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HTLV-1 bZIP factor induces T-cell lymphoma and systemic inflammation in vivo.

Satou Y, Yasunaga J, Zhao T, Yoshida M, Miyazato P, Takai K, Shimizu K, Ohshima K, Green PL, Ohkura N, Yamaguchi T, Ono M, Sakaguchi S, Matsuoka M - PLoS Pathog. (2011)

Bottom Line: As a mechanism of increased T(reg) cells, HBZ expression directly induced Foxp3 gene transcription in T cells.HBZ could physically interact with Foxp3 and NFAT, thereby impairing the suppressive function of T(reg) cells.Thus, the expression of HBZ in CD4+ T cells is a key mechanism of HTLV-1-induced neoplastic and inflammatory diseases.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Virus Control, Institute for Virus Research, Kyoto University, Kyoto, Japan.

ABSTRACT
Human T-cell leukemia virus type 1 (HTLV-1) is the causal agent of a neoplastic disease of CD4+ T cells, adult T-cell leukemia (ATL), and inflammatory diseases including HTLV-1 associated myelopathy/tropical spastic paraparesis, dermatitis, and inflammatory lung diseases. ATL cells, which constitutively express CD25, resemble CD25+CD4+ regulatory T cells (T(reg)). Approximately 60% of ATL cases indeed harbor leukemic cells that express FoxP3, a key transcription factor for T(reg) cells. HTLV-1 encodes an antisense transcript, HTLV-1 bZIP factor (HBZ), which is expressed in all ATL cases. In this study, we show that transgenic expression of HBZ in CD4+ T cells induced T-cell lymphomas and systemic inflammation in mice, resembling diseases observed in HTLV-1 infected individuals. In HBZ-transgenic mice, CD4+Foxp3+ T(reg) cells and effector/memory CD4+ T cells increased in vivo. As a mechanism of increased T(reg) cells, HBZ expression directly induced Foxp3 gene transcription in T cells. The increased CD4+Foxp3+ T(reg) cells in HBZ transgenic mice were functionally impaired while their proliferation was enhanced. HBZ could physically interact with Foxp3 and NFAT, thereby impairing the suppressive function of T(reg) cells. Thus, the expression of HBZ in CD4+ T cells is a key mechanism of HTLV-1-induced neoplastic and inflammatory diseases.

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HBZ inhibites Foxp3-mediated CTLA-4 and GITR expression in vitro.(A) Mouse CD4+CD25− T cells co-transduced with the retroviral vectors were stained with the indicated antibodies. Mean fluorescence intensity (MFI) of CD25, GITR, and CTLA-4 in GFP/NGFR double-positive cells are shown as mean ± SD. for triplicate culture. *, P<0.01; **, P<0.001 by two-tailed Student t-test. (B and C) CD4+CD25− T cells transduced with the pMXs-Ig vector encoding wild-type or mutant HBZ, and pGCSamIN-Foxp3 vector were stained with anti-GITR (B) or anti-CTLA-4 (C) antibody in addition to anti-NGFR antibody, and then analyzed by flow cytometry. Left, numbers in density plots indicate MFI of GITR (B) or CTLA-4 (C) in GFP/NGFR double-positive cells. Representative data from three independent experiments are shown. Right, relative MFI of wild type or mutated HBZ compared to mock transduced cells was shown as mean ± SD (n = 3). (D) HBZ transduction in Foxp3+ Treg cells inhibited the endogenous expression of Treg associated molecules. Mean fluorescence intensity (MFI) of CD25, GITR, and CTLA-4 in CD4+Foxp3+NGFR+ cells are shown as mean ± SD. for triplicate culture. *, P<0.05; **, P<0.01 by two-tailed Student t-test.
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ppat-1001274-g006: HBZ inhibites Foxp3-mediated CTLA-4 and GITR expression in vitro.(A) Mouse CD4+CD25− T cells co-transduced with the retroviral vectors were stained with the indicated antibodies. Mean fluorescence intensity (MFI) of CD25, GITR, and CTLA-4 in GFP/NGFR double-positive cells are shown as mean ± SD. for triplicate culture. *, P<0.01; **, P<0.001 by two-tailed Student t-test. (B and C) CD4+CD25− T cells transduced with the pMXs-Ig vector encoding wild-type or mutant HBZ, and pGCSamIN-Foxp3 vector were stained with anti-GITR (B) or anti-CTLA-4 (C) antibody in addition to anti-NGFR antibody, and then analyzed by flow cytometry. Left, numbers in density plots indicate MFI of GITR (B) or CTLA-4 (C) in GFP/NGFR double-positive cells. Representative data from three independent experiments are shown. Right, relative MFI of wild type or mutated HBZ compared to mock transduced cells was shown as mean ± SD (n = 3). (D) HBZ transduction in Foxp3+ Treg cells inhibited the endogenous expression of Treg associated molecules. Mean fluorescence intensity (MFI) of CD25, GITR, and CTLA-4 in CD4+Foxp3+NGFR+ cells are shown as mean ± SD. for triplicate culture. *, P<0.05; **, P<0.01 by two-tailed Student t-test.

