HTLV-1 Tax Oncoprotein Research Articles

Gene-to-gene coordinated regulation of transcription and alternative splicing by 3D chromatin remodeling upon NF-κB activation.

The NF-κB protein p65/RelA plays a pivotal role in coordinating gene expression in response to diverse stimuli, including viral infections. At the chromatin level, p65/RelA regulates gene transcription and alternative splicing through promoter enrichment and genomic exon occupancy, respectively. The intricate ways in which p65/RelA simultaneously governs these functions across various genes remain to be fully elucidated. In this study, we employed the HTLV-1 Tax oncoprotein, a potent activator of NF-κB, to investigate its influence on the three-dimensional organization of the genome, a key factor in gene regulation. We discovered that Tax restructures the 3D genomic landscape, bringing together genes based on their regulation and splicing patterns. Notably, we found that the Tax-induced gene-gene contact between the two master genes NFKBIA and RELA is associated with their respective changes in gene expression and alternative splicing. Through dCas9-mediated approaches, we demonstrated that NFKBIA-RELA interaction is required for alternative splicing regulation and is caused by an intragenic enrichment of p65/RelA on RELA. Our findings shed light on new regulatory mechanisms upon HTLV-1 Tax and underscore the integral role of p65/RelA in coordinated regulation of NF-κB-responsive genes at both transcriptional and splicing levels in the context of the 3D genome.

Retraction: 'Dose-dependent dual effect of HTLV-1 tax oncoprotein on p53-dependent nucleotide excision repair in human T-cells' by Yana Schavinsky-Khrapunsky, Esther Priel and Mordechai Aboud.

Schavinsky-Khrapunsky, Y., Priel, E. and Aboud, M. (2008), Dose-dependent dual effect of HTLV-1 tax oncoprotein on p53-dependent nucleotide excision repair in human T-cells. Int. J. Cancer, 122: 305-316. doi:10.1002/ijc.23091.

HTLV-1 Tax Oncoprotein Inhibits the Estrogen-Induced-ER α-Mediated BRCA1 Expression by Interaction with CBP/p300 Cofactors

BRCA1 is a multifunctional tumor suppressor, whose expression is activated by the estrogen (E2)-liganded ERα receptor and regulated by certain recruited transcriptional co-activators. Interference with BRCA1 expression and/or functions leads to high risk of breast or/and ovarian cancer. Another multifunctional protein, HTLV-1Tax oncoprotein, is widely regarded as crucial for developing adult T-cell leukemia and other clinical disorders. Tax profile reveals that it can antagonize BRCA1 expression and/or functionality. Therefore, we hypothesize that Tax expression in breast cells can sensitize them to malignant transformation by environmental carcinogens. Here we examined Tax effect on BRCA1 expression by testing its influence on E2-induced expression of BRCA1 promoter-driven luciferase reporter (BRCA1-Luc). We found that E2 strongly stimulated this reporter expression by liganding to ERα, which consequently associated with BRCA1 promoter, while ERα concomitantly recruited CBP/p300 to this complex for co-operative enhancement of BRCA1 expression. Introducing Tax into these cells strongly blocked this E2-ERα-mediated activation of BRCA1 expression. We noted, also, that Tax exerted this inhibition by binding to CBP/p300 without releasing them from their complex with ERα. Chip assay revealed that the binding of Tax to the CBP/p300-ERα complex, prevented its link to AP1 site. Interestingly, we noted that elevating the intracellular pool of CBP or p300 to excessive levels dramatically reduced the Tax-mediated inhibition of BRCA1 expression. Exploring the mechanism of this reduction revealed that the excessive co-factors were sufficient to bind separately the free Tax molecules, thus lowering their amount in the CBP/p300-ERα complex and relieving, thereby, the inhibition of BRCA1 expression.

