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Biomedical subjects

Tatsuo Shioda

Publications and source records attributed to Tatsuo Shioda.

At least 19 recordsLinked to original sources
PubMed2006

Development of a rapid and convenient method for the quantification of HIV-1 budding.

In cells, the expression of Gag protein, one of the major structural proteins of retroviruses, is sufficient for budding virus-like particles (VLPs) from the cell surface. We have previously reported that spheroplasts of Saccharomyces cerevisiae expressing HIV-1 Gag proteins from an episomal plasmid constitutively released a large amount of VLPs into culture media; however, commercially available ELISA kits which detect mature capsid of HIV-1 could not detect uncleaved 55-kDa Gag proteins released from budding yeast. We therefore developed a method to quantitate VLP levels released from budding yeast by using fusion protein from HIV-1 Gag and Firefly Luciferase. This system is useful for screening cellular factor(s) involved in retrovirus budding from S. cerevisiae.

Gene Products, gag↗
PubMed2006

A dominant-negative effect of cynomolgus monkey tripartite motif protein TRIM5alpha on anti-simian immunodeficiency virus SIVmac activity of an African green monkey orthologue.

African green monkey (AGM) tripartite motif protein (TRIM) 5alpha can inhibit both human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus SIVmac, whereas cynomolgus monkey (CM) TRIM5alpha can inhibit HIV-1, but not SIVmac. We previously reported that the 17-amino-acid region and an adjacent 20-amino-acid duplication in the SPRY(B30.2) domain of AGM TRIM5alpha determined the species specificity. In the present study, we demonstrated that CM TRIM5alpha had a dominant-negative effect on the anti-SIVmac activity of AGM TRIM5alpha. In contrast, mutant TRIM5alphas lacking the 20-amino-acid duplication did not have the dominant-negative effect, even though they failed to restrict SIVmac. These results indicated that oligomerization of the SPRY domain is required for anti-SIVmac activity and suggest that tight interaction between the viral capsid and all three molecules in one TRIM5alpha trimer may not be necessary for restriction activity.

Amino Acid Motifs↗
PubMed2006

Effects of human interleukin 7 on HIV-1 replication in monocyte-derived human macrophages.

Interleukin 7 (IL-7) contributes to development and proliferation of T cells. We investigated the effect of IL-7 on HIV-1 infected monocyte-derived human macrophages. IL-7 treatment of macrophages at a concentration of 10 ng/ml reduced replication of the R5 HIV-1 strain by approximately 50%. Meanwhile, HIV-1-infected macrophages themselves could excrete approximately 20% more IL-7 than uninfected macrophages. These results suggest that IL-7 could be used as a therapeutic modality to recover CD4 T cells.

Cell Differentiation↗
PubMed2006

Protective effects of IL4-589T and RANTES-28G on HIV-1 disease progression in infected Thai females.

OBJECTIVE: To evaluate the effect of polymorphisms in interleukin-4 (IL4) and RANTES promoters on disease progression in HIV-1 infected Thais. DESIGN: Antiretroviral (ARV) drug-free HIV-1 infected females from the prospective cohort. METHODS: A total of 246 DNA samples were genotyped for IL4 and RANTES promoter polymorphisms by PCR-RFLP. Associations of genotype with HIV-1 disease progression were assessed with respect to baseline clinical data including plasma HIV-1 load, CD4 cell counts, and proportion of symptomatic/AIDS, and survival status during 3 years of follow-up. RESULTS: Patients with homozygous IL4-589T allele showed a significantly lower HIV-1 viral load (P = 0.005) and a higher CD4 cell count (P = 0.003) than the other patients with heterozygous IL4-589C/T or homozygous IL4-589C allele. Kaplan-Meier analysis demonstrated an apparent but insignificant trend towards better survival in homozygous IL4-589T patients. On the other hand, patients with RANTES-28G allele showed a significantly better survival while those with RANTES In1.1C allele without RANTES-28G showed a significantly poorer survival compared with those who did not possess either RANTES In1.1C or RANTES-28G (P = 0.02), although those polymorphisms only weakly associated with baseline viral load and CD4 cell counts. CONCLUSIONS: Our results implicate the significant protective effect of IL4-589T and RANTES-28G on HIV disease progression in Thais. In contrast, RANTES In1.1C without RANTES-28G had an accelerating effect on HIV disease progression.

CD4 Lymphocyte Count↗
PubMed2006

Human genetic polymorphisms affecting HIV-1 diseases.

