Sequences flanking the cAMP responsive core of the HTLV-I tax response elements influence CREB protease sensitivity.
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The human T cell leukemia virus type I (HTLV-I) Tax protein activates transcription from the viral long terminal repeat and select cellular promoters by interacting with cellular DNA-binding proteins. The HTLV-I promoter contains three copies of a Tax-responsive element (TRE-1), each of which possesses a core cAMP response element (CRE). The cAMP response element-binding protein, CREB, binds TRE-1 and mediates Tax association with, and transactivation of, the viral promoter. These activities depend on DNA sequences that flank the core CRE. Although CREs are found in a variety of cellular promoters, cellular CREs vary in sequence from TRE-1, especially in the flanking regions, and are generally not Tax responsive. The molecular basis for differential Tax responsiveness of viral and cellular CREs has not been determined. Here we demonstrate that the conformation of CREB is influenced by the nucleotide sequence of its DNA-binding element. CREB showed altered sensitivity to V8, chymotrypsin, and trypsin proteases when bound to the HTLV-I TRE-1 element as compared to the rat somatostatin CRE element. The phosphorylation state of CREB did not influence its protease sensitivity on either element. Sequences flanking the core CRE-binding site in each element were found to specify protease sensitivity. Since the TRE-1-flanking sequences also modulate Tax association with CREB, and Tax transactivation of CREB-dependent LTR transcription, these results suggest that CREB conformation may determine the ability of Tax to bind CREB. (+info)
An altered peptide ligand antagonizes antigen-specific T cells of patients with human T lymphotropic virus type I-associated neurological disease.
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Human T lymphotropic virus type I (HTLV-I)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is an inflammatory neurologic disease associated with HTLV-I infection, in which chronically activated, HTLV-I-specific CD8+ CTL have been suggested to be immunopathogenic. In HLA-A2 HAM/TSP patients, CD8+ HTLV-I-specific CTLs recognize an immunodominant peptide of the HTLV-I Tax protein, Tax11-19. We examined the functional outcome on activation of both cloned peripheral blood and cerebrospinal spinal fluid-derived CTL and bulk PBMC from HAM/TSP patients by altered peptide ligands (APL) derived from HTLV-I Tax11-19. In CTL clones generated from PBMC and CSF of HLA-A2 HAM/TSP patients, an APL substituted at position 5 significantly decreased CTL responses when compared with the native peptide. Moreover, these ligands were also shown to inhibit CTL responses to the native peptide in bulk PBMC of HLA-A2 HAM/TSP patients. These data suggest that a modification of an antigenic peptide at the central position can manipulate the T cell responses in bulk PBMC from different individuals with an inflammatory disease. Additionally, these results have implications for the potential use of APL-based immunotherapy in this T cell-mediated CNS disease. (+info)
Adult T cell leukemia/lymphoma with lymphopenia in a Korean.
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We experienced a case of adult T cell leukemia/lymphoma (ATLL) in a 48-year-old Korean female, who has never been abroad since birth and no history of blood transfusion. The patient had hypercalcemia and multiple lymphadenopathy. Histopathologic study of left cervical lymph node (LN) and bone marrow (BM) revealed that infiltrates of malignant lymphoid cells were composed of small, medium and large cells with pleomorphic nuclei. Smears of peripheral blood (PB) showed lymphopenia (16%) with the appearance of a few atypical lymphoid cells (less than 2%), but not the typical clover leaf cells seen in ATLL. Immunophenotypic study of LN and BM revealed T cell phenotype. PB showed increased CD4+ T cell (T(H), CD3/CD4+, 57%) and decreased CD8+ T cell counts (T(S), CD3/CD8+, 6.7%). The sera of the patient and her family were reactive for HTLV-I antibody. The specific sequences of pol, env, and tax of HTLV-I DNA were detected in the lymphoma cells and peripheral blood mononuclear cells (PBMC) using polymerase chain reaction. Ultrastructural examination of PBMC confirmed numerous type c virus particles in extracellular space. This case was an acute type of ATLL without overt leukemic features in PB. Despite chemotherapy and intensive conservative treatment, she died 3 months after admission. (+info)
Isolation, cloning, and complete nucleotide sequence of a phenotypically distinct Brazilian isolate of human T-lymphotropic virus type II (HTLV-II).