Antagonistic effects of HTLV-1 Tax oncoprotein on BRCA1 expression and function

HTLV-1 Tax oncoprotein is considered a key factor in HTLV-1 pathogenicity. BRCA1 gene dysfunction can lead to breast cancer development. In contrast to the tumor suppressor nature of BRCA1, Tax is a potent oncoprotein, most of its activities are strictly opposing those of BRCA1. Therefore, we hypothesize that HTLV1 Tax expression in breast epithelial cells can antagonize BRCA1 expression and functionality, thereby sensitizing these cells to malignant transformation by environmental carcinogens. Our objective was to provide molecular and cellular indications to validate this hypothesis. Based on earlier findings that the milk of HTLV-1 infected women is rich in HTLV-1 infected lymphocytes that can transfer the virus into breast epithelial cells, the outcomes of this project may point that HTLV-1 can be a risk factor for the development of breast cancer, with a substantially higher risk to women who practice longterm breastfeeding. Our results showed that Tax strongly inhibited estrogen induced activation of BRCA-1 expression in breast cells by sequestering CBP/p300 co-activators. Trying to explore the CBP/p300 associated mechanism of Tax effect on BRCA1 expression, our results suggest that Tax does not prevent the binding of CBP/p300 to ERa but rather physically associates with the ERa-CBP/p300 to form a tertiary reporter. Since CBP/p300 complex has several binding domains, we believe that Tax associates with ERa-CBP/p300 complex through binding to CBP/p300 rather than to the ERa protein. We have also found that Tax inhibits BRCA1-mediated activation of p53-target promoters. These results support the possibility of HTLV-1 involvement in breast cancer development.

HTLV-1 Tax oncoprotein stimulates ROS production and apoptosis in T cells by interacting with USP10

AbstractHuman T-cell leukemia virus type 1 (HTLV-1) is the etiological agent of adult T-cell leukemia (ATL), and the viral oncoprotein Tax plays key roles in the immortalization of human T cells, lifelong persistent infection, and leukemogenesis. We herein identify the ubiquitin-specific protease 10 (USP10) as a Tax-interactor in HTLV-1–infected T cells. USP10 is an antistress factor against various environmental stresses, including viral infections and oxidative stress. On exposure to arsenic, an oxidative stress inducer, USP10 is recruited into stress granules (SGs), and USP10-containing SGs reduce reactive oxygen species (ROS) production and inhibit ROS-dependent apoptosis. We found that interaction of Tax with USP10 inhibits arsenic-induced SG formation, stimulates ROS production, and augments ROS-dependent apoptosis in HTLV-1–infected T cells. These findings suggest that USP10 is a host factor that inhibits stress-induced ROS production and apoptosis in HTLV-1–infected T cells; however, its activities are attenuated by Tax. A clinical study showed that combination therapy containing arsenic is effective against some forms of ATL. Therefore, these findings may be relevant to chemotherapy against ATL.

Kuan-Teh Jeang

Kuan-Teh Jeang

Influence of the factors OCT3/4, SOX2, KLF4 or c-Myc on cell transformation promoted by the oncoprotein tax from HTLV-I.

