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Sasada, Amane. "APOBEC3G targets human T-cell leukemia virus type 1." Kyoto University, 2006. http://hdl.handle.net/2433/143870.
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Ruggero, Katia. "Role of microRNAs in T-cell activation and transformation by human T-cell Leukemia virus type 1." Doctoral thesis, Università degli studi di Padova, 2012. http://hdl.handle.net/11577/3422191.
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Human T-Lymphotropic virus type 1 (HTLV-1) is the causative agent of two distinct pathologies, adult T-cell leukemia/lymphoma (ATLL), an aggressive malignancy of mature CD4+ T-cells, and tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM), a demyelinating neurodegenerative disease. Despite intense study, many aspects of HTLV-1 replication, persistence and pathogenesis remain to be understood. The work described in the present thesis was aimed at defining the role of microRNAs (miRNAs) in HTLV-1 infection and ATLL pathogenesis.
We generated small RNA libraries from normal CD4+ cells (resting and stimulated) and two T-cell lines chronically infected with HTLV-1 (MT-2 and C91PL). Libraries were analyzed by 454 mass sequencing and data were processed through a series of computational steps to identify known and candidate new miRNAs for each library. Comparison of frequencies of known miRNAs in the different libraries led to the identification of 14 downregulated miRNAs and 4 upregulated species in infected cell lines vs. resting CD4+ cells, while 21 miRNAs were differentially expressed (16 downregulated, 5 upregulated) in stimulated compared to resting CD4+ cells. We validated the expression of some new miRNA candidates identified by bioinformatic analysis of the libraries through end point and quantitative RT-PCR. Two sequences mapped to the HTLV-1 genome, suggesting that the virus may produce its own miRNAs under certain conditions.
We examined the profiles of known miRNA expression in ATLL cells and normal resting and activated T CD4+ lymphocytes using microarrays. On the basis of miRNA expression, cluster analysis of ATLL samples and CD4+ controls showed that the resting controls were highly related to each other, while the tumor samples exhibited some heterogeneity. Statistical analysis revealed 6 upregulated and 21 downregulated miRNAs in ATLL cells compared to CD4+ T-cell controls. Several of the differentially expressed miRNAs identified in the libraries and by microarray analysis were validated by real time RT-PCR.
Since miRNA-mRNA interactions often result in degradation of the target mRNA, integration of results from target prediction programs with expression profiles for miRNAs and mRNAs can aid in identifying genuine mRNA targets. This approach was applied to miRNA and mRNA microarray data obtained for our ATLL and resting CD4+ samples. Potential targets for 12 miRNAs differentially expressed in ATLL cells were identified by integrating miRNA and mRNA expression profiles. Functional enrichment analysis of predicted targets revealed the presence of several genes belonging to the cAMP signalling pathway, which is known to be activated upon HTLV-1 transformation. We also investigated the role of miR-34a, consistently upregulated in ATLL samples and HTLV-1 infected cells lines. Knockdown of miR-34a in infected cell lines determined an increased in cell death, suggesting that miR-34a could play an important role in the expansion of HTLV-1 infected cells and thereby in ATLL development.Il virus T-linfotropico umano di tipo 1 (HTLV-1) è l’agente eziologico della leucemia/linfoma a cellule T dell’adulto (ATLL, adult T-cell leukemia/lymphoma) e della paraparesi spastica tropicale/mielopatia associata ad HTLV (TSP/HAM, Tropical spastic paraparesis/HTLV-associated myelopathy), una patologia degenerativa del sistema nervoso centrale. Recenti evidenze suggeriscono che i microRNA (miRNA) contribuiscano a questo processo di trasformazione mediata da HTLV-1. Le ricerche condotte nel corso del mio dottorato sono state mirate ad approfondire il ruolo dei microRNA (miRNA) nell’infezione di cellule T da parte di HTLV-1 e nella patogenesi dell’ATLL. Sono state realizzate librerie di cDNA di piccoli RNA, a partire da linfociti T CD4+ normali (resting e attivati) e da due linee cellulari cronicamente infettate con HTLV-1 (C91PL e MT-2). Le librerie sono state analizzate attraverso il sequenziamento di massa 454 e l’analisi bioinformatica