently, we identified an HIV-1 CA variant that is unusually sensitive to IFNs. This variant, called the RGDA/Q112D virus, contains multiple mutations in CA: H87R, A88G, P90D, P93A, and Q112D. To investigate how an IFN-hypersensitive virus can evolve to overcome IFN-β-mediated blocks targeting the viral capsid, we adapted the RGDA/Q112D virus in IFN-β-treated cells. We successfully isolated IFN-β-resistant viruses which contained either a single Q4R substitution or the double amino acid change G94D/G116R. These two IFN-β resistance mutations variably changed the sensitivity of CA binding to human myxovirus resistance B (MxB), cleavage and polyadenylation specificity factor 6 (CPSF6), and cyclophilin A (CypA), indicating that the observed loss of sensitivity was not due to interactions with these known host CA-interacting factors. In contrast, the two mutations apparently functioned through distinct mechanisms. The Q4R mutation dramatically accelerated the kinetics of reverse transcription and initiation of uncoating of the RGDA/Q112D virus in the presence or absence of IFN-β, whereas the G94D/G116R mutations affected reverse transcription only in the presence of IFN-β, most consistent with a mechanism of the disruption of binding to an unknown IFN-β-regulated host factor. These results suggest that HIV-1 can exploit multiple, known host factor-independent pathways to avoid IFN-β-mediated restriction by altering capsid sequences and subsequent biological properties. HIV-1 infection causes robust innate immune activation in virus-infected patients. This immune activation is characterized by elevated levels of type I interferons (IFNs), which can block HIV-1 replication. Recent studies suggest that the viral capsid protein (CA) is a determinant for the sensitivity of HIV-1 to IFN-mediated restriction. Specifically, it was reported that the loss of CA interactions with CPSF6 or CypA leads to higher IFN sensitivity. However, the molecular mechanism of CA adaptation to IFN sensitivity is largely unknown. Here, we experimentally evolved an IFN-β-hypersensitive CA mutant which showed decreased binding to CPSF6 and CypA in IFN-β-treated cells. The CA mutations that emerged from this adaptation indeed conferred IFN-β resistance. Our genetic assays suggest a limited contribution of known host factors to IFN-β resistance. Strikingly, one of these mutations accelerated the kinetics of reverse transcription and uncoating. Our findings suggest that HIV-1 selected multiple, known host factor-independent pathways to avoid IFN-β-mediated restriction.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.00986-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6854511PMC
December 2019

Evaluation of novel rapid detection kits for dengue virus NS1 antigen in Dhaka, Bangladesh, in 2017.

Virol J 2019 08 15;16(1):102. Epub 2019 Aug 15.

Department of Viral Infections, Research Institute for Microbial Diseases, Osaka University, 3-1, Yamada-oka, Suita, Osaka, 565-0871, Japan.

Background: Dengue virus (DENV) infection is one of the biggest challenges for human health in the world. In addition, a secondary DENV infection sometimes causes dengue hemorrhagic fever (DHF), which frequently leads to death. For this reason, accurate diagnosis record management is useful for prediction of DHF. Therefore, the demand for DENV rapid diagnosis tests (RDTs) is increasing because these tests are easy and rapid to use. However, commercially available RDTs often show low sensitivity for DENV and cross-reactivity against other flaviviruses, especially Zika virus (ZIKV).

Methods: We developed two types of novel DENV non-structural protein 1 (NS1) detection RDTs, designated TKK-1st and TKK-2nd kits. Specificities of the monoclonal antibodies (MAbs) used in these kits were confirmed by enzyme-linked immuno-sorbent assay (ELISA), dot blot, and western blot using recombinant NS1 proteins and synthetic peptides. For evaluation of sensitivity, specificity, and cross-reactivity of the novel DENV NS1 RDTs, we first used cultured DENV and other flaviviruses, ZIKV and Japanese encephalitis virus (JEV). We then used clinical specimens obtained in Bangladesh in 2017 for further evaluation of kit sensitivity and specificity in comparison with commercially available RDTs. In addition, RNA extracted from sera were used for viral genome sequencing and genotyping.

Results: Epitopes of three out of four MAbs used in the two novel RDTs were located in amino acid positions 100 to 122 in the NS1 protein, a region that shows low levels of homology with other flaviviruses. Our new kits showed high levels of sensitivity against various serotypes and genotypes of DENV and exhibited high levels of specificity without cross-reactivity against ZIKV and JEV. In clinical specimens, our RDTs showed sensitivities of 96.0% (145/151, TKK-1st kit) and 96.7% (146/151, TKK-2nd kit), and specificities of 98.0% (98/100, TKK-1st kit and TKK-2nd kit). On the other hand, in the case of the commercially available SD Bioline RDT, sensitivity was 83.4% (126/151) and specificity was 99.0% (99/100) against the same clinical specimens.

Conclusions: Our novel DENV NS1-targeting RDTs demonstrated high levels of sensitivity and lacked cross-reactivity against ZIKV and JEV compared with commercially available RDTs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s12985-019-1204-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6694664PMC
August 2019

Genotype replacement of dengue virus type 3 and clade replacement of dengue virus type 2 genotype Cosmopolitan in Dhaka, Bangladesh in 2017.

