Human T-cell Leukemia Virus Type 1 Replication Research Articles

HTLV-1 Proliferation after CD8+ Cell Depletion by Monoclonal Anti-CD8 Antibody Administration in Latently HTLV-1-Infected Cynomolgus Macaques.

Human T-cell leukemia virus type 1 (HTLV-1) induces chronic asymptomatic latent infection with a substantial proviral load but without significant viral replication in vivo. Cumulative studies have indicated involvement of CD8-positive (CD8+) cells, including virus-specific CD8+ T cells in the control of HTLV-1 replication. However, whether HTLV-1 expression from latently infected cells in vivo occurs in the absence of CD8+ cells remains unclear. Here, we examined the impact of CD8+ cell depletion by monoclonal anti-CD8 antibody administration on proviral load in HTLV-1-infected cynomolgus macaques. Five cynomolgus macaques were infected with HTLV-1 by inoculation with HTLV-1-producing cells. Administration of monoclonal anti-CD8 antibody in the chronic phase resulted in complete depletion of peripheral CD8+ T cells for approximately 2 months. All five macaques showed an increase in proviral load following CD8+ cell depletion, which peaked just before the reappearance of peripheral CD8+ T cells. Tax-specific CD8+ T-cell responses were detected in these recovered CD8+ T cells. Importantly, anti-HTLV-1 antibodies also increased after CD8+ cell depletion, indicating HTLV-1 antigen expression. These results provide evidence indicating that HTLV-1 can proliferate from the latent phase in the absence of CD8+ cells and suggest that CD8+ cells are responsible for the control of HTLV-1 replication. IMPORTANCE HTLV-1 can cause serious diseases such as adult T-cell leukemia (ATL) in humans after chronic asymptomatic latent infection with substantial proviral load. Proviruses are detectable in peripheral lymphocytes in HTLV-1 carriers, and the association of a higher proviral load with a higher risk of disease progression has been observed. However, neither substantial viral structural protein expression nor viral replication was detectable in vivo. Cumulative studies have indicated involvement of CD8+ cells, including virus-specific CD8+ T cells in the control of HTLV-1 replication. In the present study, we showed that CD8+ cell depletion by monoclonal anti-CD8 antibody administration results in HTLV-1 expression and an increase in proviral load in HTLV-1-infected cynomolgus macaques. Our results indicate that HTLV-1 can proliferate in the absence of CD8+ cells, suggesting that CD8+ cells are responsible for the control of HTLV-1 replication. This study provides insights into the mechanism of virus-host immune interaction in latent HTLV-1 infection.

Infection of the Ex Vivo Tonsil Model by HTLV-1 Envelope-Pseudotyped Viruses

Human T-cell leukemia virus type 1 (HTLV-1) is the causal agent of adult T-cell leukemia/lymphoma and HTLV-1-associated myelopathy/tropical spastic paraparesis. Its tropism is known to be broad in cultured cell lines, while in vivo data support a more selective transmission toward CD4+ T cells and the limited targeting of other hematopoietic cell types. An essential condition for HTLV-1 infection is cell-to-cell contact, to which both virological synapse and viral biofilm have been suggested to strongly contribute. As cell lines and animal models each present their own limitations in studying HTLV-1 replication, we have explored the use of an ex vivo model based on the secondary lymphoid tonsillar tissue. HIV-1 luciferase-expressing pseudotyped viruses bearing the HTLV-1 envelope protein at their surface were first shown to recapitulate the wide spectrum of infectivity of HTLV-1 toward various cell lines. Tonsil fragments were next exposed to pseudotyped viruses and shown to be reproducibly infected. Infection by HTLV-1 Env-pseudotyped viruses was blocked by different anti-gp46 antibodies, unlike infection by HIV-1 virions. The dose-dependent infection revealed a gradual increase in luciferase activity, which was again sensitive to anti-gp46 antibodies. Overall, these results suggest that the ex vivo tonsil model represents a reliable alternative for studying HTLV-1 replication and potentially viral latency, as well as early clonal formation.