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Analysis of human T-lymphotropic virus type II (HTLV-II) isolates from North America and Europe have demonstrated the existence of two molecular subtypes of the virus, HTLV-IIa and HTLV-IIb. Recently, studies on HTLV-II infections in Brazil have revealed isolates that are related phylogenetically to the HTLV-IIa subtype but have a HTLV-IIb phenotype with respect to the transactivating protein, tax. To more clearly define this relationship, HTLV-II was isolated from peripheral blood of an IVDA from Sao Paulo, Brazil (SP-WV), and the complete provirus was cloned and sequenced. Comparison of HTLV-II(SP-WV) nucleotide sequences to other available complete HTLV-II proviral sequences revealed that HTLV-II(SP-WV) is most closely related to HTLV-II(Mo), the prototypic HTLV-IIa subtype sequence. Phylogenetic analysis of LTR, env, and tax regions unequivocally demonstrated that HTLV-II(SP-WV) and all other Brazilian sequences examined are members of the IIa subtype. The predicted amino acid sequences of the major coding regions of HTLV-II(SP-WV) are also most closely related to HTLV-II(Mo), with the important exception of tax. The tax protein encoded by HTLV-II(SP-WV) is 96-99% identical to the tax of IIb isolates and is similar in that it has an additional 25 amino acids at the carboxy-terminus compared to the HTLV-II(Mo) tax with which it shares 91% identity. Analysis of tax stop codon usage of a number of HTLV-IIa isolates from North American, Europe, and Brazil demonstrated that isolates from the last region appear to be unique in their extended tax phenotype. It could be demonstrated that the extended tax proteins in the HTLV-IIb and Brazilian isolates had equivalent ability to transactivate the viral LTR, and studies with deletion mutants indicated that the extended C-terminus is not essential for transactivation. In contrast, the HTLV-IIa tax was found to have a greatly diminished ability to transactivate the viral LTR, which appeared to be a consequence of reduced expression of the protein. The studies show that although the Brazilian strains do not represent an entirely new subtype based on nucleotide sequence analysis they are a phenotypically unique molecular variant within the HTLV-IIa subtype. (+info)
HTLV-I Tax transrepresses the human c-Myb promoter independently of its interaction with CBP or p300.
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The c-Myb proto-oncogene is preferentially expressed in hematopoietic lineages, and highly expressed in several leukemia types. The Human T-cell Leukemia Virus Type I (HTLV-I) is the etiological agent of Adult T-cell Leukemia/Lymphoma (ATLL). A previous report suggested that Tax, the viral transactivator, is able to suppress the transactivation potential of c-Myb protein by competing for recruitment of CBP. We tested whether such a competition could affect transcription from the c-Myb promoter in Tax expressing T-cells. Using several c-Myb promoter reporter constructs carrying mutations in various regions, we demonstrate that Tax suppression of c-Myb transactivation results in transrepression of the c-Myb promoter through the Myb responsive elements in Jurkat T-cells. The ability of Tax mutants M22, M47 and V89A to interact with the full-length CBP and p300 proteins in vitro, and their ability to repress the c-Myb promoter, was then evaluated. Although both M47 and M22 bind to CBP and p300 to a similar extent, only M47 was able to repress the c-Myb promoter, suggesting that competition for CBP/p300 binding was not the mechanism underlying Tax's effect. This concept was further supported by the fact that the Tax mutant V89A transrepresses the c-Myb promoter efficiently in spite of an impaired binding to CBP and p300. Therefore, Tax-mediated repression of the c-Myb promoter appears to be independent from a direct competition between c-Myb and Tax for recruitment of CBP/p300. Interestingly, a decreased transcription from the endogenous c-Myb promoter was observed in several HTLV-I transformed T-cell lines. Finally, the ability of Tax to directly repress the endogenous c-Myb promoter was demonstrated in a Jurkat cell line stably transfected with a tax gene driven by a cadmium-inducible promoter. (+info)
HTLV-1 Tax protein sensitizes cells to apoptotic cell death induced by DNA damaging agents.