Infect Genet Evol 2019 11 24;75:103977. Epub 2019 Jul 24.

Research Institute for Microbial Diseases, Osaka University, 3-1, Yamada-oka, Suita, Osaka 565-0871, Japan; Mahidol-Osaka Center for Infectious Diseases, Mahidol University, 420/6 Ratchawithi road, Ratchathewi, Bangkok 10400, Thailand.

Dengue is a mosquito-borne disease that has spread to >100 countries and is caused by the dengue virus (DENV), which belongs to the Flavivirus genus of the family Flaviviridae. DENV comprises 4 serotypes (DENV-1 to -4), and each serotype is further divided into distinct genotypes. In India, it is reported that all 4 serotypes of DENV co-circulate. Although Bangladesh is a neighboring country of India, very few reports have published DENV sequence data for the country, especially after 2012. To understand the current distribution of DENV genotypes in Bangladesh, we determined the nucleotide sequences of envelope regions obtained from 58 DENV-positive patients diagnosed at Apollo Hospitals Dhaka during the period between September 2017 and February 2018. We found 5 DENV-1, 47 DENV-2, and 6 DENV-3 serotypes. A phylogenetic analysis of the obtained viral sequences revealed that DENV-3 genotype I was present instead of DENV-3 genotype II, which was predominant in Bangladesh between 2000 and 2009. Furthermore, we found two distinct lineages of the Cosmopolitan genotype of DENV-2, one of which was closely related to strains from Southeast Asia and has never been reported previously in Bangladesh. These results indicated that DENVs in Bangladesh have increased in genotypic diversity and suggest that the DENV genotypic shift observed in other Asian countries also might have been taking place in Bangladesh.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.meegid.2019.103977DOI Listing
November 2019

A Novel Phenotype Links HIV-1 Capsid Stability to cGAS-Mediated DNA Sensing.

J Virol 2019 08 30;93(16). Epub 2019 Jul 30.

Aaron Diamond AIDS Research Center, New York, New York, USA

The HIV-1 capsid executes essential functions that are regulated by capsid stability and host factors. In contrast to increasing knowledge on functional roles of capsid-interacting host proteins during postentry steps, less is known about capsid stability and its impact on intracellular events. Here, using the antiviral compound PF-3450074 (PF74) as a probe for capsid function, we uncovered a novel phenotype of capsid stability that has a profound effect on innate sensing of viral DNA by the DNA sensor cGAS. A single mutation, R143A, in the capsid protein conferred resistance to high concentrations of PF74, without affecting capsid binding to PF74. A cell-free assay showed that the R143A mutant partially counteracted the capsid-destabilizing activity of PF74, pointing to capsid stabilization as a resistance mechanism for the R143A mutant. In monocytic THP-1 cells, the R143A virus, but not the wild-type virus, suppressed cGAS-dependent innate immune activation. These results suggest that capsid stabilization improves the shielding of viral DNA from innate sensing. We found that a naturally occurring transmitted founder (T/F) variant shares the same properties as the R143A mutant with respect to PF74 resistance and DNA sensing. Imaging assays revealed delayed uncoating kinetics of this T/F variant and the R143A mutant. All these phenotypes of this T/F variant were controlled by a genetic polymorphism located at the trimeric interface between capsid hexamers, thus linking these capsid-dependent properties. Overall, this work functionally connects capsid stability to innate sensing of viral DNA and reveals naturally occurring phenotypic variation in HIV-1 capsid stability. The HIV-1 capsid, which is made from individual viral capsid proteins (CA), is a target for a number of antiviral compounds, including the small-molecule inhibitor PF74. In the present study, we utilized PF74 to identify a transmitted/founder (T/F) strain that shows increased capsid stability. Interestingly, PF74-resistant variants prevented cGAS-dependent innate immune activation under a condition where the other T/F strains induced type I interferon. These observations thus reveal a new CA-specific phenotype that couples capsid stability to viral DNA recognition by cytosolic DNA sensors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.00706-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6675898PMC
August 2019

HIV-1 is more dependent on the K182 capsid residue than HIV-2 for interactions with CPSF6.

Virology 2019 06 27;532:118-126. Epub 2019 Apr 27.

Research Institute for Microbial Diseases, Osaka University, Osaka, Japan.

The HIV-1 capsid (CA) utilizes CPSF6 for nuclear entry and integration site targeting. Previous studies demonstrated that the HIV-1 CA C-terminal domain (CTD) contains a highly conserved K182 residue involved in interaction with CPSF6. In contrast, certain HIV-2 strains possess a substitution at this residue (K182R). To assess whether CA-CPSF6 interaction via the CA CTD is conserved among primate lentiviruses, we examined resistance of several HIV-1- and HIV-2-lineage viruses to a truncated form of CPSF6, CPSF6-358. The results demonstrated that viruses belonging to the HIV-2-lineage maintain interaction with CPSF6 regardless of the presence of the K182R substitution, in contrast to the case with HIV-1-lineage viruses. Our structure-guided mutagenesis indicated that the differential requirement for CA-CPSF6 interaction is regulated in part by residues near the 182nd amino acid of CA. These results demonstrate a previously unrecognized distinction between HIV-1 and HIV-2, which may reflect differences in their evolutionary histories.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.virol.2019.04.012DOI Listing
June 2019

CA Mutation N57A Has Distinct Strain-Specific HIV-1 Capsid Uncoating and Infectivity Phenotypes.