HTLV-1 persistent infection and ATLL oncogenesis.

Human T-cell leukemia virus type 1 (HTLV-1) is an oncogenic retrovirus; whereas HTLV-1 mainly persists in the infected host cell as a provirus, it also causes a malignancy called adult T-cell leukemia/lymphoma (ATLL) in about 5% of infection. HTLV-1 replication is in most cases silent in vivo and viral de novo infection rarely occurs; HTLV-1 rather relies on clonal proliferation of infected T cells for viral propagation as it multiplies the number of the provirus copies. It is mechanistically elusive how leukemic clones emerge during the course of HTLV-1 infection in vivo and eventually cause the onset of ATLL. This review summarizes our current understanding of HTLV-1 persistence and oncogenesis, with the incorporation of recent cutting-edge discoveries obtained by high-throughput sequencing.

Is the HTLV-1 Retrovirus Targeted by Host Restriction Factors?

Human T cell leukemia virus type 1 (HTLV-1), the etiological agent of adult T cell leukemia/lymphoma (ATLL) and of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), was identified a few years before Human Immunodeficiency Virus (HIV). However, forty years later, our comprehension of HTLV-1 immune detection and the host immune responses to HTLV-1 is far more limited than for HIV. In addition to innate and adaptive immune responses that rely on specialized cells of the immune system, host cells may also express a range of antiviral factors that inhibit viral replication at different stages of the cycle, in a cell-autonomous manner. Multiple antiviral factors allowing such an intrinsic immunity have been primarily and extensively described in the context HIV infection. Here, we provide an overview of whether known HIV restriction factors might act on HTLV-1 replication. Interestingly, many of them do not exert any antiviral activity against HTLV-1, and we discuss viral replication cycle specificities that could account for these differences. Finally, we highlight future research directions that could help to identify antiviral factors specific to HTLV-1.

Characterizing the Interaction between the HTLV-1 Transactivator Tax-1 with Transcription Elongation Factor ELL2 and Its Impact on Viral Transactivation.

The human T-cell leukemia virus type 1 (HTLV-1)-encoded transactivator and oncoprotein Tax-1 is essential for HTLV-1 replication. We recently found that Tax-1 interacts with transcription elongation factor for RNA polymerase II 2, ELL2, which enhances Tax-1-mediated transactivation of the HTLV-1 promotor. Here, we characterize the Tax-1:ELL2 interaction and its impact on viral transactivation by confocal imaging, co-immunoprecipitation, and luciferase assays. We found that Tax-1 and ELL2 not only co-precipitate, but also co-localize in dot-like structures in the nucleus. Tax-1:ELL2 complex formation occurred independently of Tax-1 point mutations, which are crucial for post translational modifications (PTMs) of Tax-1, suggesting that these PTMs are irrelevant for Tax-1:ELL2 interaction. In contrast, Tax-1 deletion mutants lacking either N-terminal (aa 1–37) or C-terminal regions (aa 150–353) of Tax-1 were impaired in interacting with ELL2. Contrary to Tax-1, the related, non-oncogenic Tax-2B from HTLV-2B did not interact with ELL2. Finally, we found that ELL2-R1 (aa 1–353), which carries an RNA polymerase II binding domain, and ELL2-R3 (aa 515–640) are sufficient to interact with Tax-1; however, only ELL2-truncations expressing R1 could enhance Tax-1-mediated transactivation of the HTLV-1 promoter. Together, this study identifies domains in Tax-1 and ELL2 being required for Tax-1:ELL2 complex formation and for viral transactivation.

Essential Role of Human T Cell Leukemia Virus Type 1 orf-I in Lethal Proliferation of CD4+ Cells in Humanized Mice.