delle sequenze ottenute ha permesso l’identificazione dei miRNA noti e nuovi miRNA candidati presenti in ciascuna libreria. Il confronto delle frequenze dei miRNA noti nelle diverse librerie ha evidenziato la presenza di 14 e 4 miRNA rispettivamente downregolati e upregolati nelle linee cellulari infettare rispetto ai linofociti T CD4+ resting, mentre 21 miRNA sono risultati differenzialmente espressi in linfociti T CD4+ stimolati in confronto ai linfociti T CD4+ resting (16 downregolati, 5 upregolati). L’espressione di diversi nuovi miRNA, individuati dall’analisi bioinformatica delle librerie, è stata validata attraverso RT-PCR end-point o RT-PCR quantitativa. Inoltre la nostra analisi ha rivelato nelle librerie da cellule infettate 2 sequenze che mappano in regioni trascritte del genoma di HTLV-1 e che potrebbero rappresentare dei miRNA virali. Attraverso l’impiego di microarray il profilo di espressione dei miRNA noti è stato analizzato in pazienti ATLL e in linfociti T CD4+ resting e stimolati. In base ai profili di espressione di miRNA ottenuti i campioni sono stati raggruppati in cluster che indicano una forte similitudine all’interno dei campioni di linfocititi T CD4+ resting, mentre i campioni di ATLL hanno profili di espressione di miRNA più eterogenei. L’analisi statistica ha evidenziato 21 miRNA downregolati e 6 upregolati nei pazienti ATLL vs linfociti T CD4+ resting. Diversi miRNA differenzialmente espressi identificati attraverso l’analisi delle librerie e dei microarray sono stati validati tramite RT-PCR quantitativa. Dal momento che l’interazione miRNA-mRNA spesso comporta la degradazione del messaggero bersaglio, l’analisi integrata dei risultati dei programmi di predizione di bersagli con i profili di espressione di miRNA e geni può aiutare nell’identificazione di target. Abbiamo applicato questo approccio ai dati di espressione di miRNA e geni ottenuti per i nostri campioni di ATLL e linfociti T CD4+ resting. Dall’integrazione dei profili di espressione di miRNA e mRNA sono stati identificati i target putativi per 12 miRNA differenzialmente espressi nei pazienti ATLL. L’arricchimento funzionale dei geni bersaglio predetti ha evidenziato la presenza di diversi geni coinvolti nella via di segnale di cAMP, noto per essere presente ad alti livelli in cellule trasformate da HTLV-1. Infine abbiamo indagato il significato funzionale di miR-34a, che risulta essere consistentemente upregolato in pazienti ATLL e linee cellulari infettate. Il silenziamento di miR-34a in linee cellulari infettate determina un aumento della morte cellulare, suggerendo che la deregolazione di questo miRNA possa svolgere un ruolo importante nell’espansione della popolazione di cellule infettate da HTLV-1 e quindi nello sviluppo dell’ATLL.
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Furuta, Rie. "Human T-cell leukemia virus type 1 infects multiple lineage hematopoietic cells in vivo." Kyoto University, 2018. http://hdl.handle.net/2433/232110.
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Younis, Ihab H. "Molecular analysis of human t-cell leukemia virus regulatory and accessory proteins." The Ohio State University, 2005. http://rave.ohiolink.edu/etdc/view?acc_num=osu1123168747.
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Li, Min. "Kinetic analysis of Human T-cell leukemia virus type 1 gene expression." The Ohio State University, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=osu1228156327.
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Miura, Michi. "Characterization of simian T-cell leukemia virus type 1 in naturally infected Japanese macaques as a model of HTLV-1 infection." Kyoto University, 2014. http://hdl.handle.net/2433/188641.
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Yamamoto, Brenda Michiyo. "Molecular Analysis of Human T-cell Leukemia Virus Type 2 Accessory Protein p28." The Ohio State University, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=osu1241708950.
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Doueiri, Rami. "CHARACTERIZATION OF THE HUMAN T-CELL LEUKEMIA VIRUS TYPE-2 P28 ACCESSORY PROTEIN." The Ohio State University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=osu1343453789.
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Newbound, Garret C. "Transcriptional control of human t-cell leukemia virus type-1 in primary lymphocytes /." The Ohio State University, 1997. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487948440826361.