J Virol 2019 05 17;93(9). Epub 2019 Apr 17.

Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA

The ability of human immunodeficiency virus type 1 (HIV-1) to transduce nondividing cells is key to infecting terminally differentiated macrophages, which can serve as a long-term reservoir of HIV-1 infection. The mutation N57A in the viral CA protein renders HIV-1 cell cycle dependent, allowing examination of HIV-1 infection of nondividing cells. Here, we show that the N57A mutation confers a postentry infectivity defect that significantly differs in magnitude between the common lab-adapted molecular clones HIV-1 (>10-fold) and HIV-1 (2- to 5-fold) in multiple human cell lines and primary CD4 T cells. Capsid permeabilization and reverse transcription are altered when N57A is incorporated into HIV-1 but not HIV-1 The N57A infectivity defect is significantly exacerbated in both virus strains in the presence of cyclosporine (CsA), indicating that N57A infectivity is dependent upon CA interacting with host factor cyclophilin A (CypA). Adaptation of N57A HIV-1 selected for a second CA mutation, G94D, which rescued the N57A infectivity defect in HIV-1 but not HIV-1 The rescue of N57A by G94D in HIV-1 is abrogated by CsA treatment in some cell types, demonstrating that this rescue is CypA dependent. An examination of over 40,000 HIV-1 CA sequences revealed that the four amino acids that differ between HIV-1 and HIV-1 CA are polymorphic, and the residues at these positions in the two strains are widely prevalent in clinical isolates. Overall, a few polymorphic amino acid differences between two closely related HIV-1 molecular clones affect the phenotype of capsid mutants in different cell types. The specific mechanisms by which HIV-1 infects nondividing cells are unclear. A mutation in the HIV-1 capsid protein abolishes the ability of the virus to infect nondividing cells, serving as a tool to examine cell cycle dependence of HIV-1 infection. We have shown that two widely used HIV-1 molecular clones exhibit significantly different N57A infectivity phenotypes due to fewer than a handful of CA amino acid differences and that these clones are both represented in HIV-infected individuals. As such minor differences in closely related HIV-1 strains may impart significant infectivity differences, careful consideration should be given to drawing conclusions from one particular HIV-1 clone. This study highlights the potential for significant variation in results with the use of multiple strains and possible unanticipated effects of natural polymorphisms.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.00214-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6475785PMC
May 2019

Naturally Occurring Mutations in HIV-1 CRF01_AE Capsid Affect Viral Sensitivity to Restriction Factors.

AIDS Res Hum Retroviruses 2018 04 13;34(4):382-392. Epub 2018 Feb 13.

3 National Institute of Health , Department of Medical Science, Ministry of Public Health, Nonthaburi, Thailand .

TRIM5α and MxB are known as restriction factors that inhibit the early step of intracellular HIV-1 replication cycle. Both factors are believed to interact with the incoming virus core to suppress HIV-1 infection. The extreme diversity of HIV-1 is thought to be a consequence of its propensity to mutate to escape immune responses and host restriction factors. We recently determined the capsid sequences for 144 HIV-1 CRF01_AE viruses obtained in Thailand from 2005 to 2011. In this study, we further analyzed the amino acid variations among the capsid sequences of 204 HIV-1 CRF01_AE obtained in Thailand and China, including 84 of the aforementioned 144 viruses, to detect mutations permitting escape from restriction by host factors. We found a characteristic combination of E79D, V83T, and H87Q in sequences from Chinese viruses and subsequently showed that this combination conferred partial resistance to MxB. Interestingly, this combination conferred resistance to human TRIM5α as well. The H87Q mutation alone conferred resistance to MxB in the CRF01_AE background, but not in subtype B virus. In contrast, the H87Q mutation alone conferred resistance to human TRIM5α in both the CFR01_AE and subtype B backgrounds. BLAST analysis revealed the presence of the E79D, V83T, and H87Q combination in CRF01_AE viruses isolated not only in China but also in many other countries. Although the mechanistic details as well as precise role of MxB antiviral activity in infected individuals remain to be clarified, our data suggest an interaction between MxB and the HIV-1 capsid in vivo.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1089/AID.2017.0212DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5899301PMC
April 2018

Human T-cell leukemia virus type 1 infects multiple lineage hematopoietic cells in vivo.

PLoS Pathog 2017 Nov 29;13(11):e1006722. Epub 2017 Nov 29.

Laboratory of Virus Control, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan.