Human T cell leukemia virus type 1 (HTLV-1) is the ethological agent of adult T cell leukemia/lymphoma (ATLL) and a number of lymphocyte-mediated inflammatory conditions, including HTLV-1-associated myelopathy/tropical spastic paraparesis. HTLV-1 orf-I encodes two proteins, p8 and p12, whose functions in humans are to counteract innate and adaptive responses and to support viral transmission. However, the in vivo requirements for orf-I expression vary in different animal models. In macaques, the ablation of orf-I expression by mutation of its ATG initiation codon abolishes the infectivity of the molecular clone HTLV-1p12KO In rabbits, HTLV-1p12KO is infective and persists efficiently. We used humanized mouse models to assess the infectivity of both wild-type HTLV-1 (HTLV-1WT) and HTLV-1p12KO We found that NOD/SCID/γC-/- c-kit+ mice engrafted with human tissues 1 day after birth (designated NSG-1d mice) were highly susceptible to infection by HTLV-1WT, with a syndrome characterized by the rapid polyclonal proliferation and infiltration of CD4+ CD25+ T cells into vital organs, weight loss, and death. HTLV-1 clonality studies revealed the presence of multiple clones of low abundance, confirming the polyclonal expansion of HTLV-1-infected cells in vivo HTLV-1p12KO infection in a bone marrow-liver-thymus (BLT) mouse model prone to graft-versus-host disease occurred only following reversion of the orf-I initiation codon mutation within weeks after exposure and was associated with high levels of HTLV-1 DNA in blood and the expansion of CD4+ CD25+ T cells. Thus, the incomplete reconstitution of the human immune system in BLT mice may provide a window of opportunity for HTLV-1 replication and the selection of viral variants with greater fitness.IMPORTANCE Humanized mice constitute a useful model for studying the HTLV-1-associated polyclonal proliferation of CD4+ T cells and viral integration sites in the human genome. The rapid death of infected animals, however, appears to preclude the clonal selection typically observed in human ATLL, which normally develops in 2 to 5% of individuals infected with HTLV-1. Nevertheless, the expansion of multiple clones of low abundance in these humanized mice mirrors the early phase of HTLV-1 infection in humans, providing a useful model to investigate approaches to inhibit virus-induced CD4+ T cell proliferation.

Pentosan Polysulfate Demonstrates Anti-human T-Cell Leukemia Virus Type 1 Activities In Vitro and In Vivo.

Human T-cell leukemia virus type 1 (HTLV-1) infection causes T-cell leukemia and inflammatory diseases, most notably including HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). The underlying mechanism for the pathogenesis of HAM/TSP remains unclear. According to a recent clinical trial, a humanized antibody that targets CCR4+ cells ameliorates inflammation by reducing the number of infected cells in the central nervous system; this result suggests that the transmigration of HTLV-1-infected cells plays a crucial role in HAM/TSP. Partly due to the blood-brain barrier, current treatments for HAM/TSP are mostly palliative. Pentosan polysulfate (PPS), a semisynthetic glycosaminoglycan, has recently been used to treat HAM/TSP and was found to alleviate the symptoms. In this study, we investigated the effect of PPS on HTLV-1-infected cells and provide evidence for its efficacy in HAM/TSP. PPS was cytotoxic to certain HTLV-1-infected cells and significantly suppressed HTLV-1 virion production. PPS also efficiently inhibited HTLV-1 cell-cell transmission in T cells. In addition, PPS blocked HTLV-1 infection of primary endothelial cells (human umbilical vascular endothelial cells) and suppressed the subsequent induction of proinflammatory cytokine expression. Furthermore, PPS was found to inhibit the adhesion and transmigration of HTLV-1-infected cells. We also confirmed the anti-HTLV-1 effect of PPS in vivo using two mouse models. PPS blocked HTLV-1 infection in a mouse model with peripheral blood mononuclear cell (PBMC)-humanized NOD-scid IL2Rgammanull (huPBMC NSG) mice. PPS was also found to suppress the development of dermatitis and lung damage in HTLV-1 bZIP factor (HBZ)-transgenic (HBZ-Tg) mice, an HTLV-1 transgenic mouse model in which the mice develop systemic inflammation.IMPORTANCE HTLV-1 is the first human retrovirus to have been identified and is endemic in certain areas worldwide. HTLV-1 infection leads to the development of an inflammatory disease called HAM/TSP, a myelopathy characterized by slowly progressive spastic paraparesis. There have been no effective therapeutics available for HAM/TSP, but recently, a semisynthetic glycosaminoglycan, named pentosan polysulfate (PPS), has been found to alleviate the symptoms of HAM/TSP. Here we conducted a comprehensive study on the effect of PPS both in vitro and in vivo PPS demonstrated anti-HTLV-1 potential in infected cell lines, as shown by its suppressive effects on HTLV-1 replication and transmission and on the transmigration of infected T cells. Moreover, results obtained from two HTLV-1 mouse models demonstrate that PPS inhibits HTLV-1 infection and inflammation development in vivo Our work offers insights into the treatment of HAM/TSP by PPS and also suggests its possible use for treating other HTLV-1-induced inflammatory diseases.

