The HTLV-I oncoprotein Tax inactivates the tumor suppressor FBXW7.

Human T-cell leukemia virus type 1 (HTLV-I) is the etiological agent of adult T-cell leukemia (ATL). Mutational analysis has demonstrated that the tumor suppressor, F-box and WD repeat domain containing 7 (FBXW7/FBW7/CDC4), is mutated in primary ATL patients. However, even in the absence of genetic mutations, FBXW7 substrates are stabilized in ATL cells, suggesting additional mechanisms can prevent FBXW7 functions. Here, we report that the viral oncoprotein Tax represses FBXW7 activity, resulting in the stabilization of activated Notch intracellular domain, c-MYC, Cyclin E, and myeloid cell leukemia sequence 1 (BCL2-related) (Mcl-1). Mechanistically, we demonstrate that Tax directly binds to FBXW7 in the nucleus, effectively outcompeting other targets for binding to FBXW7, resulting in decreased ubiquitination and degradation of FBXW7 substrates. In support of the nuclear role of Tax, a non-degradable form of the nuclear factor kappa B subunit 2 (NFκB2/p100) was found to delocalize Tax to the cytoplasm, thereby preventing Tax interactions with FBXW7 and Tax-mediated inhibition of FBXW7. Finally, we characterize a Tax mutant that is unable to interact with FBXW7, unable to block FBXW7 tumor suppressor functions, and unable to effectively transform fibroblasts. These results demonstrate that HTLV-I Tax can inhibit FBXW7 functions without genetic mutations to promote an oncogenic state. These results suggest that Tax-mediated inhibition of FBXW7 is likely critical during the early stages of the cellular transformation process. IMPORTANCE: F-box and WD repeat domain containing 7 (FBXW7), a critical tumor suppressor of human cancers, is frequently mutated or epigenetically suppressed. Loss of FBXW7 functions is associated with stabilization and increased expression of oncogenic factors such as Cyclin E, c-Myc, Mcl-1, mTOR, Jun, and Notch. In this study, we demonstrate that the human retrovirus human T-cell leukemia virus type 1 oncoprotein Tax directly interacts with FBXW7, effectively outcompeting other targets for binding to FBXW7, resulting in decreased ubiquitination and degradation of FBXW7 cellular substrates. We further demonstrate that a Tax mutant unable to interact with and inactivate FBXW7 loses its ability to transform primary fibroblasts. Collectively, our results describe a novel mechanism used by a human tumor virus to promote cellular transformation.

Increased H19/miR-675 Expression in Adult T-Cell Leukemia Is Associated with a Unique Notch Signature Pathway.

The Notch pathway is a key cancer driver and is important in tumor progression. Early research suggested that Notch activity was highly dependent on the expression of the intracellular cleaved domain of Notch-1 (NICD). However, recent insights into Notch signaling reveal the presence of Notch pathway signatures, which may vary depending on different cancer types and tumor microenvironments. Herein, we perform a comprehensive investigation of the Notch signaling pathway in adult T-cell leukemia (ATL) primary patient samples. Using gene arrays, we demonstrate that the Notch pathway is constitutively activated in ATL patient samples. Furthermore, the activation of Notch in ATL cells remains elevated irrespective of the presence of activating mutations in Notch itself or its repressor, FBXW7, and that ATL cells are dependent upon Notch-1 expression for proliferation and survival. We demonstrate that ATL cells exhibit the expression of pivotal Notch-related genes, including notch-1, hes1, c-myc, H19, and hes4, thereby defining a critical Notch signature associated with ATL disease. Finally, we demonstrate that lncRNA H19 is highly expressed in ATL patient samples and ATL cells and contributes to Notch signaling activation. Collectively, our results shed further light on the Notch pathway in ATL leukemia and reveal new therapeutic approaches to inhibit Notch activation in ATL cells.

HTLV-1 Tax Tug-of-War: Cellular Senescence and Death or Cellular Transformation.