Event Abstract Back to Event Influence of the factors OCT3/4, SOX2, KLF4 or c-Myc on cell transformation promoted by the oncoprotein tax from HTLV-I. Gustavo E. Brito-Melo1* 1 Federal University of the Jequitinhonha and Mucuri Valleys - UFVM, Pharmacy, Brazil Human T-cell leukemia virus type 1 (HTLV-I) infects approximately 15-20 milion people worldwide and is able to cause Adult T-cell Leukemia (ATL). The virus encodes the oncoprotein Tax wich can immortalize human T-lymphocytes and transform rodent cells in vitro. However the mechanism involved in cell tranformation by Tax are not fully understood. It has been shown that cancers are comprised of two broad categories of cells: those with high- and those with limited- proliferative potential. Cells in the former category are termed cancer stem cells. CSCs share several properties with adult stem cells, particularly the abilities to self-renew and differentiate into multiple cell types. Our new research proposal addresses a key question "Can the HTLV-1 Tax oncoprotein transform embryonic stem cells better than primary cells?” In the present study NIH3T3 cells were transfected with both Tax and individual pluripotence-associated factors (OCT3/4, SOX2, KLF4 and cMYC into CMV3.1 expression plasmid) communly used for cell reprograming. Hella cells were used as tranformed cell control. The cell growing of the transfected cells was kept on soft agar and the cell colony counting (UFC) was achieved using ImageJ software. According to the results, cell transfection using Tax plus KLF4 (mean=197±13.4) and Tax plus OCT3/4 (mean=94.7±18.0) promoted a higher UFC counting as compared to the cells trasfected with Tax (mean=21,3±8,7) or the other factors, isolately. Althouth very preliminar, the results presented here point to an interessting way for future investigations on cell transformation pathways observed in HTLV-I infection. Acknowledgements We are greatfull to Fapemig, CNPq and CAPES financial agencies for supporting the study. Keywords: HTLV, pluripotency, factors, immunology, tax Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Translational immunology and immune intervention Citation: Brito-Melo GE (2013). Influence of the factors OCT3/4, SOX2, KLF4 or c-Myc on cell transformation promoted by the oncoprotein tax from HTLV-I.. Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.00719 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 17 Jun 2013; Published Online: 22 Aug 2013. * Correspondence: Prof. Gustavo E Brito-Melo, Federal University of the Jequitinhonha and Mucuri Valleys - UFVM, Pharmacy, Diamantina, Not Applicable, 39100000, Brazil, gbrito1998@yahoo.com Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Gustavo E Brito-Melo Google Gustavo E Brito-Melo Google Scholar Gustavo E Brito-Melo PubMed Gustavo E Brito-Melo Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Physical and functional interaction between the HTLV-1 Tax oncoprotein and the cellular LIM domain protein FHL3

HTLV-1 is the ethiologic agent of adult T-cell leukemia (ATL). The viral regulatory protein Tax1 plays a pivotal role in T-cell transformation by deregulating several signalling pathways including CREB, NF-κB and AP-1 leading to the abnormal expression of several cellular proteins involved in cell survival and proliferation. Previous studies in the lab, using the yeast-2-hybrid approach to screen a T-cell library for Tax1 interacting partners identified the cellular Four and a Half Lim domain protein 3 (FHL3) as a possible Tax1 interacting candidate. FHL3 can regulate the actin-cytoskeleton and functions as a transcriptional activator. The aim of this study is to investigate the physical and functional interaction between Tax1 and FHL3. We have found that Tax1 and FHL3 interact both in vitro and in vivo and deletion analysis of FHL3 revealed that each LIM domain of FHL3 can interact with Tax1. Deletion analysis of Tax1 determined that the interaction with FHL3 occurs at both the N and C terminus, namely amino acids 1-50 and a fragment which encompasses the predicted LIM binding domain 207-353aa. We have demonstrated that FHL3 enhances Tax1 mediated LTR transcriptional activation without affecting basal activity. In contrast to this we show that FHL3 down regulates NF-κB activation by Tax1. Using confocal microscopy, we show that FHL3 co localizes with Tax in the nucleus and at the periphery of co transfected cells. In addition we demonstrate that FHL3 induces nanotubules in Cos7 cells and may be capable of transporting Tax1 from one cell to another. Overall our results so far suggest that the interaction between Tax1 and FHL3 alters both the transactivating activity and the sub cellular localization of Tax1 and provide new insights into molecular mechanisms that underlie the oncogenic nature of this HTLV-1 protein.

Human T Cell Leukemia Virus Type 1 Tax Inhibits Innate Antiviral Signaling via NF-κB-Dependent Induction of SOCS1