Human immunodeficiency virus type 1 (HIV-1) infection is generally characterized by a long-term, chronic disease course that gradually progresses to acquired immunodeficiency syndrome (AIDS). However, a small fraction of HIV-1-infected individuals remain both clinically and immunologically healthy for 10 years or more after seroconversion. Conversely, the disease of another significant fraction is characterized by an extremely rapid progression to AIDS within 1 year. There are also individuals not infected with HIV-1 who have had repetitive sexual exposure to HIV-1 in extremely high-risk situations. Determining the host factors involved in these different susceptibility and disease courses would be helpful for better understanding AIDS and its control. Today, a worldwide scientific endeavor called the Human Genome Project has completed the production of a full-length sequence of the 3 billion base pairs that make up the human genome. The Human Genome Project has also revealed the presence of variation in human genomes. Relating these genetic differences to human phenotypes offers a very attractive prospect for a genetic understanding of the different sensitivities to various human diseases, including cancer, brain and heart diseases, allergy, and infectious diseases. In this review, we present examples of human genetic variation that can affect HIV-1 infection and disease progression.

Acquired Immunodeficiency Syndrome↗
PubMed2005

A specific region of 37 amino acid residues in the SPRY (B30.2) domain of African green monkey TRIM5alpha determines species-specific restriction of simian immunodeficiency virus SIVmac infection.

Human immunodeficiency virus type 1 (HIV-1) efficiently enters cells of Old World monkeys but encounters a block before reverse transcription. This restriction is mediated by a dominant repressive factor. Recently, a member of the tripartite motif (TRIM) family proteins, TRIM5alpha, was identified as a blocking factor in a rhesus macaque cDNA library. Among Old World monkey cell lines, the African green monkey kidney cell line CV1 is highly resistant to not only HIV-1 but also simian immunodeficiency virus SIVmac infection. We analyzed TRIM5alpha of CV1 cells and HSC-F cells, a T-cell line from a cynomolgus monkey, and found that both CV1- and HSC-F-TRIM5alphas could inhibit CD4-dependent HIV-1 infection, as well as vesicular stomatitis virus glycoprotein-mediated infection. CV1-TRIM5alpha could also inhibit SIVmac infection, whereas HSC-F-TRIM5alpha could not. In the SPRY (B30.2) domain of CV1-TRIM5alpha, there was a 20-amino-acid duplication that was not present in HSC-F-TRIM5alpha. A chimeric TRIM5alpha containing 37 amino acid residues from CV1-TRIM5alpha, which spanned the 20-amino-acid duplication, in the background of HSC-F-TRIM5alpha fully gained the ability to inhibit SIVmac infection. Conversely, the mutant CV1-TRIM5alpha lacking the 20-amino-acid duplication completely lost the ability to restrict SIVmac infection. These findings clearly indicated that a specific region of 37 amino acid residues in the SPRY domain of CV1-TRIM5alpha contained a determinant of species-specific restriction of SIVmac.

Amino Acid Sequence↗
PubMed2005

Influence of glycosylation on the efficacy of an Env-based vaccine against simian immunodeficiency virus SIVmac239 in a macaque AIDS model.

The envelope glycoprotein (Env) of human immunodeficiency viruses (HIVs) and simian immunodeficiency viruses (SIVs) is heavily glycosylated, and this feature has been speculated to be a reason for the insufficient immune control of these viruses by their hosts. In a macaque AIDS model, we demonstrated that quintuple deglycosylation in Env altered a pathogenic virus, SIVmac239, into a novel attenuated mutant virus (delta5G). In delta5G-infected animals, strong protective immunity against SIVmac239 was elicited. These HIV and SIV studies suggested that an understanding of the role of glycosylation is critical in defining not only the virological properties but also the immunogenicity of Env, suggesting that glycosylation in Env could be modified for the development of effective vaccines. To examine the effect of deglycosylation, we constructed prime-boost vaccines consisting of Env from SIVmac239 and delta5G and compared their immunogenicities and vaccine efficacies by challenge infection with SIVmac239. Vaccination-induced immune responses differed between the two vaccine groups. Both Env-specific cellular and humoral responses were higher in wild-type (wt)-Env-immunized animals than in delta5G Env-immunized animals. Following the challenge, viral loads in SIVmac239 Env (wt-Env)-immunized animals were significantly lower than in vector controls, with controlled viral replication in the chronic phase. Unexpectedly, viral loads in delta5G Env-immunized animals were indistinguishable from those in vector controls. This study demonstrated that the prime-boost Env vaccine was effective against homologous SIVmac239 challenge. Changes in glycosylation affected both cell-mediated and humoral immune responses and vaccine efficacy.

Animals↗
PubMed2005

[TRIM5alpha].