Human T cell leukemia virus type 1 (HTLV-1) is a retrovirus associated with a lymphoproliferative disease known as adult T cell leukemia/lymphoma (ATLL). HTLV-1 infection efficiently transforms human T cells in vivo and in vitro. The virus does not transduce a proto-oncogene, nor does it integrate into tumor-promoting genomic sites. Instead, HTLV-1 uses a random mutagenesis model, resulting in cellular transformation. Expression of the viral protein Tax is critical for the immortalization of infected cells by targeting specific cellular signaling pathways. However, Tax is highly immunogenic and represents the main target for the elimination of virally infected cells by host cytotoxic T cells (CTLs). In addition, Tax expression in naïve cells induces pro-apoptotic signals and has been associated with the induction of non-replicative cellular senescence. This review will explore these conundrums and discuss the mechanisms used by the Tax viral oncoprotein to influence life-and-death cellular decisions and affect HTLV-1 pathogenesis.

Targeting Pim kinases in hematological cancers: molecular and clinical review.

Decades of research has recognized a solid role for Pim kinases in lymphoproliferative disorders. Often up-regulated following JAK/STAT and tyrosine kinase receptor signaling, Pim kinases regulate cell proliferation, survival, metabolism, cellular trafficking and signaling. Targeting Pim kinases represents an interesting approach since knock-down of Pim kinases leads to non-fatal phenotypes in vivo suggesting clinical inhibition of Pim may have less side effects. In addition, the ATP binding site offers unique characteristics that can be used for the development of small inhibitors targeting one or all Pim isoforms. This review takes a closer look at Pim kinase expression and involvement in hematopoietic cancers. Current and past clinical trials and in vitro characterization of Pim kinase inhibitors are examined and future directions are discussed. Current studies suggest that Pim kinase inhibition may be most valuable when accompanied by multi-drug targeting therapy.

Clinical significance of FBXW7 loss of function in human cancers.

FBXW7 (F-Box and WD Repeat Domain Containing 7) (also referred to as FBW7 or hCDC4) is a component of the Skp1-Cdc53 / Cullin-F-box-protein complex (SCF/ß-TrCP). As a member of the F-box protein family, FBXW7 serves a role in phosphorylation-dependent ubiquitination and proteasome degradation of oncoproteins that play critical role(s) in oncogenesis. FBXW7 affects many regulatory functions involved in cell survival, cell proliferation, tumor invasion, DNA damage repair, genomic instability and telomere biology. This thorough review of current literature details how FBXW7 expression and functions are regulated through multiple mechanisms and how that ultimately drives tumorigenesis in a wide array of cell types. The clinical significance of FBXW7 is highlighted by the fact that FBXW7 is frequently inactivated in human lung, colon, and hematopoietic cancers. The loss of FBXW7 can serve as an independent prognostic marker and is significantly correlated with the resistance of tumor cells to chemotherapeutic agents and poorer disease outcomes. Recent evidence shows that genetic mutation of FBXW7 differentially affects the degradation of specific cellular targets resulting in a distinct and specific pattern of activation/inactivation of cell signaling pathways. The clinical significance of FBXW7 mutations in the context of tumor development, progression, and resistance to therapies as well as opportunities for targeted therapies is discussed.

Feedback Loop Regulation between Pim Kinases and Tax Keeps Human T-Cell Leukemia Virus Type 1 Viral Replication in Check.