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Anderson, Matthew David. "Studies with the human t-cell leukemia virus tax and rex positive trans-regulatory proteins." Connect to this title online, 2004. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1083092375.
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Thesis (Ph. D.)--Ohio State University, 2004.Title from first page of PDF file. Document formatted into pages; contains xv, 129 p.; also includes graphics Includes bibliographical references (p. 105-129). Available online via OhioLINK's ETD Center
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Ding, Yan Shirley. "Expression, purification and charaterization of recombinant human T-cell leukemia virus type I protease." Diss., Georgia Institute of Technology, 1998. http://hdl.handle.net/1853/29992.
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Hiraragi, Hajime. "Study of lentiviral vector for in utero gene transfer and functional analysis of human T-lymphotropic virus type p13(II)." Connect to resource, 2005. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1116532636.
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Thesis (Ph. D.)--Ohio State University, 2005.Title from first page of PDF file. Document formatted into pages; contains xvii, 230 p.; also includes graphics. Includes bibliographical references (p. 200-230). Available online via OhioLINK's ETD Center
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Dennison, Kelly J. "Development of a structural model of human T-cell leukemia virus type-I protease." Thesis, Georgia Institute of Technology, 2002. http://hdl.handle.net/1853/30060.
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Manicone, Mariangela. "Functional interactions of the Tax and p13 proteins of Human T-cell Leukemia Virus Type I." Doctoral thesis, Università degli studi di Padova, 2013. http://hdl.handle.net/11577/3422623.
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Human T-cell leukemia virus type 1 (HTLV-1) establishes a lifelong persistent infection in humans. Approximately 3% of the infected individuals will develop adult T-cell leukemia/lymphoma (ATLL), an aggressive malignancy of mature CD4+ T-cells. The viral protein Tax plays a major role in HTLV-1 pathogenicity by activating the NF-κB pathway. Tax activates both the canonical and non-canonical NF-κB pathways, promoting NF-κB translocation to the nucleus and transcription of genes that favour T-cell proliferation and survival. Our previous studies showed that the p13 protein of HTLV-1 enhances mitochondrial ROS production, resulting in activation of normal T-cells. ROS constitute a homeostatic rheostat that controls the activity of several key pathways, including the NF-κB pathway.Thus, we hypothesized that the effects of p13 on ROS production could affect the activation of the NF-κB pathway by Tax in primary T-cells.
The work described in the present thesis was aimed at testing the hypothesis that Tax and p13 might act in concert to activate the NF-κB signal transduction pathway in primary T-cells. To this end, we optimized a transfection protocol for primary T-cells using an innovative approach based on the electroporation of in vitro-transcribed RNA. Activation of the NF-κB pathway was then analysed by measuring expression of the NF-κB target genes CD25 and 4-1BB.
Results showed that the co-transfection of Tax and p13 resulted in a synergistic activation of the NF-κB pathway in primary T-cells measured as an increase in the expression levels of both CD25 and 4-1BB. In addition to being a transcriptional target of NF-κB, CD25 is also an early marker of T-cell activation. To further test the effects of Tax and p13 on cell activation, we measured CD38 expression by flow cytometry. Jurkat T-cells, which exhibit a constitutively activated CD38 positive phenotype, were used as a control. Results of this analysis confirmed the synergy of Tax and p13, although the effect was not so prominent as that observed for the CD25 marker, suggesting that, within the time frame of our experiments, Tax and p13 drove T-cells to an early-intermediate stage of activation.
Taken together, these findings suggest that, in contrast to the well-established role of Tax as an activator of the NF-κB pathway in tumor cell lines, in the context of normal T-cells, the induction of NF-κB target genes requires the concerted action of Tax and p13.