Human T-cell leukemia virus type 1 (HTLV-1) infects mainly CD4+CCR4+ effector/memory T cells in vivo. However, it remains unknown whether HTLV-1 preferentially infects these T cells or this virus converts infected precursor cells to specialized T cells. Expression of viral genes in vivo is critical to study viral replication and proliferation of infected cells. Therefore, we first analyzed viral gene expression in non-human primates naturally infected with simian T-cell leukemia virus type 1 (STLV-1), whose virological attributes closely resemble those of HTLV-1. Although the tax transcript was detected only in certain tissues, Tax expression was much higher in the bone marrow, indicating the possibility of de novo infection. Furthermore, Tax expression of non-T cells was suspected in bone marrow. These data suggest that HTLV-1 infects hematopoietic cells in the bone marrow. To explore the possibility that HTLV-1 infects hematopoietic stem cells (HSCs), we analyzed integration sites of HTLV-1 provirus in various lineages of hematopoietic cells in patients with HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP) and a HTLV-1 carrier using the high-throughput sequencing method. Identical integration sites were detected in neutrophils, monocytes, B cells, CD8+ T cells and CD4+ T cells, indicating that HTLV-1 infects HSCs in vivo. We also detected Tax protein in myeloperoxidase positive neutrophils. Furthermore, dendritic cells differentiated from HTLV-1 infected monocytes caused de novo infection to T cells, indicating that infected monocytes are implicated in viral spreading in vivo. Certain integration sites were re-detected in neutrophils from HAM/TSP patients at different time points, indicating that infected HSCs persist and differentiate in vivo. This study demonstrates that HTLV-1 infects HSCs, and infected stem cells differentiate into diverse cell lineages. These data indicate that infection of HSCs can contribute to the persistence and spread of HTLV-1 in vivo.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.ppat.1006722DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5724899PMC
November 2017

Sequence diversity of dengue virus type 2 in brain and thymus of infected interferon receptor ko mice: implications for dengue virulence.

Virol J 2016 Nov 30;13(1):199. Epub 2016 Nov 30.

Department of Virology I, National Institute for Infectious Diseases, Tokyo, Japan.

Background: We previously reported that a clinical isolate of dengue virus (DENV) is capable of causing acute-phase systemic infection in mice harboring knockouts of the genes encoding type-I and -II interferon IFN receptors (IFN-α/β/γR KO mice); in contrast, other virulent DENV isolates exhibited slow disease progression in this mice, yielding lethal infection around 20 days post-infection (p.i.). In the present study, we sought to clarify the dynamics of slow disease progression by examining disease progression of a type-2 DENV clinical isolate (DV2P04/08) in mice.

Methods: The tissue distributions of DV2P04/08 in several organs of infeted mice were examined at different time points. Whole genome viral sequences from organs were determined.

Results: At day 6 p.i., high levels of viral RNA (vRNA) were detected in non-neuronal organs (including peritoneal exudate cells (PECs), spleen, kidney, liver, lung, and bone marrow) but not in brain. By day 14 p.i, vRNA levels subsequently decreased in most organs, with the exception of thymus and brain. Sequence analysis of the whole genome of the original P04/08 and those of viruses recovered from mouse brain and thymus demonstrated the presence of both synonymous and non-synonymous mutations. Individual mice showed different virus populations in the brain. The vRNA sequence derived from brain of one mouse was nearly identical to the original DV2P04/08 inoculum, suggesting that there was no need for adaptation of DV2P04/08 for growth in the brain. However, quasispecies (that is, mixed populations, detected as apparent nucleotide mixtures during sequencing) were observed in the thymus of another mouse, and interestingly only mutant population invaded the brain at a late stage of infection.

Conclusions: These results suggested that the mouse nearly succeeded in eliminating virus from non-neuronal organs but failed to do so from brain. Although the cause of death by DV2P04/08 infection is likely to be the result of virus invasion to brain, its processes to the death are different in individual mice. This study will provide a new insight into disease progression of DENV in mice.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s12985-016-0658-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5129197PMC
November 2016

Epidemiological Surveillance of Lymphocryptovirus Infection in Wild Bonobos.

Front Microbiol 2016 12;7:1262. Epub 2016 Aug 12.

Primate Research Institute, Kyoto UniversityKyoto, Japan; Institute for Virus Research, Kyoto UniversityKyoto, Japan.

Lymphocryptovirus (LCV) is one of the major gena in the herpesvirus family and is widely disseminated among primates. LCVs of human and rhesus macaques are shown to be causative agents of a number of malignant diseases including lymphoma and carcinoma. Bonobos (Pan paniscus) are highly endangered and the least studied species of the great apes. Considering the potential pathogenicity of the LCV that might threaten the fate of wild bonobos, population-based epidemiological information in terms of LCV prevalence in different location of Bonobo's habitats will help propose improved conservation strategies for the bonobos. However, such data are not available yet because it is very difficult to collect blood samples in the wild and thus virtually impossible to conduct sero-epidemiological study on the wild ape. In order to overcome this issue, we focused on evaluating anti-LCV IgA in the feces of bonobos, which are available in a non-invasive manner. Preliminary study showed that anti-LCV IgA but not IgG was efficiently and reproducibly detected in the feces of captive chimpanzees. It is noteworthy that the fecal IgA-positive individuals were seropositive for both anti-LCV IgG and IgA and that the IgA antibodies in both sera and feces were also detectable by Western blotting assay. These results indicate that the detection of fecal anti-LCV IgA is likely a reliable and feasible for epidemiological surveillance of LCV prevalence in the great apes. We then applied this method and found that 31% of wild bonobos tested were positive for anti-LCV IgA antibody in the feces. Notably, the positivity rates varied extensively among their sampled populations. In conclusion, our results in this study demonstrate that LCV is highly disseminated among wild bonobos while the prevalence is remarkably diverse in their population-dependent manner.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fmicb.2016.01262DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4981747PMC
August 2016