Human T cell leukemia virus type 1 (HTLV-1) inhibits host antiviral signaling pathways although the underlying mechanisms are unclear. Here we found that the HTLV-1 Tax oncoprotein induced the expression of SOCS1, an inhibitor of interferon signaling. Tax required NF-κB, but not CREB, to induce the expression of SOCS1 in T cells. Furthermore, Tax interacted with SOCS1 in both transfected cells and in HTLV-1-transformed cell lines. Although SOCS1 is normally a short-lived protein, in the presence of Tax, the stability of SOCS1 was greatly increased. Accordingly, Tax enhanced the replication of a heterologous virus, vesicular stomatitis virus (VSV), in a SOCS1-dependent manner. Surprisingly, Tax required SOCS1 to inhibit RIG-I-dependent antiviral signaling, but not the interferon-induced JAK/STAT pathway. Inhibition of SOCS1 by RNA-mediated interference in the HTLV-1-transformed cell line MT-2 resulted in increased IFN-β expression accompanied by reduced HTLV-1 replication and p19(Gag) levels. Taken together, our results reveal that Tax inhibits antiviral signaling, in part, by hijacking an interferon regulatory protein.

Constitutive Activation of TAK1 by HTLV-1 Tax-dependent Overexpression of TAB2 Induces Activation of JNK-ATF2 but Not IKK-NF-κB

HTLV-1 Tax oncoprotein induces persistent activation of the transcription factor NF-kappaB and CREB (cAMP-response element-binding protein)/ATF. Transforming growth factor-beta-activated kinase 1 (TAK1) has been shown to play a critical role in these transcription factors. Here, we found that TAK1 was constitutively activated in Tax-positive HTLV-1-transformed T cells. Tax induced persistent overexpression of TAK1-binding protein 2 (TAB2), but not TAB3, which is essential for TAK1 activation. Surprisingly, TAK1 was not involved in the activation of NF-kappaB. On the other hand, JNK and p38 mitogen-activated protein kinases were activated by TAK1. In addition, ATF2, but not CREB, was a target for the TAK1-JNK pathway, and p38 negatively regulated TAK1 activity through TAB1 phosphorylation. These results indicate that Tax-mediated TAK1 activation is important for the activation of ATF2 rather than NF-kappaB.

Human T-cell leukemia virus type-1 Tax oncoprotein regulates G-protein signaling

AbstractHuman T-cell leukemia virus type-1 (HTLV-1) is associated with adult T-cell leukemia (ATL) and neurological syndromes. HTLV-1 encodes the oncoprotein Tax-1, which modulates viral and cellular gene expression leading to T-cell transformation. Guanine nucleotide–binding proteins (G proteins) and G protein–coupled receptors (GPCRs) constitute the largest family of membrane proteins known and are involved in the regulation of most biological functions. Here, we report an interaction between HTLV-1 Tax oncoprotein and the G-protein β subunit. Interestingly, though the G-protein β subunit inhibits Tax-mediated viral transcription, Tax-1 perturbs G-protein β subcellular localization. Functional evidence for these observations was obtained using conditional Tax-1–expressing transformed T-lymphocytes, where Tax expression correlated with activation of the SDF-1/CXCR4 axis. Our data indicated that HTLV-1 developed a strategy based on the activation of the SDF-1/CXCR4 axis in the infected cell; this could have tremendous implications for new therapeutic strategies.

Negative regulation of the SH2-homology-containing protein-tyrosine phosphatase-1 (Shp-1) P2 promoter by the HTLV-1 tax oncoprotein

Expression of SH2-Homology-Containing Protein-Tyrosine Phosphatase-1 (Shp-1), a candidate tumor suppressor, is repressed in HTLV-1 transformed lymphocyte cell lines, leukemic cells of Adult T-cell Leukemia (ATL) patients and other hematologic malignancies. However, the mechanisms underlying regulation and repression of Shp-1 remain unclear. Herein, we cloned the putative full-length, hematopoietic cell specific Shp-1 P2 promoter and identified a 277 bp promoter region. In a dose-dependent manner, wild type HTLV-1 Tax and its M22/M47 mutant derivatives profoundly inhibited Shp-1 P2 promoter activity. NF-kB, not CBP or p300, was found to markedly stimulate basal P2 promoter activity and reverse Tax-induced Promoter Silencing (TIPS) in Jurkat cells. Mutagenesis of two NF-kB binding sites in the core promoter region led to only modest inhibition of basal promoter activity but to complete loss of TIPS. Further studies show that TIPS is potentiated by the histone deacetylase-1 (HDAC1) and in co-immunoprecipitation studies NF-kB could compete with HDAC1 for association with Tax protein. In addition, chromatin immunoprecipitation (ChIP) studies showed that Tax recruits HDAC1 to the core promoter and displaces NF-kB binding. We propose that in TIPS, Tax recruits HDAC1 to the Shp-1 P2 promoter which results in deacetylation and dissociation of NF-kB from the promoter leading to attenuation of Shp-1 expression, thus providing a possible first step toward leukemogenesis through its silencing of this key immediate early negative regulator of IL-2 signaling.