Mentions: In HBZ-Tg mice, the expression of Treg-associated molecules including CTLA-4, GITR and CD25 was suppressed when compared with their expression in Treg cells from non-Tg mice (Figure 3B and E). This finding may account for the impaired function of Treg cells since these molecules, in particular CTLA-4, play a critical role in Treg-mediated suppression [39]. To further study the effect of HBZ on the expression of Treg-associated molecules, we transduced HBZ along with Foxp3 into naive CD4+ T cells in vitro using retrovirus vectors (Figure 4A). HBZ expression suppressed Foxp3-induced GITR and CTLA-4 expression whereas it did not inhibit CD25 expression (Figure 6A). Expression of HBZ alone increased CD25 expression (Figure 6A), which might obscure the suppressive effect of HBZ under these conditions. Suppression of GITR and CTLA-4 expression required both the activation and the central domains of HBZ (Figure 6, B and C), which correspond to the binding sites of HBZ to Foxp3 (Figure 5C) and NFAT (Figure S12). Since both Foxp3 and NFAT are critical for Treg function [35], it is likely that HBZ suppresses the expression of GITR and CTLA-4 by interacting with Foxp3 and NFAT and thereby interfering with their transcriptional regulation in Treg cells. To examine suppressive effect of HBZ on expression of GITR, CTLA-4 and CD25, we isolated Treg cells from wild type mice and expressed HBZ using retroviral vectors. As shown in Figure 6D, HBZ suppressed endogenous expression of CD25, GITR and CTLA-4 in Treg cells, confirming that HBZ is responsible for suppressed expression of these molecules.


HTLV-1 bZIP factor induces T-cell lymphoma and systemic inflammation in vivo.

Satou Y, Yasunaga J, Zhao T, Yoshida M, Miyazato P, Takai K, Shimizu K, Ohshima K, Green PL, Ohkura N, Yamaguchi T, Ono M, Sakaguchi S, Matsuoka M - PLoS Pathog. (2011)

HBZ inhibites Foxp3-mediated CTLA-4 and GITR expression in vitro.(A) Mouse CD4+CD25− T cells co-transduced with the retroviral vectors were stained with the indicated antibodies. Mean fluorescence intensity (MFI) of CD25, GITR, and CTLA-4 in GFP/NGFR double-positive cells are shown as mean ± SD. for triplicate culture. *, P<0.01; **, P<0.001 by two-tailed Student t-test. (B and C) CD4+CD25− T cells transduced with the pMXs-Ig vector encoding wild-type or mutant HBZ, and pGCSamIN-Foxp3 vector were stained with anti-GITR (B) or anti-CTLA-4 (C) antibody in addition to anti-NGFR antibody, and then analyzed by flow cytometry. Left, numbers in density plots indicate MFI of GITR (B) or CTLA-4 (C) in GFP/NGFR double-positive cells. Representative data from three independent experiments are shown. Right, relative MFI of wild type or mutated HBZ compared to mock transduced cells was shown as mean ± SD (n = 3). (D) HBZ transduction in Foxp3+ Treg cells inhibited the endogenous expression of Treg associated molecules. Mean fluorescence intensity (MFI) of CD25, GITR, and CTLA-4 in CD4+Foxp3+NGFR+ cells are shown as mean ± SD. for triplicate culture. *, P<0.05; **, P<0.01 by two-tailed Student t-test.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3037353&req=5