Capsid-CPSF6 Interaction Is Dispensable for HIV-1 Replication in Primary Cells but Is Selected during Virus Passage In Vivo.

J Virol 2016 08 11;90(15):6918-6935. Epub 2016 Jul 11.

Aaron Diamond AIDS Research Center, New York, New York, USA

Unlabelled: Cleavage and polyadenylation specificity factor subunit 6 (CPSF6), a host factor that interacts with the HIV-1 capsid (CA) protein, is implicated in diverse functions during the early part of the HIV-1 life cycle, including uncoating, nuclear entry, and integration targeting. Preservation of CA binding to CPSF6 in vivo suggests that this interaction is fine-tuned for efficient HIV-1 replication in physiologically relevant settings. Nevertheless, this possibility has not been formally examined. To assess the requirement for optimal CPSF6-CA binding during infection of primary cells and in vivo, we utilized a novel CA mutation, A77V, that significantly reduced CA binding to CPSF6. The A77V mutation rendered HIV-1 largely independent from TNPO3, NUP358, and NUP153 for infection and altered the integration site preference of HIV-1 without any discernible effects during the late steps of the virus life cycle. Surprisingly, the A77V mutant virus maintained the ability to replicate in monocyte-derived macrophages, primary CD4(+) T cells, and humanized mice at a level comparable to that for the wild-type (WT) virus. Nonetheless, revertant viruses that restored the WT CA sequence and hence CA binding to CPSF6 emerged in three out of four A77V-infected animals. These results suggest that the optimal interaction of CA with CPSF6, though not absolutely essential for HIV-1 replication in physiologically relevant settings, confers a significant fitness advantage to the virus and thus is strictly conserved among naturally circulating HIV-1 strains.

Importance: CPSF6 interacts with the HIV-1 capsid (CA) protein and has been implicated in nuclear entry and integration targeting. Preservation of CPSF6-CA binding across various HIV-1 strains suggested that the optimal interaction between CA and CPSF6 is critical during HIV-1 replication in vivo Here, we identified a novel HIV-1 capsid mutant that reduces binding to CPSF6, is largely independent from the known cofactors for nuclear entry, and alters integration site preference. Despite these changes, virus carrying this mutation replicated in humanized mice at levels indistinguishable from those of the wild-type virus. However, in the majority of the animals, the mutant virus reverted back to the wild-type sequence, hence restoring the wild-type level of CA-CPSF6 interactions. These results suggest that optimal binding of CA to CPSF6 is not absolutely essential for HIV-1 replication in vivo but provides a fitness advantage that leads to the widespread usage of CPSF6 by HIV-1 in vivo.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.00019-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4944271PMC
August 2016

Roles of Capsid-Interacting Host Factors in Multimodal Inhibition of HIV-1 by PF74.

J Virol 2016 06 27;90(12):5808-5823. Epub 2016 May 27.

Aaron Diamond AIDS Research Center, New York, New York, USA

Unlabelled: The viral capsid of HIV-1 interacts with a number of host factors to orchestrate uncoating and regulate downstream events, such as reverse transcription, nuclear entry, and integration site targeting. PF-3450074 (PF74), an HIV-1 capsid-targeting low-molecular-weight antiviral compound, directly binds to the capsid (CA) protein at a site also utilized by host cell proteins CPSF6 and NUP153. Here, we found that the dose-response curve of PF74 is triphasic, consisting of a plateau and two inhibitory phases of different slope values, consistent with a bimodal mechanism of drug action. High PF74 concentrations yielded a steep curve with the highest slope value among different classes of known antiretrovirals, suggesting a dose-dependent, cooperative mechanism of action. CA interactions with both CPSF6 and cyclophilin A (CypA) were essential for the unique dose-response curve. A shift of the steep curve at lower drug concentrations upon blocking the CA-CypA interaction suggests a protective role for CypA against high concentrations of PF74. These findings, highlighting the unique characteristics of PF74, provide a model in which its multimodal mechanism of action of both noncooperative and cooperative inhibition by PF74 is regulated by interactions of cellular proteins with incoming viral capsids.

Importance: PF74, a novel capsid-targeting antiviral against HIV-1, shares its binding site in the viral capsid protein (CA) with the host factors CPSF6 and NUP153. This work reveals that the dose-response curve of PF74 consists of two distinct inhibitory phases that are differentially regulated by CA-interacting host proteins. PF74's potency depended on these CA-binding factors at low doses. In contrast, the antiviral activity of high PF74 concentrations was attenuated by cyclophilin A. These observations provide novel insights into both the mechanism of action of PF74 and the roles of host factors during the early steps of HIV-1 infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.03116-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4886767PMC
June 2016

Novel mutant human immunodeficiency virus type 1 strains with high degree of resistance to cynomolgus macaque TRIMCyp generated by random mutagenesis.