The Pim family of serine/threonine kinases promote tumorigenesis by enhancing cell survival and inhibiting apoptosis. Three isoforms exist, Pim-1, -2, and -3, that are highly expressed in hematological cancers, including Pim-1 in adult T-cell leukemia (ATL). Human T-cell leukemia virus type-1 (HTLV-1) is the etiological agent of ATL, a dismal lymphoproliferative disease known as adult T-cell leukemia. The HTLV-1 virally encoded oncogene Tax promotes CD4+ T-cell transformation through disruption of DNA repair pathways and activation of survival and cellular proliferation pathways. In this study, we found Tax increases the expression of Pim-1 and Pim-3, while decreasing Pim-2 expression. Furthermore, we discovered that Pim-1, -2, and -3 bind Tax protein to reduce its expression thereby creating a feedback regulatory loop between these two oncogenes. The loss of Tax expression triggered by Pim kinases led to loss in Tax-mediated transactivation of the HTLV-1 long terminal repeat (LTR) and reductions in HTLV-1 virus replication. Because Tax is also the immunodominant cytotoxic T cell lymphocytes (CTL) target, our data suggest that Pim kinases may play an important role in immune escape of HTLV-1-infected cells. IMPORTANCE The Pim family of protein kinases have established pro-oncogenic functions. They are often upregulated in cancer; especially leukemias and lymphomas. In addition, a role for Pim kinases in control of virus expression and viral latency is important for Kaposi sarcoma-associated herpesvirus (KSHV) and human immunodeficiency virus type 1 (HIV-1). Our data demonstrate that HTLV-1 encodes viral genes that promote and maintain Pim kinase activation, which in turn may stimulate T-cell transformation and maintain ATL leukemic cell growth. HTLV-1 Tax increases expression of Pim-1 and Pim-3, while decreasing expression of Pim-2. In ATL cells, Pim expression is maintained through extended protein half-life and heat shock protection. In addition, we found that Pim kinases have a new role during HTLV-1 infection. Pim-1, -2, and -3 can subvert Tax expression and HTLV-1 virus production. This may lead to partial suppression of the host immunogenic responses to Tax and favor immune escape of HTLV-1-infected cells. Therefore, Pim kinases have not only pro-oncogenic roles but also favor persistence of the virus-infected cell.

Germinal epimutation of Fragile Histidine Triad (FHIT) gene is associated with progression to acute and chronic adult T-cell leukemia diseases.

BACKGROUND: Human T cell Leukemia virus type 1 (HTLV-I) is etiologically linked to adult T cell leukemia/lymphoma (ATL) and an inflammatory neurodegenerative disease called HTLV-I-associated myelopathy or tropical spastic paraparesis (HAM/TSP). The exact genetic or epigenetic events and/or environmental factors that influence the development of ATL, or HAM/TSP diseases are largely unknown. The tumor suppressor gene, Fragile Histidine Triad Diadenosine Triphosphatase (FHIT), is frequently lost in cancer through epigenetic modifications and/or deletion. FHIT is a tumor suppressor acting as genome caretaker by regulating cellular DNA repair. Indeed, FHIT loss leads to replicative stress and accumulation of double DNA strand breaks. Therefore, loss of FHIT expression plays a key role in cellular transformation. METHODS: Here, we studied over 400 samples from HTLV-I-infected individuals with ATL, TSP/HAM, or asymptomatic carriers (AC) for FHIT loss and expression. We examined the epigenetic status of FHIT through methylation specific PCR and bisulfite sequencing; and correlated these results to FHIT expression in patient samples. RESULTS: We found that epigenetic alteration of FHIT is specifically found in chronic and acute ATL but is absent in asymptomatic HTLV-I carriers and TSP/HAM patients' samples. Furthermore, the extent of FHIT methylation in ATL patients was quantitatively comparable in virus-infected and virus non-infected cells. We also found that longitudinal HTLV-I carriers that progressed to smoldering ATL and descendants of ATL patients harbor FHIT methylation. CONCLUSIONS: These results suggest that germinal epigenetic mutation of FHIT represents a preexisting mark predisposing to the development of ATL diseases. These findings have important clinical implications as patients with acute ATL are rarely cured. Our study suggests an alternative strategy to the current "wait and see approach" in that early screening of HTLV-I-infected individuals for germinal epimutation of FHIT and early treatment may offer significant clinical benefits.

FHIT as a biomarker for early screening of adult T-cell leukemia.

Adult T-cell leukemia (ATL) is an incurable leukemia deriving from human T-cell leukemia virus (HTLV-I) infected cells. In our most recent study, we discovered that methylation of the tumor suppressor, fragile histidine triad gene (FHIT), exists in the majority of acute and chronic ATL patients. Methylation was seen in non-tumorigenic cells, in cells with low levels of HTLV-I integrated DNA, in longitudinal samples from HTLV-I carriers, in a percentage of HTLV-I carriers, and in direct descendants of ATL patients. Overall, this suggests that FHIT methylation is likely present in patients, prior to HTLV-I infection, and predisposes HTLV-I carriers to ATL development. In this commentary we discuss the importance of developing diagnostic tools for the early detection of FHIT methylation and the possibility that prior FHIT methylation may predispose any individual to the development of cancer.