Current studies are aimed at verifying the ROS-dependence of this effect and testing the functional interaction of Tax and p13 in the context of the complete HTLV-1 genome using wild type HTLV-1 and a p13-knock-out HTLV-1 molecular clone. These experiments will be carried out in primary T-cells as well as in dendritic cells, which have recently emerged as an important target of the virus in vivo.Il virus T-linfotropico umano di tipo 1 (HTLV-1) stabilisce un'infezione persistente negli uomini. Circa il 3% degli individui infettati sviluppa la leucemia/linfoma a cellule T dell'adulto (ATLL), un'aggressiva neoplasia a carico dei linfociti T CD4+ maturi. L'attivazione della via di segnale di NF-κB mediata dalla proteina virale Tax è un evento cruciale nella patogenesi dell’infezione da HTLV-1. Tax attiva entrambe le vie di segnale di NF-κB, canonica e non-canonica, promuovendo la traslocazione nucleare di NF-κB e la trascrizione di geni che favoriscono la proliferazione e la sopravvivenza delle cellule T. I nostri studi precedenti hanno rivelato che la proteina virale p13 favorisce la produzione di specie reattive dell'ossigeno (ROS) a livello mitocondriale, causando l'attivazione di cellule T normali. I ROS possono essere paragonati ad un reostato che controlla l'attività di diverse vie di trasduzione del segnale, inclusa la via di NF-κB. Lo scopo primario di questa tesi è stato quindi di verificare l'ipotesi che Tax e p13 potessero attivare sinergicamente la via di trasduzione del segnale di NF-κB in cellule T normali. A tal fine, è stato ottimizzato un protocollo di trasfezione di cellule T primarie utilizzando un approccio innovativo basato sull'elettroporazione di RNA trascritto in vitro. L'attivazione della via di NF-κB è stata analizzata misurando l'espressione dei geni target di NF-κB CD25, mediante analisi citofluorimetrica, e 4-1BB, mediante RT-PCR quantitativa. I risultati ottenuti hanno mostrato che in cellule T normali, la co-trasfezione di Tax e p13 causa l'attivazione sinergica della via di NF-κB misurata come incremento dei livelli di espressione di entrambi i geni target. Oltre ad essere un target trascrizionale di NF-κB, il CD25 è anche un marcatore precoce di attivazione di cellule T. Per verificare il possibile effetto di Tax e p13 sull'attivazione cellulare, abbiamo misurato mediante analisi citofluorimetrica l'espressione del CD38, un marcatore intermedio-tardivo di attivazione. La linea T-cellulare leucemica Jurkat, caratterizzata da un fenotipo costitutivamente CD38 positivo, è stata utilizzata come controllo. I risultati di questa analisi hanno confermato la sinergia di Tax e p13, nonostante l'effetto sull’espressione del CD38 non fosse così prominente come quello osservato per il CD25, suggerendo che, nel nostro contesto sperimentale, Tax e p13 spingano le cellule T in uno stadio precoce-intermedio di attivazione. In complesso questi risultati suggeriscono che, in contrasto con il ruolo ben stabilito di Tax nell'attivazione della via di NF-κB in linee cellulari tumorali, nel contesto delle cellule T normali, l'induzione dei geni target di NF-κB necessita l'azione sinergica di Tax e p13. Gli studi attualmente in corso sono volti a verificare la ROS-dipendenza dell'effetto sinergico di Tax e p13 sulla via di segnale di NF-κB. Inoltre, verificheremo la validità dell'interazione funzionale di Tax e p13 nel contesto del intero genoma di HTLV-1. A tal fine, paragoneremo l'attivazione della via di NF-κB indotta da un clone molecolare di HTLV-1 wild type, con quella indotta da un clone molecolare di HTLV-1 p13-knock-out. Questi esperimenti verranno condotti in cellule T primarie ed in cellule dendritiche, che rappresentano il principale target infezione da HTLV-1 in vivo.
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Gao, Weiwei, and 高蔚为. "Salt-inducible kinases function as a host restriction to human T-cell leukemia virus type 1 transcription." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2012. http://hub.hku.hk/bib/B4818309X.