J Gen Virol 2016 Apr 20;97(4):963-976. Epub 2016 Jan 20.

Department of Viral Infections, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, Japan.

Old World monkey TRIM5α strongly suppresses human immunodeficiency virus type 1 (HIV-1) replication. A fusion protein comprising cynomolgus macaque (CM) TRIM5 and cyclophilin A (CM TRIMCyp) also potently suppresses HIV-1 replication. However, CM TRIMCyp fails to suppress a mutant HIV-1 that encodes a mutant capsid protein containing a SIVmac239-derived loop between α-helices 4 and 5 (L4/5). There are seven amino acid differences between L4/5 of HIV-1 and SIVmac239. Here, we investigated the minimum numbers of amino acid substitutions that would allow HIV-1 to evade CM TRIMCyp-mediated suppression. We performed random PCR mutagenesis to construct a library of HIV-1 variants containing mutations in L4/5, and then we recovered replication-competent viruses from CD4+ MT4 cells that expressed high levels of CM TRIMCyp. CM TRIMCyp-resistant viruses were obtained after three rounds of selection in MT4 cells expressing CM TRIMCyp and these were found to contain four amino acid substitutions (H87R, A88G, P90D and P93A) in L4/5. We then confirmed that these substitutions were sufficient to confer CM TRIMCyp resistance to HIV-1. In a separate experiment using a similar method, we obtained novel CM TRIM5α-resistant HIV-1 strains after six rounds of selection and rescue. Analysis of these mutants revealed that V86A and G116E mutations in the capsid region conferred partial resistance to CM TRIM5α without substantial fitness cost when propagated in MT4 cells expressing CM TRIM5α. These results confirmed and further extended the previous notion that CM TRIMCyp and CM TRIM5α recognize the HIV-1 capsid in different manners.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1099/jgv.0.000408DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4854368PMC
April 2016

Emergence of infectious malignant thrombocytopenia in Japanese macaques (Macaca fuscata) by SRV-4 after transmission to a novel host.

Sci Rep 2015 Mar 6;5:8850. Epub 2015 Mar 6.

Department of Molecular Biology, Primate Research Institute, Kyoto University, Inuyama, Aichi 484-8506, Japan.

We discovered a lethal hemorrhagic syndrome arising from severe thrombocytopenia in Japanese macaques kept at the Primate Research Institute, Kyoto University. Extensive investigation identified that simian retrovirus type 4 (SRV-4) was the causative agent of the disease. SRV-4 had previously been isolated only from cynomolgus macaques in which it is usually asymptomatic. We consider that the SRV-4 crossed the so-called species barrier between cynomolgus and Japanese macaques, leading to extremely severe acute symptoms in the latter. Infectious agents that cross the species barrier occasionally amplify in virulence, which is not observed in the original hosts. In such cases, the new hosts are usually distantly related to the original hosts. However, Japanese macaques are closely related to cynomolgus macaques, and can even hybridize when given the opportunity. This lethal outbreak of a novel pathogen in Japanese macaques highlights the need to modify our expectations about virulence with regards crossing species barriers.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/srep08850DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4351523PMC
March 2015

Seroprevalence of Japanese encephalitis virus infection in captive Japanese macaques (Macaca fuscata).

Primates 2014 Jul 19;55(3):441-5. Epub 2014 Apr 19.

Laboratory of Veterinary Microbiology, Joint Faculty of Veterinary Medicine, Yamaguchi University, 1677-1 Yoshida, Yamaguchi, Yamaguchi, 753-8515, Japan.

Japanese encephalitis virus (JEV), which is transmitted by mosquitoes, infects many animal species and causes serious acute encephalitis in humans and horses. In this study, a serosurvey of JEV in Japanese macaques (Macaca fuscata) reared in Aichi Prefecture was conducted using purified JEV as an antigen for ELISA. The results revealed that 146 of 332 monkeys (44 %) were seropositive for JEV. In addition, 35 of 131 monkeys (27 %) born in the facility were seropositive, and the annual infection rate in the facility was estimated as 13 %. Our results provide evidence of the frequent exposure of many Japanese macaques to JEV, suggesting that there is a risk of JEV transmission to humans by mosquitoes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10329-014-0421-7DOI Listing
July 2014

Characterization of simian T-cell leukemia virus type 1 in naturally infected Japanese macaques as a model of HTLV-1 infection.

Retrovirology 2013 Oct 24;10:118. Epub 2013 Oct 24.

Laboratory of Virus Control, Institute for Virus Research, Kyoto University, Shogoin Kawahara-cho 53, Sakyo-ku, Kyoto 606-8507, Japan.