Loss of FBXW7-mediated degradation of BRAF elicits resistance to BET inhibitors in adult T cell leukemia cells.

BACKGROUND: Human T cell leukemia virus type 1 (HTLV-1)-associated adult T cell leukemia (ATL) has a very poor prognosis with a median survival of 8 months and a 4-year overall survival of 11% for acute ATL. Present treatment options are limited and there is no curative therapy for ATL. Ubiquitin ligase FBXW7 is commonly mutated or functionally inactivated in human cancers. Consistent with the notion that FBXW7 controls the degradation of many oncoproteins, loss of FBXW7 has been linked to increased drug resistance or sensitivity in cancer cells. METHOD: In this study, we have characterized FBXW7 mutants previously identified in HTLV-I-infected ATL patient samples. TET-inducible ATL cells carrying wild type or mutated FBXW7 were analyzed for target degradation and for drug sensitivity. RESULTS: Our results demonstrate that mutations in FBXW7 can selectively disrupt ubiquitination and proteasome degradation of target proteins: c-MYC, cyclin E and MCL1. Both c-MYC and MYCN were highly expressed in uncultured ATL patient's samples and ATL-derived cell lines; and ATL cells demonstrated sensitivity to BET inhibitors in vitro and in vivo. High-throughput reverse phase protein array revealed BRAF as a novel target of FBXW7 and further experiments showed that mutations in FBXW7 preventing degradation of BRAF provided resistance to BET inhibitors. In contrast to R465, hot spot FBXW7 mutations at R505C retained degradation of BRAF but not NOTCH1, c-MYC, cyclin E, or MCL1. Finally, a combination therapy using BET inhibitors along with a BRAF or an ERK inhibitor prevented tumor cell resistance and growth. CONCLUSION: Our results suggest that FBXW7 status may play an important role in drug therapy resistance of cancer cells. Genetic characterization of FBXW7 may be one factor included in future personalized cancer treatment identification.

JAG1 overexpression contributes to Notch1 signaling and the migration of HTLV-1-transformed ATL cells.

BACKGROUND: HTLV-1 is a retrovirus that infects over 20 million people worldwide and is responsible for the hematopoietic malignancy adult T cell leukemia (ATL). We previously demonstrated that Notch is constitutively activated in ATL cells. Activating genetic mutations were found in Notch; however, Notch signaling was also activated in the absence of genetic mutations suggesting the existence of other mechanisms. METHODS: We analyzed the expression of Notch receptor ligands in HTLV-I-transformed cells, ATL patient-derived cell lines, and fresh uncultured ATL samples by RT-PCR, FACS, and immunohistochemistry. We then investigated viral and cellular molecular mechanisms regulating expression of JAG1. Finally, using shRNA knock-down and neutralizing antibodies, we investigated the function of JAG1 in ATL cells. RESULTS: Here, we report the overexpression of the Notch ligand, JAG1, in freshly uncultured ATL patient samples compared to normal PBMCs. We found that in ATL cells, JAG1 overexpression relies upon the viral protein Tax and cellular miR-124a, STAT3, and NFATc1. Interestingly, our data show that blockade of JAG1 signaling dampens Notch1 downstream signaling and limits cell migration of transformed ATL cells. CONCLUSIONS: Our results suggest that targeting JAG1 can block Notch1 activation in HTLV-I-transformed cells and represents a new target for immunotherapy in ATL patients.

FBXW7: a critical tumor suppressor of human cancers.