Acidic polysaccharides isolated from marine algae inhibit the early step of viral infection

We examined the effects of various acidic polysaccharides isolated from marine algae on the infection and replication of human immunodeficiency virus type-1 (HIV-1), hepatitis B virus (HBV), hepatitis C virus (HCV), and human T-cell leukemia virus type-1 (HTLV-1). It was found that sulfated fucan polysaccharides, ascophyllan, and two fucoidans derived from different sources significantly inhibited the early step of HIV-1 (R9 and JR-FL) infection, while they did not affect the late step. The alginate oligomer consisted of uronic acids and sulfated-galactan porphyran showed no significant inhibitory effects. In addition, ascophyllan and two fucoidans inhibited the early step of HBV infection in a dose-dependent manner. Furthermore, these polysaccharides inhibited the early step of HCV infection but had no inhibitory effects on HTLV-1 replication. To further examine the specificity of these polysaccharides in viral infections, we used vesicular stomatitis virus (VSV)-G-pseudotyped HIV-1 infection. Ascophyllan, the two fucoidans, and alginate oligomer also potently inhibited VSV-G-pseudotyped HIV-1 infection in HeLa cells. Taken together, these results suggest that the acidic polysaccharides used in this study are capable of inhibiting the early step of viral infections depending on the polysaccharides but not in a strict species-specific manner.

How to Control HTLV-1-Associated Diseases: Preventing de Novo Cellular Infection Using Antiviral Therapy.

Five to ten million individuals are infected by Human T-cell Leukemia Virus type 1 (HTLV-1). HTLV-1 is transmitted through prolonged breast-feeding, by sexual contacts and by transmission of infected T lymphocytes through blood transfusion. One to ten percent of infected carriers will develop a severe HTLV-1-associated disease: Adult-T-cell leukemia/lymphoma (ATLL), or a neurological disorder named Tropical Spastic Paraparesis/HTLV-1 Associated Myelopathy (TSP/HAM). In vivo, HTLV-1 is mostly detected in CD4+ T-cells, and to a lesser extent in CD8+ T cells and dendritic cells. There is a strong correlation between HTLV-1 proviral load (PVL) and clinical status of infected individuals. Thus, reducing PVL could be part of a strategy to prevent or treat HTLV-1-associated diseases among carriers. Treatment of ATLL patients using conventional chemotherapy has very limited benefit. Some chronic and acute ATLL patients are, however, efficiently treated with a combination of interferon α and zidovudine (IFN-α/AZT), to which arsenic trioxide is added in some cases. On the other hand, no efficient treatment for TSP/HAM patients has been described yet. It is therefore crucial to develop therapies that could either prevent the occurrence of HTLV-1-associated diseases or at least block the evolution of the disease in the early stages. In vivo, reverse transcriptase (RT) activity is low in infected cells, which is correlated with a clonal mode of viral replication. This renders infected cells resistant to nucleoside RT inhibitors such as AZT. However, histone deacetylase inhibitors (HDACi) associated to AZT efficiently induces viral expression and prevent de novo cellular infection. In asymptomatic STLV-1 infected non-human primates, HDACi/AZT combination allows a strong decrease in the PVL. Unfortunately, rebound in the PVL occurs when the treatment is stopped, highlighting the need for better antiviral compounds. Here, we review previously used strategies targeting HTLV-1 replication. We also tested a series of HIV-1 RT inhibitors in an in vitro anti-HTLV-1 screen, and report that bis-POM-PMEA (adefovir dipivoxil) and bis-POC-PMPA (tenofovir disoproxil) are much more efficient compared to AZT to decrease HTLV-1 cell-to-cell transmission in vitro. Our results suggest that revisiting already established antiviral drugs is an interesting approach to discover new anti-HTLV-1 drugs.