Background: Human T-cell leukemia virus type 1 (HTLV-1) causes chronic infection leading to development of adult T-cell leukemia (ATL) and inflammatory diseases. Non-human primates infected with simian T-cell leukemia virus type 1 (STLV-1) are considered to constitute a suitable animal model for HTLV-1 research. However, the function of the regulatory and accessory genes of STLV-1 has not been analyzed in detail. In this study, STLV-1 in naturally infected Japanese macaques was analyzed.

Results: We identified spliced transcripts of STLV-1 corresponding to HTLV-1 tax and HTLV-1 bZIP factor (HBZ). STLV-1 Tax activated the NFAT, AP-1 and NF-κB signaling pathways, whereas STLV-1 bZIP factor (SBZ) suppressed them. Conversely, SBZ enhanced TGF-β signaling and induced Foxp3 expression. Furthermore, STLV-1 Tax activated the canonical Wnt pathway while SBZ suppressed it. STLV-1 Tax enhanced the viral promoter activity while SBZ suppressed its activation. Then we addressed the clonal proliferation of STLV-1⁺ cells by massively sequencing the provirus integration sites. Some clones proliferated distinctively in monkeys with higher STLV-1 proviral loads. Notably, one of the monkeys surveyed in this study developed T-cell lymphoma in the brain; STLV-1 provirus was integrated in the lymphoma cell genome. When anti-CCR4 antibody, mogamulizumab, was administered into STLV-1-infected monkeys, the proviral load decreased dramatically within 2 weeks. We observed that some abundant clones recovered after discontinuation of mogamulizumab administration.

Conclusions: STLV-1 Tax and SBZ have functions similar to those of their counterparts in HTLV-1. This study demonstrates that Japanese macaques naturally infected with STLV-1 resemble HTLV-1 carriers and are a suitable model for the investigation of persistent HTLV-1 infection and asymptomatic HTLV-1 carrier state. Using these animals, we verified that mogamulizumab, which is currently used as a drug for relapsed ATL, is also effective in reducing the proviral load in asymptomatic individuals.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/1742-4690-10-118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4016002PMC
October 2013

Efficient in vivo depletion of CD8(+) T lymphocytes in common marmosets by novel CD8 monoclonal antibody administration.

Immunol Lett 2013 Jul-Aug;154(1-2):12-7. Epub 2013 Aug 19.

Tsukuba Primate Research Center, National Institute of Biomedical Innovation, Hachimandai, Tsukuba, Ibaraki 305-0843, Japan; Primate Research Institute, Kyoto University, Inuyama, Aichi 484-8506, Japan. Electronic address:

In order to directly demonstrate the roles of CD8(+) T lymphocytes in non-human primates, in vivo depletion of the CD8(+) T cells by administration of a CD8-specific monoclonal antibody (mAb) is one of the crucial techniques. Recently, the common marmoset (Callithrix jacchus), which is classified as a New World monkey, has been shown useful as an experimental animal model for various human diseases such as multiple sclerosis, Parkinson's disease and a number of infectious diseases. Here we show that an anti-marmoset CD8 mAb 6F10, which we have recently established, efficiently depletes the marmoset CD8(+) T lymphocytes in vivo, i.e., the administration of 6F10 induces drastic and specific reduction in the ratio of the CD8(+) T cell subset for at least three weeks or longer. Our finding will help understand the pivotal role of CD8(+) T cells in vivo in the control of human diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.imlet.2013.08.005DOI Listing
July 2014

Generation of rhesus macaque-tropic HIV-1 clones that are resistant to major anti-HIV-1 restriction factors.

J Virol 2013 Nov 21;87(21):11447-61. Epub 2013 Aug 21.

Department of Microbiology, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima, Tokushima, Japan.

Human immunodeficiency virus type 1 (HIV-1) replication in macaque cells is restricted mainly by antiviral cellular APOBEC3, TRIM5α/TRIM5CypA, and tetherin proteins. For basic and clinical HIV-1/AIDS studies, efforts to construct macaque-tropic HIV-1 (HIV-1mt) have been made by us and others. Although rhesus macaques are commonly and successfully used as infection models, no HIV-1 derivatives suitable for in vivo rhesus research are available to date. In this study, to obtain novel HIV-1mt clones that are resistant to major restriction factors, we altered Gag and Vpu of our best HIV-1mt clone described previously. First, by sequence- and structure-guided mutagenesis, three amino acid residues in Gag-capsid (CA) (M94L/R98S/G114Q) were found to be responsible for viral growth enhancement in a macaque cell line. Results of in vitro TRIM5α susceptibility testing of HIV-1mt carrying these substitutions correlated well with the increased viral replication potential in macaque peripheral blood mononuclear cells (PBMCs) with different TRIM5 alleles, suggesting that the three amino acids in HIV-1mt CA are involved in the interaction with TRIM5α. Second, we replaced the transmembrane domain of Vpu of this clone with the corresponding region of simian immunodeficiency virus SIVgsn166 Vpu. The resultant clone, MN4/LSDQgtu, was able to antagonize macaque but not human tetherin, and its Vpu effectively functioned during viral replication in a macaque cell line. Notably, MN4/LSDQgtu grew comparably to SIVmac239 and much better than any of our other HIV-1mt clones in rhesus macaque PBMCs. In sum, MN4/LSDQgtu is the first HIV-1 derivative that exhibits resistance to the major restriction factors in rhesus macaque cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.01549-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3807366PMC
November 2013

Macaque-tropic human immunodeficiency virus type 1: breaking out of the host restriction factors.