Binding of HTLV-1 tax oncoprotein to the precursor of interleukin-16, a T cell PDZ domain-containing protein

HTLV-1 Tax oncoprotein interacts with various cellular factors and modulates transcription and the cell cycle. In that role it is sufficient to create T cell malignancies in the absence of HTLV-1 infection. HTLV-1 Tax protein has been reported to bind to cellular proteins containing PDZ domains in vitro. The precursor of human interleukin 16, pro-IL-16, is an abundant cellular protein present in human peripheral blood T cells. Pro-IL-16 contains three PDZ domains. It has been shown that expression of pro-IL-16 in pro-IL-16 negative cells induces a G 0/G 1 arrest in the cell cycle. The current studies demonstrate that Tax binds to pro-IL-16 in HTLV-1 infected human T cells. We mapped the Tax binding site to the first PDZ domain of pro-IL-16. Over-expression of Tax in COS cells resulted in fewer cells in G 0/G 1 consistent with its activity to induce G 1- to S-phase progression in lymphocytes, while over-expression of pro-IL-16 in COS cells resulted in G 0/G 1 arrest. Co-expression of wild type Tax with pro-IL-16 in COS cells negated the effects of pro-IL-16, an effect not observed with Tax mutated at its PDZ binding C-terminus. These results suggest that one of the effects of Tax on growth deregulation in HTLV-1 infected T cells might be mediated by its binding to pro-IL-16.

The HTLV-1 Tax Oncoprotein Attenuates DNA Damage Induced G1 Arrest and Enhances Apoptosis in p53 Null Cells

Transformation of cells by the human T cell leukemia virus type 1 occurs via mechanisms unique among oncogenic retroviruses. A prevailing hypothesis for HTLV-1-mediated cellular transformation is that expression of the viral transactivator, Tax, induces genomic instability. Tax-mediated failure in the cellular repair response is one possible mechanism for loss in genomic integrity. Here we have examined the in vivo repair response of Tax-expressing cells to determine the underlying defects that contribute to loss of genomic integrity. In these studies we examined the effects of de novo Tax-expression in naive “pre-neoplastic” REF52 cells. DNA-damage-induced p53 stabilization and concomitant transient stabilization of p21 were clearly evident in Tax-expressing cells. Likewise, the damage-induced apoptotic response of Tax-expressing cells was normal. However, the damage-induced G1 checkpoint was abrogated in either p53+ or p53− cellular backgrounds. Although nucleotide excision repair (NER) of asynchronous Tax-expressing cells was impaired, cell-cycle-independent assessment of NER in the global excision repair assay demonstrated comparable NER activity in Tax-expressing cells, suggesting that the failure of G1 checkpoint contributes to NER deficiency. Interestingly, we observed a dramatic increase in apoptosis and UV sensitivity of Tax-expressing p53−/− cells when compared to Tax-expressing p53+/+ cells. These data demonstrate that Tax-mediated cellular genomic instability arises from attenuation of cell-cycle checkpoint and imply a clonal dependence on p53 status separate from genomic integrity.

HTLV type 1 Tax oncoprotein binds to DNA topoisomerase I and inhibits its catalytic activity.