The ubiquitin-proteasome system (UPS) is involved in multiple aspects of cellular processes, such as cell cycle progression, cellular differentiation, and survival (Davis RJ et al., Cancer Cell 26:455-64, 2014; Skaar JR et al., Nat Rev Drug Discov 13:889-903, 2014; Nakayama KI and Nakayama K, Nat Rev Cancer 6:369-81, 2006). F-box and WD repeat domain containing 7 (FBXW7), also known as Sel10, hCDC4 or hAgo, is a member of the F-box protein family, which functions as the substrate recognition component of the SCF E3 ubiquitin ligase. FBXW7 is a critical tumor suppressor and one of the most commonly deregulated ubiquitin-proteasome system proteins in human cancer. FBXW7 controls proteasome-mediated degradation of oncoproteins such as cyclin E, c-Myc, Mcl-1, mTOR, Jun, Notch and AURKA. Consistent with the tumor suppressor role of FBXW7, it is located at chromosome 4q32, a genomic region deleted in more than 30% of all human cancers (Spruck CH et al., Cancer Res 62:4535-9, 2002). Genetic profiles of human cancers based on high-throughput sequencing have revealed that FBXW7 is frequently mutated in human cancers. In addition to genetic mutations, other mechanisms involving microRNA, long non-coding RNA, and specific oncogenic signaling pathways can inactivate FBXW7 functions in cancer cells. In the following sections, we will discuss the regulation of FBXW7, its role in oncogenesis, and the clinical implications and prognostic value of loss of function of FBXW7 in human cancers.

Telomere Dynamics in Immune Senescence and Exhaustion Triggered by Chronic Viral Infection.

The progressive loss of immunological memory during aging correlates with a reduced proliferative capacity and shortened telomeres of T cells. Growing evidence suggests that this phenotype is recapitulated during chronic viral infection. The antigenic volume imposed by persistent and latent viruses exposes the immune system to unique challenges that lead to host T-cell exhaustion, characterized by impaired T-cell functions. These dysfunctional memory T cells lack telomerase, the protein capable of extending and stabilizing chromosome ends, imposing constraints on telomere dynamics. A deleterious consequence of this excessive telomere shortening is the premature induction of replicative senescence of viral-specific CD8+ memory T cells. While senescent cells are unable to expand, they can survive for extended periods of time and are more resistant to apoptotic signals. This review takes a closer look at T-cell exhaustion in chronic viruses known to cause human disease: Epstein-Barr virus (EBV), Hepatitis B/C/D virus (HBV/HCV/HDV), human herpesvirus 8 (HHV-8), human immunodeficiency virus (HIV), human T-cell leukemia virus type I (HTLV-I), human papillomavirus (HPV), herpes simplex virus-1/2(HSV-1/2), and Varicella-Zoster virus (VZV). Current literature linking T-cell exhaustion with critical telomere lengths and immune senescence are discussed. The concept that enduring antigen stimulation leads to T-cell exhaustion that favors telomere attrition and a cell fate marked by enhanced T-cell senescence appears to be a common endpoint to chronic viral infections.

Oncogenic mutations in the FBXW7 gene of adult T-cell leukemia patients.

Human T-cell leukemia virus type 1 (HTLV-I) is associated with adult T-cell leukemia (ATL), an aggressive lymphoproliferative disease with a dismal prognosis. We have previously described the presence of Notch1 activating mutations and constitutive Notch1 signaling in patients with acute ATL. In this study, we report a high frequency of F-box and WD repeat domain containing 7 (FBXW7)/hCDC4 mutations within the WD40 substrate-binding domain in 8 of 32 acute ATL patients (25%). Functionally, ATL FBXW7 mutants lost their ability to interact with intracellular Notch (NICD), resulting in increased protein stability and constitutive Notch1 signaling. Consistent with the loss-of-function found in ATL patients, expression of WT FBXW7 in several patient-derived ATL lines demonstrated strong tumor-suppressor activity characterized by reduced proliferation of ATL cells. Remarkably, two FBXW7 mutants, D510E and D527G, demonstrated oncogenic activity when expressed in the presence of HTLV-I Tax, mutated p53 R276H, or c-Myc F138C found in human cancers. Transforming activity was further demonstrated by the ability of the FBXW7 D510E mutant to provide IL-2-independent growth of Tax-immortalized human T cells and increase the tumor formation in a xenograft mouse model of ATL. This study suggests that FBXW7, normally a tumor suppressor, can act as an oncogene when mutated and may play an important role in the pathogenesis of ATL.

Constitutive activation of Pim1 kinase is a therapeutic target for adult T-cell leukemia.