Human immunodeficiency virus type 1 (HIV-1) shows a very narrow host range limited only to humans and chimpanzees. HIV-1 dose not experimentally infect Old World monkeys, such as rhesus and cynomolgus monkeys, and fails to replicate in activated CD4 positive T lymphocytes obtained from those monkeys. Several lines of evidence have suggested that the block of HIV-1 replication in Old World monkey cells occurred at a post-entry step and appeared to result from a failure to initiate reverse transcription. Recently, the screening of a rhesus monkey cDNA library identified tripartite motif 5 (TRIM5) alpha, a component of cytoplasmic bodies, as a factor that confers resistance to HIV-1 infection. Shortly after, TRIM5alpha of African green monkey, another Old World monkey, was also shown to restrict HIV-1 infection, while human TRIM5alpha was reported to restrict N-tropic murine leukemia virus. Small amino acid differences in the SPRY domain among human and monkey TRIM5alphas were reported to determine species-specific restriction. This review discusses about anti-viral activity of TRIM5alpha.

Animals↗
PubMed2004

A CCR2-V64I polymorphism affects stability of CCR2A isoform.

OBJECTIVE: A valine to isoleucine substitution at position 64 of CCR2 (CCR2-64I) is associated with a delay in progression to AIDS in HIV-1-infected individuals. The aim of the present study is to elucidate the molecular mechanism underlying the effect of this allele. DESIGN: We analysed the effect of the 64I substitution on levels of expression of CCR2A and CCR2B, two CCR2 isoforms produced by alternative splicing. METHODS: Sendai virus vector was used to express CCR2 molecules. RESULTS: While CCR2B trafficked well to the cell surface, CCR2A, which differs from CCR2B only by the sequence of its C-terminal cytoplasmic tail, was detected predominantly in the cytoplasm. The level of expression of CCR2A-64I was significantly higher than that of CCR2A without the substitution. On the other hand, the 64I substitution did not affect levels of CCR2B expression. Pulse-chase experiments revealed that the 64I substitution increased the half-life of CCR2A in cells. When co-expressed with CCR5, CCR2A-64I interfered more severely with cell surface expression of CCR5 than did wild-type CCR2A. Furthermore, immunoprecipitation experiments showed that CCR2A co-precipitated with an immature form of CCR5. CONCLUSION: These results suggest that CCR2A binds to CCR5 in the cytoplasm and down-modulates its surface expression. We propose that the increased ability of CCR2A-64I to down-modulate CCR5 expression might be a possible cause of a delay in HIV-1 disease progression in patients with this allele.

Amino Acid Sequence↗
PubMed2003

Possible role of dimerization in human immunodeficiency virus type 1 genome RNA packaging.

The dimer initiation site/dimer linkage sequence (DIS/DLS) region in the human immunodeficiency virus type 1 (HIV-1) RNA genome is suggested to play important roles in various steps of the virus life cycle. However, due to the presence of a putative DIS/DLS region located within the encapsidation signal region (E/psi), it is difficult to perform a mutational analysis of DIS/DLS without affecting the packaging of RNA into virions. Recently, we demonstrated that duplication of the DIS/DLS region in viral RNA caused the production of partially monomeric RNAs in virions, indicating that the region indeed mediated RNA-RNA interaction. We utilized this system to assess the precise location of DIS/DLS in the 5' region of the HIV-1 genome with minimum effect on RNA packaging. We found that the entire lower stem of the U5/L stem-loop was required for packaging, whereas the region important for dimer formation was only 10 bases long within the lower stem of the U5/L stem-loop. The R/U5 stem-loop was required for RNA packaging but was completely dispensable for dimer formation. The SL1 lower stem was important for both dimerization and packaging, but surprisingly, deletion of the palindromic sequence at the top of the loop only partially affected dimerization. These results clearly indicated that the E/psi of HIV-1 is much larger than the DIS/DLS and that the primary DIS/DLS is completely included in the E/psi. Therefore, it is suggested that RNA dimerization is a part of RNA packaging, which requires multiple steps.

Base Sequence↗
PubMed2002

New Rev-transport inhibitor with anti-HIV activity from Valerianae Radix.

Bioassay-guided separation by use of the fission yeast expressing NES of Rev, a HIV-1 viral regulatory protein, resulted in isolation of valtrate (1) as a new Rev-transport inhibitor from the nucleus to cytoplasm from Valerianae Radix. Valtrate (1) also inhibited the p-24 production of HIV-1 virus without showing any cytotoxicity against the host MT-4 cells.

Biotin↗
PubMed2002

Protective effect of interleukin-4 -589T polymorphism on human immunodeficiency virus type 1 disease progression: relationship with virus load.

The interleukin (IL)-4 -589T allele bears a single nucleotide polymorphism at position -589 upstream from the open-reading frame of the IL-4 gene. To determine the influence of this allele on human immunodeficiency virus (HIV) type 1 disease, disease progression and serum virus load were assessed by IL-4 genotype in 427 white patients with known seroconversion dates who were followed in the French SEROCO cohort between 1988 and 1996. Serum virus load was 0.20 log lower during the 6-24-month plateau phase after seroconversion in patients with IL-4 -589T than in those without this allele (P=.02). Kaplan-Meier analysis survival curves showed a slower progression to clinical AIDS in carriers of IL-4 -589T (P=.04). Adjustment for early serum virus load greatly diminished the strength of this association. These results suggest that IL-4 -589T protects against HIV-1 disease progression by reducing virus load.