Sporadic on/off switching of HTLV-1 Tax expression is crucial to maintain the whole population of virus-induced leukemic cells

Viruses causing chronic infection artfully manipulate infected cells to enable viral persistence in vivo under the pressure of immunity. Human T-cell leukemia virus type 1 (HTLV-1) establishes persistent infection mainly in CD4+ T cells in vivo and induces leukemia in this subset. HTLV-1-encoded Tax is a critical transactivator of viral replication and a potent oncoprotein, but its significance in pathogenesis remains obscure due to its very low level of expression in vivo. Here, we show that Tax is expressed in a minor fraction of leukemic cells at any given time, and importantly, its expression spontaneously switches between on and off states. Live cell imaging revealed that the average duration of one episode of Tax expression is ∼19 hours. Knockdown of Tax rapidly induced apoptosis in most cells, indicating that Tax is critical for maintaining the population, even if its short-term expression is limited to a small subpopulation. Single-cell analysis and computational simulation suggest that transient Tax expression triggers antiapoptotic machinery, and this effect continues even after Tax expression is diminished; this activation of the antiapoptotic machinery is the critical event for maintaining the population. In addition, Tax is induced by various cytotoxic stresses and also promotes HTLV-1 replication. Thus, it seems that Tax protects infected cells from apoptosis and increases the chance of viral transmission at a critical moment. Keeping the expression of Tax minimal but inducible on demand is, therefore, a fundamental strategy of HTLV-1 to promote persistent infection and leukemogenesis.

Perturbation of Human T-Cell Leukemia Virus Type 1 Particle Morphology by Differential Gag Co-Packaging

Human T-cell leukemia virus type 1 (HTLV-1) is an important cancer-causing human retrovirus that has infected approximately 15 million individuals worldwide. Many aspects of HTLV-1 replication, including virus particle structure and assembly, are poorly understood. Group-specific antigen (Gag) proteins labeled at the carboxy terminus with a fluorophore protein have been used extensively as a surrogate for fluorescence studies of retroviral assembly. How these tags affect Gag stoichiometry and particle morphology has not been reported in detail. In this study, we used an HTLV-1 Gag expression construct with the yellow fluorescence protein (YFP) fused to the carboxy-terminus as a surrogate for the HTLV-1 Gag-Pol to assess the effects of co-packaging of Gag and a Gag-YFP on virus-like particle (VLP) morphology and analyzed particles by cryogenic transmission electron microscopy (cryo-TEM). Scanning transmission electron microscopy (STEM) and fluorescence fluctuation spectroscopy (FFS) were also used to determine the Gag stoichiometry. We found that ratios of 3:1 (Gag:Gag-YFP) or greater resulted in a particle morphology indistinguishable from that of VLPs produced with the untagged HTLV-1 Gag, i.e., a mean diameter of ~113 nm and a mass of 220 MDa as determined by cryo-TEM and STEM, respectively. Furthermore, FFS analysis indicated that HTLV-1 Gag-YFP was incorporated into VLPs in a predictable manner at the 3:1 Gag:Gag-YFP ratio. Both STEM and FFS analyses found that the Gag copy number in VLPs produced with a 3:1 ratio of Gag:Gag-YFP was is in the range of 1500–2000 molecules per VLP. The observations made in this study indicate that biologically relevant Gag–Gag interactions occur between Gag and Gag-YFP at ratios of 3:1 or higher and create a Gag lattice structure in VLPs that is morphologically indistinguishable from that of VLPs produced with just untagged Gag. This information is useful for the quantitative analysis of Gag–Gag interactions that occur during virus particle assembly and in released immature particles.