Human T-cell leukemia virus type 1 (HTLV-1)-associated adult T-cell leukemia and T-cell lymphoma (ATL) are aggressive diseases with poor prognoses, limited therapeutic options, and no curative treatment. In this study, we used a mouse model of ATL and restored expression of the microRNA, miR-124a, to identify in vivo downstream effectors responsible for its tumor-suppressive functions in ATL cells. Our results revealed that STAT3, a direct target of miR-124a, is constitutively activated in HTLV-I-transformed cells and ATL cells, and activating STAT3 mutations were detected in 25.5% of primary ATL patients. Interestingly, we found that the STAT3 downstream kinase effector, Pim1, is constitutively activated in ATL cells. The dependence of ATL cells to Pim1 activity was demonstrated using 2 Pim1 small inhibitors, SMI-4a and AZD1208. These studies indicated that HTLV-I-transformed and ATL cells, but not normal peripheral blood mononuclear cells, are highly sensitive to AZD1208, and the inhibition of Pim1 signaling triggers an apoptotic signal in leukemic cells. Finally, preclinical testing of AZD1208 in a mouse model of ATL resulted in significant prevention of tumor growth in vivo. In conclusion, our studies suggest that constitutive activation of the STAT3-Pim1 pathway represents a novel therapeutic target for the treatment of ATL.

Multiple Pathways Control the Reactivation of Telomerase in HTLV-I-Associated Leukemia.

While telomerase (hTERT) activity is absent from normal somatic cells, reactivation of hTERT expression is a hallmark of cancer cells. Telomerase activity is required for avoiding replicative senescence and supports immortalization of cellular proliferation. Only a minority of cancer cells rely on a telomerase-independent process known as alternative lengthening of telomeres, ALT, to sustain cancer cell proliferation. Multiple genetic, epigenetic, and viral mechanisms have been found to de-regulate telomerase gene expression, thereby increasing the risk of cellular transformation. Here, we review the different strategies used by the Human T-cell leukemia virus type 1, HTLV-I, to activate hTERT expression and stimulate its enzymatic activity in virally infected CD4 T cells. The implications of hTERT reactivation in HTLV-I pathogenesis and disease treatment are discussed.

STAT1: A Novel Target of miR-150 and miR-223 Is Involved in the Proliferation of HTLV-I-Transformed and ATL Cells.

We have previously reported on the deregulation of cellular microRNAs involved in hematopoiesis and inflammation in human T-cell lymphotropic virus type 1 (HTLV-I)-transformed cells. In this study, we demonstrate that miR-150 and miR-223 specifically target the signal transducer and activator of transcription 1 (STAT1) 3' untranslated region, reducing STAT1 expression and dampening STAT1-dependent signaling in human T cells. The effects of miR-150 and miR-223 on endogenous STAT1 were confirmed using inducible cell lines. Our studies also showed that miR-150 expression is upregulated by interleukin-2 signaling in adult T cell leukemia/lymphoma (ATL) cells. HTLV-I-transformed and ATL-derived cells have reduced levels of miR150 and miR223 expression, which coincide with increased STAT1 expression and STAT1-dependent signaling. Knockdown of STAT1 by short hairpin RNA demonstrated that the constitutive activation of STAT1 is required for the continuous proliferation of HTLV-I-transformed cells. Our studies further demonstrate that increased expression of STAT1 in ATL cells is associated with higher levels of major histocompatibility complex class I expression. Previous studies have demonstrated that the pressure exerted by natural killer (NK) cells in vivo can edit leukemic tumor cells by forcing an increased expression of major histocompatibility complex class I to escape immune clearance. STAT1-expressing tumor cells produce more aggressive tumors because they cannot be eliminated by NK cells. Our results suggest that therapeutic approaches using combined targeting of STAT1 and MHC class I may be an effective approach to activate NK cell-mediated clearance of ATL tumor cells.

Tax impairs DNA replication forks and increases DNA breaks in specific oncogenic genome regions.