HTLV-1 Tax oncoprotein exerts pleiotropic effects on cellular regulatory systems, trans-activation and trans-repression of transcription and promotion of the cell cycle, through interaction with various cellular factors. Here, we report Tax-mediated inhibition of DNA topoisomerase I. The interaction of Tax with topoisomerase I was demonstrated in vitro and also in vivo, and inhibition of DNA relaxation activity of topoisomerase I was clearly shown. These results suggested that Tax has a novel potential to affect various cellular processes, such as transcription and maintenance of genomic stability, in which DNA topoisomerase I is involved.

HTLV-1 Tax Oncoprotein Binds to DNA Topoisomerase I and Inhibits Its Catalytic Activity

HTLV-1 Tax oncoprotein exerts pleiotropic effects on cellular regulatory systems, such as transcription and the cell cycle, through the interaction with various cellular factors. During our search for additional cellular targets of Tax using a yeast two-hybrid screening system, we isolated a cDNA encoding human DNA topoisomerase I. Tax was demonstrated to bind to topoisomerase I in vitro, and the Tax-topoisomerase I complex was also detected in HTLV-1-infected T-cells by immunoprecipitation. Furthermore, Tax inhibited the catalytic activity of topoisomerase I as measured by relaxation of supercoiled DNA, although complete inhibition was not observed under the conditions used. The binding of topoisomerase I to DNA was inhibited by the addition of the wild type of Tax but not by a mutant of Tax that cannot bind to topoisomerase I. Consistent with these observations, expression of Tax induced an in vivo reduction of the covalent association of topoisomerase I with chromosomal DNA, which accumulates in the presence of camptothecin. These results suggest that Tax has a novel potential to affect various cellular processes such as transcription and maintenance of genomic stability, in which DNA topoisomerase I is involved.

Tax oncoprotein of HTLV-1 binds to the human homologue of Drosophila discs large tumor suppressor protein, hDLG, and perturbs its function in cell growth control.

HTLV-1 Tax oncoprotein interacts with various cellular factors and modulates transcription and the cell cycle. To identify more cellular targets, we employed the yeast two hybrid system with Tax using a human cDNA library, and isolated a cDNA encoding the human counterpart of Drosophila discs large tumor suppressor protein, hDLG. Tax binding to hDLG was confirmed in vitro and also in HTLV-1-infected T-cells. Furthermore, hDLG was found to be efficiently phosphorylated in Tax-transfected cells and HTLV-1-infected T-cells. The C-terminus of Tax and the PDZ domain of hDLG were responsible for the binding of Tax to hDLG. The C-terminal peptide of Tax prevented the binding of hDLG to APC tumor suppressor gene product, suggesting inhibition of hDLG function by Tax. Over-expression of hDLG in NIH3T3 cells by microinjection induced a reduction of BrdU incorporation into DNA, but co-expression of Tax suppressed this inhibitory effect of hDLG. These results suggest that hDLG arrested the cell cycle and that Tax canceled this inhibitory action of hDLG through targeting hDLG. Therefore, Tax affects this novel regulatory pathway of the cell cycle alteration, of which seems to play a role in the development of human cancer.

The C-terminus of the HTLV-1 Tax oncoprotein mediates interaction with the PDZ domain of cellular proteins.

Infection by HTLV-1 has been correlated with the appearance of various proliferative or degenerative diseases. Some of these disorders have been observed in transgenic mice expressing the Tax protein, which is known to transactivate various viral and cellular promoters through interactions with several transcription factors. In this study we show that the C-terminus of this viral oncoprotein represents a motif permitting binding of Tax to the PDZ domains of several cellular proteins. A two-hybrid screen with Tax as bait indeed yielded complementary DNAs coding for six proteins including PDZ domains. Two of them correspond to truncated forms of the PSD-95 and beta1-syntrophin proteins, another clone codes for a protein homologous to the product of the C. elegans gene lin-7. The other three clones code for new human members of the PDZ family of cellular proteins. The interaction of Tax with the products of these clones was confirmed by immunoprecipitation assays in mammalian cells, and analysis of various mutants of Tax established the importance of the C-terminal amino acids for several of these interactions. These data suggest that Tax could perturb the normal function of targeted cellular proteins by strongly interacting with their PDZ domains.