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Transient HTLV-1 Tax expression suppresses cellular nucleotide excision repair, and this effect correlates with Tax transactivation of the proliferating cell nuclear antigen promoter. The inability to repair DNA damage typically induces apoptotic cell death. Therefore, we investigated the effect of Tax-mediated suppression of DNA repair on apoptosis in stable Tax-expressing cells. Constitutive Tax expression reduced cellular nucleotide excision repair activity compared with parental and control cells. Tax-expressing cells were also more sensitive to apoptosis induced by DNA damaging agents than control cells. Even though Tax-expressing cells displayed reduced DNA repair, they showed increased DNA replication following UV damage. These results suggest that Tax suppresses the cell's ability to repair DNA damage and stimulates DNA replication even in the presence of damage. The inability to repair DNA damage is likely to stimulate apoptotic cell death in the majority of Tax-expressing cells while the ability to promote DNA replication may also allow the survival of a small population of cells. We propose that together these effects contribute to the monoclonal nature and low efficiency of HTLV-1 transformation. (+info)
Prostaglandin E(2) increases bovine leukemia virus tax and pol mRNA levels via cyclooxygenase 2: regulation by interleukin-2, interleukin-10, and bovine leukemia virus.
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Prostaglandin E(2) (PGE(2)), produced by macrophages, has important immune regulatory functions, suppressing a type 1 immune response and stimulating a type 2 immune response. Type 1 cytokines (interleukin-2 [IL-2], IL-12, and gamma interferon) increase in freshly isolated peripheral blood mononuclear cells (PBMCs) of animals with an early disease stage of bovine leukemia virus (BLV) infection, while IL-10 increases in animals with a late disease stage. Although IL-10 has an immunosuppressive role in the host immune system, IL-10 also inhibits BLV tax and pol mRNA levels in vitro. In contrast, IL-2 stimulates BLV tax and pol mRNA and p24 protein expression in cultured PBMCs. The inhibitory effect of IL-10 on BLV expression depends on soluble factors secreted by macrophages. Thus, we hypothesized that PGE(2), a cyclooxygenase 2 (COX-2) product of macrophages, may regulate BLV expression. Here, we show that the level of COX-2 mRNA was decreased in PBMCs treated with IL-10, while IL-2 enhanced the level of COX-2 mRNA. Addition of PGE(2) stimulated BLV tax and pol mRNA levels and reversed the IL-10 inhibition of BLV mRNA. In addition, the specific COX-2 inhibitor, NS-398, inhibited the amount of BLV mRNA detected. Addition of PGE(2) increased BLV tax mRNA regardless of NS-398 addition. PGE(2) inhibited antigen-specific PBMC stimulation, suggesting that stimulation of BLV tax and pol mRNA levels by PGE(2) is independent of cell proliferation. These findings suggest that macrophage-derived COX-2 products, such as PGE(2), regulate virus expression and disease progression in BLV infection. (+info)
Transcriptional activation of JC virus by human T-lymphotropic virus type I Tax protein in human neuronal cell lines.
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Polyomavirus JC (JCV) causes the human demyelinating disease, progressive multifocal leukoencephalopathy (PML). The recent demonstration of cases of PML in association with human T-lymphotropic virus type I (HTLV-I) infection prompted us to examine whether the HTLV-I-encoded regulatory protein Tax activates JCV transcription. By employing a dual luciferase assay, we initially found that the expression of Tax activated the transcriptional potential of both early and late promoters of JCV in human neuronal but not in non-neuronal cells. We subsequently analyzed the mechanism of Tax-induced activation of the JCV promoter in neuronal cells with the following results: 1) the JCV promoter that lacks the NF-kappaB-binding motif could not be activated by Tax; 2) the overexpression of IkappaBalpha abolished Tax-induced transcriptional activation of the JCV promoter; 3) a Tax mutant (M22) lacking the potential for activation via the NF-kappaB pathway did not activate the JCV promoter. Furthermore, Tax enhances the gene expression of JCV T antigen and VP1. We examined mechanisms of the cell-specific activation of the JCV promoter by Tax. Electrophoretic mobility shift assay demonstrated the presence of Tax-bound protein(s) that were specifically present in non-neuronal cells. This study is the first demonstration of the activation of JCV promoter by HTLV-I Tax in an NF-kappaB-dependent manner. (+info)