Front Microbiol 2013 9;4:187. Epub 2013 Jul 9.

Center for Human Evolution Modeling Research, Primate Research Institute, Kyoto University Inuyama, Japan ; Japan Foundation for AIDS Prevention Chiyoda-ku, Japan.

Macaque monkeys serve as important animal models for understanding the pathogenesis of lentiviral infections. Since human immunodeficiency virus type 1 (HIV-1) hardly replicates in macaque cells, simian immunodeficiency virus (SIV) or chimeric viruses between HIV-1 and SIV (SHIV) have been used as challenge viruses in this research field. These viruses, however, are genetically distant from HIV-1. Therefore, in order to evaluate the efficacy of anti-HIV-1 drugs and vaccines in macaques, the development of a macaque-tropic HIV-1 (HIV-1mt) having the ability to replicate efficiently in macaques has long been desired. Recent studies have demonstrated that host restriction factors, such as APOBEC3 family and TRIM5, impose a strong barrier against HIV-1 replication in macaque cells. By evading these restriction factors, others and we have succeeded in developing an HIV-1mt that is able to replicate in macaques. In this review, we have attempted to shed light on the role of host factors that affect the susceptibility of macaques to HIV-1mt infection, especially by focusing on TRIM5-related factors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fmicb.2013.00187DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3705164PMC
July 2013

TRIM5 genotypes in cynomolgus monkeys primarily influence inter-individual diversity in susceptibility to monkey-tropic human immunodeficiency virus type 1.

J Gen Virol 2013 Jun 13;94(Pt 6):1318-1324. Epub 2013 Mar 13.

Center for Human Evolution Modeling Research, Primate Research Institute, Kyoto University, 41-2 Kanrin, Inuyama, Aichi 484-8506, Japan.

TRIM5α restricts human immunodeficiency virus type 1 (HIV-1) infection in cynomolgus monkey (CM) cells. We previously reported that a TRIMCyp allele expressing TRIM5-cyclophilin A fusion protein was frequently found in CMs. Here, we examined the influence of TRIM5 gene variation on the susceptibility of CMs to a monkey-tropic HIV-1 derivative (HIV-1mt) and found that TRIMCyp homozygotes were highly susceptible to HIV-1mt not only in vitro but also in vivo. These results provide important insights into the inter-individual differences in susceptibility of macaques to HIV-1mt.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1099/vir.0.050252-0DOI Listing
June 2013

Systemic biological analysis of the mutations in two distinct HIV-1mt genomes occurred during replication in macaque cells.

Microbes Infect 2013 Apr 4;15(4):319-28. Epub 2013 Feb 4.

Department of Microbiology, Institute of Health Biosciences, The University of Tokushima Graduate School, 3-18-15 Kuramoto, Tokushima 770-8503, Japan.

Fundamental property of viruses is to rapidly adapt themselves under changing conditions of virus replication. Using HIV-1 derivatives that poorly replicate in macaque cells as model viruses, we studied here mechanisms for promoting viral replication in non-natural host cells. We found that the HIV-1s could evolve to grow better in both macaque and human cells by the continuous culture in macaque lymphocyte cell lines. Notably, only several mutations at defined sites of the Pol-integrase and/or the Env-gp120 reproducibly appeared in repeated adaptation experiments and were sufficient to cause the phenotypic change. Meanwhile, no amino acid changes to enhance viral replication in macaque cells were found in interaction sites for the known anti-retroviral proteins. These findings disclose a hitherto unappreciated evolutionary pathway to augment HIV-1 replication in primate cells, where tuning of viral interactions with positive rather than negative factors for replication can play a dominant role.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.micinf.2013.01.005DOI Listing
April 2013

Dynamics of cellular immune responses in the acute phase of dengue virus infection.

Arch Virol 2013 Jun 5;158(6):1209-20. Epub 2013 Feb 5.

Tsukuba Primate Research Center, National Institute of Biomedical Innovation, 1-1 Hachimandai, Tsukuba, Ibaraki 305-0843, Japan.

In this study, we examined the dynamics of cellular immune responses in the acute phase of dengue virus (DENV) infection in a marmoset model. Here, we found that DENV infection in marmosets greatly induced responses of CD4/CD8 central memory T and NKT cells. Interestingly, the strength of the immune response was greater in animals infected with a dengue fever strain than in those infected with a dengue hemorrhagic fever strain of DENV. In contrast, when animals were re-challenged with the same DENV strain used for primary infection, the neutralizing antibody induced appeared to play a critical role in sterilizing inhibition against viral replication, resulting in strong but delayed responses of CD4/CD8 central memory T and NKT cells. The results in this study may help to better understand the dynamics of cellular and humoral immune responses in the control of DENV infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00705-013-1618-6DOI Listing
June 2013