Molecular Studies of HTLV-1 Replication: An Update.

Human T-cell leukemia virus type 1 (HTLV-1) was the first human retrovirus discovered. Studies on HTLV-1 have been instrumental for our understanding of the molecular pathology of virus-induced cancers. HTLV-1 is the etiological agent of an adult T-cell leukemia (ATL) and can lead to a variety of neurological pathologies, including HTLV-1-associated-myelopathy/tropical spastic paraparesis (HAM/TSP). The ability to treat the aggressive ATL subtypes remains inadequate. HTLV-1 replicates by (1) an infectious cycle involving virus budding and infection of new permissive target cells and (2) mitotic division of cells harboring an integrated provirus. Virus replication initiates host antiviral immunity and the checkpoint control of cell proliferation, but HTLV-1 has evolved elegant strategies to counteract these host defense mechanisms to allow for virus persistence. The study of the molecular biology of HTLV-1 replication has provided crucial information for understanding HTLV-1 replication as well as aspects of viral replication that are shared between HTLV-1 and human immunodeficiency virus type 1 (HIV-1). Here in this review, we discuss the various stages of the virus replication cycle—both foundational knowledge as well as current updates of ongoing research that is important for understanding HTLV-1 molecular pathogenesis as well as in developing novel therapeutic strategies.

Modes of Human T Cell Leukemia Virus Type 1 Transmission, Replication and Persistence.

Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus that causes cancer (Adult T cell Leukemia, ATL) and a spectrum of inflammatory diseases (mainly HTLV-associated myelopathy—tropical spastic paraparesis, HAM/TSP). Since virions are particularly unstable, HTLV-1 transmission primarily occurs by transfer of a cell carrying an integrated provirus. After transcription, the viral genomic RNA undergoes reverse transcription and integration into the chromosomal DNA of a cell from the newly infected host. The virus then replicates by either one of two modes: (i) an infectious cycle by virus budding and infection of new targets and (ii) mitotic division of cells harboring an integrated provirus. HTLV-1 replication initiates a series of mechanisms in the host including antiviral immunity and checkpoint control of cell proliferation. HTLV-1 has elaborated strategies to counteract these defense mechanisms allowing continuous persistence in humans.

153 Molecular pathogenesis by HTLV-1 bZIP factor

Human T-cell leukemia virus type 1 (HTLV-1) mainly transmits by cell-to-cell contact. Therefore, this virus induces proliferation of infected cells to facilitate transmission. In HTLV-1 infected individuals, infected cells proliferate in vivo. This proliferation of infected cells leads to the development of adult T-cell leukemia (ATL) and inflammatory diseases including HTLV-1 associated myelopathy (HAM). HTLV-1 bZIP factor (HBZ) is consistently expressed in ATL cells and HTLV-1 infected cells, and plays a critical role in their proliferation. Transgenic expression of HBZ causes T-cell lymphoma and inflammatory diseases, indicating that HBZ is critical for the pathogenesis of HTLV-1.We analyzed the function of HBZ and found that it possessed opposite functions to Tax in many pathways. We found that Tax could activate the classical Wnt pathway while HBZ inhibited it by interacting with TCF1/LEF1. The canonical Wnt pathway is activated in the thymus, and its transcription factors, TCF1/LEF1, suppressed the viral gene transcription by Tax. Conversely, the canonical Wnt pathway is suppressed in the peripheral T cells in which the non-canonical Wnt pathway is activated. HBZ enhanced the expression of Wnt5a, which is a ligand in non-canonical Wnt signaling. Knockdown of Wnt5a suppressed the proliferation and migration of ATL cells. Thus, activation of the non-canonical Wnt pathway by HBZ is linked with proliferation of ATL cells.These findings suggest that HTLV-1 is adapted to, not thymic T cells, but peripheral T cells. HBZ modulates the intracellular environment suitable for HTLV-1 replication and proliferation of infected cells.