Acquired Immunodeficiency Syndrome↗
PubMed2002

Maximal HIV-1 replication in alveolar macrophages during tuberculosis requires both lymphocyte contact and cytokines.

HIV-1 replication is markedly upregulated in alveolar macrophages (AM) during pulmonary tuberculosis (TB). This is associated with loss of an inhibitory CCAAT enhancer binding protein beta (C/EBPbeta) transcription factor and activation of nuclear factor (NF)-kappaB. Since the cellular immune response in pulmonary TB requires lymphocyte--macrophage interaction, a model system was developed in which lymphocytes were added to AM. Contact between lymphocytes and AM reduced inhibitory C/EBPbeta, activated NF-kappaB, and enhanced HIV-1 replication. If contact between lymphocytes and macrophages was prevented, inhibitory C/EBPbeta expression was maintained and the HIV-1 long terminal repeat (LTR) was not maximally stimulated although NF-kappaB was activated. Antibodies that cross-linked macrophage expressed B-7, and vascular cell adhesion molecule and CD40 were used to mimic lymphocyte contact. All three cross-linking antibodies were required to abolish inhibitory C/EBPbeta expression. However, the HIV-1 LTR was not maximally stimulated and NF-kappaB was not activated. Maximal HIV-1--LTR stimulation required both lymphocyte-derived soluble factors, and cross-linking of macrophage expressed costimulatory molecules. High level HIV-1--LTR stimulation was also achieved when IL-1beta, IL-6, and TNF-beta were added to macrophages with cross-linked costimulatory molecules. Contact between activated lymphocytes and macrophages is necessary to down-regulate inhibitory C/EBPbeta, thereby derepressing the HIV-1 LTR. Lymphocyte-derived cytokines activate NF-kappaB, further enhancing the HIV-1 LTR.

Blotting, Western↗
PubMed2002

An efficient and versatile mammalian viral vector system for major histocompatibility complex class I/peptide complexes.

We report a Sendai virus (SeV) vector system for expression of major histocompatibility complex (MHC) class I/peptide complexes. We cloned the extracellular domain of a human MHC class I heavy chain, HLA-A*2402, and human beta-2 microglobulin (beta2m) fused with HLA-A*2402-restricted human immunodeficiency virus type 1 (HIV-1) cytotoxic T-lymphocyte (CTL) epitopes (e-beta2m) in separate SeV vectors. When we coinfected nonhuman mammalian cells with the SeVs, naturally folded human MHC class I/peptide complexes were secreted in the culture supernatants. Biotin binding peptide sequences on the C terminus of the heavy chain were used to tetramerize the complexes. These tetramers made in the SeV system recognized specific CD8-positive T cells in peripheral blood mononuclear cells of HIV-1-positive patients with a specificity and sensitivity similar to those of MHC class I tetramers made in an Escherichia coli system. Solo infection of e-beta2m/SeV produced soluble e-beta2m in the culture supernatant, and cells pulsed with the soluble protein were recognized by specific CTLs. Furthermore, when cells were infected with e-beta2m/SeV, these cells were recognized by the specific CTLs more efficiently than the protein pulse per se. SeV is nonpathogenic for humans, can transduce foreign genes into nondividing cells, and may be useful for immunotherapy to enhance antigen-specific immune responses. Our system can be used not only to detect but also to stimulate antigen-specific cellular immune responses.

Animals↗
PubMed2002

Dissociation of genome dimerization from packaging functions and virion maturation of human immunodeficiency virus type 1.

The dimer initiation site/dimer linkage sequence (DIS/DLS) region of the human immunodeficiency virus type 1 (HIV-1) RNA genome is thought to play important roles at various stages of the virus life cycle. Recently we showed that the DIS/DLS region affects RNA-RNA interaction in intact virus particles, by demonstrating that duplication of the region in viral RNA caused the production of virus particles containing partially monomeric RNAs. We have extended this finding and succeeded for the first time in creating mutant particles which contain only monomeric RNAs without modifying any viral proteins. In terms of RNA encapsidation ability, virion density, and protein processing, the mutant particles were comparable to wild-type particles. The level of production of viral DNA by the mutant virus construct in infected cells was also comparable to that of the constructs that produced exclusively dimeric RNA, indicating that monomeric viral RNA could be the template for strand transfer. These results indicated that the RNA dimerization of HIV-1 could be separated from viral RNA packaging and was not absolutely required for RNA packaging, virion maturation, and reverse transcription.

Cell Line, Transformed↗