ppat-1001274-g006: HBZ inhibites Foxp3-mediated CTLA-4 and GITR expression in vitro.(A) Mouse CD4+CD25− T cells co-transduced with the retroviral vectors were stained with the indicated antibodies. Mean fluorescence intensity (MFI) of CD25, GITR, and CTLA-4 in GFP/NGFR double-positive cells are shown as mean ± SD. for triplicate culture. *, P<0.01; **, P<0.001 by two-tailed Student t-test. (B and C) CD4+CD25− T cells transduced with the pMXs-Ig vector encoding wild-type or mutant HBZ, and pGCSamIN-Foxp3 vector were stained with anti-GITR (B) or anti-CTLA-4 (C) antibody in addition to anti-NGFR antibody, and then analyzed by flow cytometry. Left, numbers in density plots indicate MFI of GITR (B) or CTLA-4 (C) in GFP/NGFR double-positive cells. Representative data from three independent experiments are shown. Right, relative MFI of wild type or mutated HBZ compared to mock transduced cells was shown as mean ± SD (n = 3). (D) HBZ transduction in Foxp3+ Treg cells inhibited the endogenous expression of Treg associated molecules. Mean fluorescence intensity (MFI) of CD25, GITR, and CTLA-4 in CD4+Foxp3+NGFR+ cells are shown as mean ± SD. for triplicate culture. *, P<0.05; **, P<0.01 by two-tailed Student t-test.
Mentions: In HBZ-Tg mice, the expression of Treg-associated molecules including CTLA-4, GITR and CD25 was suppressed when compared with their expression in Treg cells from non-Tg mice (Figure 3B and E). This finding may account for the impaired function of Treg cells since these molecules, in particular CTLA-4, play a critical role in Treg-mediated suppression [39]. To further study the effect of HBZ on the expression of Treg-associated molecules, we transduced HBZ along with Foxp3 into naive CD4+ T cells in vitro using retrovirus vectors (Figure 4A). HBZ expression suppressed Foxp3-induced GITR and CTLA-4 expression whereas it did not inhibit CD25 expression (Figure 6A). Expression of HBZ alone increased CD25 expression (Figure 6A), which might obscure the suppressive effect of HBZ under these conditions. Suppression of GITR and CTLA-4 expression required both the activation and the central domains of HBZ (Figure 6, B and C), which correspond to the binding sites of HBZ to Foxp3 (Figure 5C) and NFAT (Figure S12). Since both Foxp3 and NFAT are critical for Treg function [35], it is likely that HBZ suppresses the expression of GITR and CTLA-4 by interacting with Foxp3 and NFAT and thereby interfering with their transcriptional regulation in Treg cells. To examine suppressive effect of HBZ on expression of GITR, CTLA-4 and CD25, we isolated Treg cells from wild type mice and expressed HBZ using retroviral vectors. As shown in Figure 6D, HBZ suppressed endogenous expression of CD25, GITR and CTLA-4 in Treg cells, confirming that HBZ is responsible for suppressed expression of these molecules.

Bottom Line: As a mechanism of increased T(reg) cells, HBZ expression directly induced Foxp3 gene transcription in T cells.HBZ could physically interact with Foxp3 and NFAT, thereby impairing the suppressive function of T(reg) cells.Thus, the expression of HBZ in CD4+ T cells is a key mechanism of HTLV-1-induced neoplastic and inflammatory diseases.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Virus Control, Institute for Virus Research, Kyoto University, Kyoto, Japan.

ABSTRACT
Human T-cell leukemia virus type 1 (HTLV-1) is the causal agent of a neoplastic disease of CD4+ T cells, adult T-cell leukemia (ATL), and inflammatory diseases including HTLV-1 associated myelopathy/tropical spastic paraparesis, dermatitis, and inflammatory lung diseases. ATL cells, which constitutively express CD25, resemble CD25+CD4+ regulatory T cells (T(reg)). Approximately 60% of ATL cases indeed harbor leukemic cells that express FoxP3, a key transcription factor for T(reg) cells. HTLV-1 encodes an antisense transcript, HTLV-1 bZIP factor (HBZ), which is expressed in all ATL cases. In this study, we show that transgenic expression of HBZ in CD4+ T cells induced T-cell lymphomas and systemic inflammation in mice, resembling diseases observed in HTLV-1 infected individuals. In HBZ-transgenic mice, CD4+Foxp3+ T(reg) cells and effector/memory CD4+ T cells increased in vivo. As a mechanism of increased T(reg) cells, HBZ expression directly induced Foxp3 gene transcription in T cells. The increased CD4+Foxp3+ T(reg) cells in HBZ transgenic mice were functionally impaired while their proliferation was enhanced. HBZ could physically interact with Foxp3 and NFAT, thereby impairing the suppressive function of T(reg) cells. Thus, the expression of HBZ in CD4+ T cells is a key mechanism of HTLV-1-induced neoplastic and inflammatory diseases.

Show MeSH
Related in: MedlinePlus