Examination of the interactions between the HTLV-1-encoded Tax oncoprotein and the cellular coactivators CBP/p300

Human T-cell Leukemia virus type-1 (HTLV-1) causes several diseases including aggressive Adult T-cell Leukemia (ATL). A major player in the malignant transformation of infected cells is expression of Tax, the virally-encoded oncoprotein. Tax is a potent activator of HTLV-1 transcription. To activate viral transcription, Tax, together with the cellular protein CREB, bind to the viral CRE enhancer elements located in the HTLV-1 promoter. Together, these activators recruit the homologous coactivator proteins CBP to initiate strong transcription of the provirus. Both CBP/p300 are involved in multiple cellular pathways, and deregulation of these proteins is associated with numerous pathologies. Despite extensive work on Tax-stimulated recruitment of CBP/p300 to the HTLV-1 promoter, primarily via their KIX domain, potential interactions between Tax and other domains of CBP/p300 have not been fully investigated. Recently, we have found that Tax interacts with several additional domains of the coactivators, however the significance of these interactions remains incompletely understood. We hypothesize that Tax interaction with various domains of CBP/p300 stimulates the potency of the coactivators and enhanced transcription. Our studies utilize protein-protein and protein-DNA interaction assays using individual domains of the coactivators, together with mutations and deletions that specifically target the Tax-CBP/p300 interface. Competition and activity assays enable us to functionally and biophysically characterize Tax-coactivator interactions. These studies have implications for both viral and cellular mechanisms of gene expression, and expand our understanding of the role of Tax and the cellular coactivators in HTLV-1 transcription and replication.

PA28γ is a novel corepressor of HTLV-1 replication and controls viral latency

AbstractThe establishment of a latent reservoir by human tumor viruses is a vital step in initiating cellular transformation and represents a major shortcoming to current therapeutic strategies and the ability to eradicate virus-infected cells. Human T-cell leukemia virus type 1 (HTLV-1) establishes a lifelong infection and is linked to adult T-cell leukemia lymphoma (ATLL). Here, we demonstrate that HTLV-1 p30 recruits the cellular proteasome activator PA28γ onto the viral tax/rex mRNA to prevent its nuclear export and suppress virus replication. Interaction of p30 with a PA28γ retaining fully functional proteasome activity is required for p30's ability to repress HTLV-1. Consistently, HTLV-1 molecular clones replicate better and produce more virus particles in PA28γ-deficient cells. These results define a unique and novel role for the cellular factor PA28γ in the control of nuclear RNA trafficking and HTLV-1–induced latency. Importantly, knockdown of PA28γ expression in ATLL cells latently infected with HTLV-1 reactivates expression of viral tax/rex RNA and the Tax protein. Because Tax is the most immunogenic viral antigen and triggers strong CTL responses, our results suggest that PA28γ-targeted therapy may reactivate virus expression from latently infected cells and allow their eradication from the host.

Suppression of HTLV-1 replication by Tax-mediated rerouting of the p13 viral protein to nuclear speckles

AbstractDisease development in human T-cell leukemia virus type 1 (HTLV-1)–infected individuals is positively correlated with the level of integrated viral DNA in T cells. HTLV-1 replication is positively regulated by Tax and Rex and negatively regulated by the p30 and HBZ proteins. In the present study, we demonstrate that HTLV-1 encodes another negative regulator of virus expression, the p13 protein. Expressed separately, p13 localizes to the mitochondria, whereas in the presence of Tax, part of it is ubiquitinated, stabilized, and rerouted to the nuclear speckles. The p13 protein directly binds Tax, decreases Tax binding to the CBP/p300 transcriptional coactivator, and, by reducing Tax transcriptional activity, suppresses viral expression. Because Tax stabilizes its own repressor, these findings suggest that HTLV-1 has evolved a complex mechanism to control its own replication. Further, these results highlight the importance of studying the function of the HTLV-1 viral proteins, not only in isolation, but also in the context of full viral replication.