BACKGROUND: Human T-cell leukemia virus type 1 (HTLV-I) is a human retrovirus associated with adult T-cell leukemia (ATL), an aggressive CD4 T-cell proliferative disease with dismal prognosis. The long latency preceding the development of the disease and the low incidence suggests that the virus itself is not sufficient for transformation and that genetic defects are required to create a permissive environment for leukemia. In fact, ATL cells are characterized by profound genetic modifications including structural and numerical chromosome alterations. RESULTS: In this study we used molecular combing techniques to study the effect of the oncoprotein Tax on DNA replication. We found that replication forks have difficulties replicating complex DNA, fork progression is slower, and they pause or stall more frequently in the presence of Tax expression. Our results also show that Tax-associated replication defects are partially compensated by an increase in the firing of back-up origins. Consistent with these effects of Tax on DNA replication, an increase in double strand DNA breaks (DDSB) was seen in Tax expressing cells. Tax-mediated increases in DDSBs were associated with the ability of Tax to activate NF-kB and to stimulate intracellular nitric oxide production. We also demonstrated a reduced expression of human translesion synthesis (TLS) DNA polymerases Pol-H and Pol-K in HTLV-I-transformed T cells and ATL cells. This was associated with an increase in DNA breaks induced by Tax at specific genome regions, such as the c-Myc and the Bcl-2 major breakpoints. Consistent with the notion that the non-homologous end joining (NHEJ) pathway is hyperactive in HTLV-I-transformed cells, we found that inhibition of the NHEJ pathway induces significant killing of HTLV-I transformed cells and patient-derived leukemic ATL cells. CONCLUSION: Our results suggest that, replication problems increase genetic instability in HTLV-I-transformed cells. As a result, abuse of NHEJ and a defective homologous repair (HR) DNA repair pathway can be targeted as a new therapeutic approach for the treatment of adult T-cell leukemia.

Adult T-cell leukemia cells overexpress Wnt5a and promote osteoclast differentiation.

Adult T-cell leukemia/lymphoma (ATL) is etiologically linked to infection with the human T-cell leukemia/lymphoma virus type 1 (HTLV-I). ATL is classified into 4 distinct clinical diseases: acute, lymphoma, chronic, and smoldering. Acute ATL is the most aggressive form, representing 60% of cases and has a 4-year survival of < 5%. A frequent complication and cause of death in acute ATL patients is the presence of lytic bone lesions and hypercalcemia. We analyzed the Wnt/ß-catenin pathway because of its common role in cancer and bone remodeling. Our study demonstrated that ATL cells do not express high levels of ß-catenin but displayed high levels of LEF-1/TCF genes along with elevated levels of ß-catenin (LEF-1/TCF target genes) responsive genes. By profiling Wnt gene expression, we discovered that ATL patient leukemia cells shifted expression toward the noncanonical Wnt pathway. Interestingly, ATL cells overexpressed the osteolytic-associated genes-Wnt5a, PTHLH, and RANKL. We further show that Wnt5a secreted by ATL cells favors osteoclast differentiation and expression of RANK. Our results suggest that Wnt5a is a major contributing factor to the increase in osteolytic bone lesions and hypercalcemia found in ATL patients. Anti-Wnt5a therapy may prevent or reduce osteolytic lesions found in ATL patients and improve therapy outcome.

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

UNLABELLED: The 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-1­induced 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. KEY POINTS: PA28γ acts as a co-repressor of HTLV-1 p30 to suppress virus replication and is required for the maintenance of viral latency. HTLV-1 has evolved a unique function mediated by its posttranscriptional repressor p30, which is not found in HTLV-2.

Tumor suppressors p53, p63TAα, p63TAy, p73α, and p73ß use distinct pathways to repress telomerase expression.

The promoter of the telomerase catalytic subunit (TERT) is subject to tight regulation and remains repressed in somatic cells to ensure their limited life span and to prevent tumor initiation. Here we report that the hTERT promoter is strongly repressed by p53 and the related family members p63 and p73. We found that p53-mediated repression was different in human and mouse cells and occurred through p53-dependent transcription inhibition of c-Myc or through E-box/E2F pathways, respectively. Although p63TAα-mediated repression occurred through SP1, p63TAy-mediated repression occurred through E2F signaling. Finally, p73α- and p73ß-mediated repression occurred through NF-YB2. Our results show a complex multifactorial mechanism used by p53 and its family members to keep hTERT expression under tight control.