New insights into HTLV-1 particle structure, assembly, and Gag-Gag interactions in living cells.

Human T-cell leukemia virus type 1 (HTLV-1) has a reputation for being extremely difficult to study in cell culture. The challenges in propagating HTLV-1 has prevented a rigorous analysis of how these viruses replicate in cells, including the detailed steps involved in virus assembly. The details for how retrovirus particle assembly occurs are poorly understood, even for other more tractable retroviral systems. Recent studies on HTLV-1 using state-of-the-art cryo-electron microscopy and fluorescence-based biophysical approaches explored questions related to HTLV-1 particle size, Gag stoichiometry in virions, and Gag-Gag interactions in living cells. These results provided new and exciting insights into fundamental aspects of HTLV-1 particle assembly—which are distinct from those of other retroviruses, including HIV-1. The application of these and other novel biophysical approaches promise to provide exciting new insights into HTLV-1 replication.

Tax-mediated re-routing of the HTLV-1 p13 protein to nuclear speckles

Human T-cell Leukemia Virus type-1 (HTLV-1) is highly immunogenic and has low variability so tight regulation of its expression is essential for virus maintenance in vivo. HTLV-1 replication is positively regulated by Tax and Rex, and negatively by the p30 and HBZ proteins. Here, we demonstrate that HTLV-1 encodes another negative regulator of virus expression, the p13 protein. Expressed separately, p13 localizes to the mitochondria, but in the presence of Tax, p13 is stabilized, and re-routed to the nuclear speckles. The p13 protein directly binds Tax, decreases Tax binding to the CBP/ p300 transcriptional co-activator and, by reducing Tax transcriptional activity, suppresses viral expression. These findings suggest that HTLV-1 has evolved a complex mechanism to control its own replication through regulation of positive and negative viral proteins. Further, these results emphasize the importance of studying the function of the HTLV-1 viral proteins, not only in isolation, but also in the context of full viral replication.

HTLV-1 Evades Type I Interferon Antiviral Signaling by Inducing the Suppressor of Cytokine Signaling 1 (SOCS1)

Human T cell leukemia virus type 1 (HTLV-1) is the etiologic agent of Adult T cell Leukemia (ATL) and the neurological disorder HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Although the majority of HTLV-1–infected individuals remain asymptomatic carriers (AC) during their lifetime, 2–5% will develop either ATL or HAM/TSP, but never both. To better understand the gene expression changes in HTLV-1-associated diseases, we examined the mRNA profiles of CD4+ T cells isolated from 7 ATL, 12 HAM/TSP, 11 AC and 8 non-infected controls. Using genomic approaches followed by bioinformatic analysis, we identified gene expression pattern characteristic of HTLV-1 infected individuals and particular disease states. Of particular interest, the suppressor of cytokine signaling 1—SOCS1—was upregulated in HAM/TSP and AC patients but not in ATL. Moreover, SOCS1 was positively correlated with the expression of HTLV-1 mRNA in HAM/TSP patient samples. In primary PBMCs transfected with a HTLV-1 proviral clone and in HTLV-1-transformed MT-2 cells, HTLV-1 replication correlated with induction of SOCS1 and inhibition of IFN-α/β and IFN-stimulated gene expression. Targeting SOCS1 with siRNA restored type I IFN production and reduced HTLV-1 replication in MT-2 cells. Conversely, exogenous expression of SOCS1 resulted in enhanced HTLV-1 mRNA synthesis. In addition to inhibiting signaling downstream of the IFN receptor, SOCS1 inhibited IFN-β production by targeting IRF3 for ubiquitination and proteasomal degradation. These observations identify a novel SOCS1 driven mechanism of evasion of the type I IFN antiviral response against HTLV-1.