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Human T-cell leukemia virus type 1 (HTLV-1) is the causative agent of adult T-cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). HTLV-1 Tax is the major viral transactivator and transforming protein centrally involved in the proviral transcription, transformation and proliferation of infected T-cells as well as progression of diseases caused by HTLV-1 infection. Salt-inducible kinases (SIKs) are serine/threonine protein kinases belonging to the AMPK-related kinase (AMPK-RK) family. SIK subfamily consists of three isoforms named SIK1, SIK2 and SIK3 respectively. We have previously demonstrated the negative regulatory role of SIK1 in Tax-mediated activation of proviral transcription from long terminal repeats (LTR). In this study, we reported that both SIK2 and SIK3 exhibited a kinase-dependent suppressive effect on Tax-activated LTR transcription. We also found that SIK1, SIK2 and SIK3 act additively to suppress Tax activation of LTR. We further demonstrated that the SIK2- and SIK3-mediated suppression on LTR transcription was achieved through phosphorylation of TORC1, an essential transcriptional coactivator of CREB required for Tax-mediated transcriptional activation of LTR. Our findings revealed a new function of SIK2 and SIK3 in host restriction to HTLV-1 transcription. Pharmaceutical activation of SIKs or upstream kinase such as LKB1 may provide a new strategy for anti-HTLV-1 therapy.published_or_final_version
Biochemistry
Master
Master of Medical Sciences
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Pais, Correia Ana Monica. "Biofilm-like extracellular viral assemblies mediate HTLV-1 (human T cell leukemia virus type-1) cell-to-cell transmission at virological synapses." Paris 7, 2009. http://www.theses.fr/2009PA077233.
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Près de 20 millions d'individus sont infectés par HTLV-1 (human T cell leukemia virus type-1). La plupart des patients n'ont aucun symptôme, seulement 5-10% d'entre eux développeront des maladies associées à l'infection virale. La particularité d'HTLV-1 est de se transmettre par des contacts cellulaires désignés « synapses virologiques », qui partagent certaines caractéristiques avec les synapses immunologiques. Ce projet a permis de mieux comprendre le processus de transmission cellule à cellule et introduit un nouveau concept de transmission virale par des « biofilms viraux ». Contrairement à ce qui avait été suggéré, nous démontrons qu'HTLV-1 s'accumule à la surface des cellules infectées, dans des structures composées de matrice extracellulaire (HSPG agrine, collagène) et de molécules de type « linker » (galectine, tetherine). Ces structures facilitent l'adhésion et la cohésion du virus et sont essentielles pour une transmission virale efficace, car leur détachement diminue fortement l'infection de cellules cibles («80%). Les structures que nous avons décrites concentrent et protègent le virus, permettent son stockage et aident à sa propagation. Ces caractéristiques ressemblent à celles des biofilms bactériens ou des champignons. Ainsi, nous proposons le nom « biofilm viral» pour désigner ce nouveau processus de transmission viraleApproximately 20 million people are infected by HTLV-1 (human T cell leukemia virus type-1). Although the majority of infected individuals remain asymptomatic, around 10% will develop HTLV-1-associated disease. HTLV-1 has been described to efficiently propagate through cell-cell contacts. Designated as virological synapses, these cellular contacts exhibit some of the characteristics of immunological synapses. This project has allowed a better comprehension of how HTLV-1 propagates from cell-to-cell and proposes a completely novel concept of virus transmission through 'viral biofilms'. Contrary to what was previously suggested we have shown that HTLV-1 is accumulated at the surface of infected cells, in extracellular viral assemblies. Viral assemblies are composed of extracellular matrix (HSPG agrin, collagen) and linker molecules (galectin, tetherin) that provide the adhesion and cohesion forces needed to maintain viruses at the cell surface. More importantly, we have shown that HTLV- 1 viral assemblies contribute to the majority of virus cell-cell transmission since their removal significantly inhibits infection (« 80%). Viral assemblies, store, concentrate, disseminate and protect the virus. These characteristics recapitulate most of the features of bacterial or fungal biofilms. Therefore, we propose the use of the term 'viral biofilm' to designate this new mode of cell-to-cell virus spread
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Ye, Jianxin. "Transformation studies of human t-cell leukemia virus with emplhasis on the role of tax and rex." The Ohio State University, 2003. http://rave.ohiolink.edu/etdc/view?acc_num=osu1060451751.
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Zhao, Tiejun. "Human T-cell leukemia virus type 1 bZIP factor selectively suppresses the classical pathway of NF-κB." Kyoto University, 2010. http://hdl.handle.net/2433/120547.
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Je, Jianxin. "Transformation studies of human t-cell leukemia virus with emphasis on the role of tax and rex." Columbus, Ohio : Ohio State University, 2003. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1060451751.
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Thesis (Ph. D.)--Ohio State University, 2003.Title from first page of PDF file. Document formatted into pages; contains xii, 133 p.; also includes graphics. Includes abstract and vita. Advisor:, Dept. of Molecular, Cellular, and Developmental Biology. Includes bibliographical references (p. 108-133).
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Miyazato, Paola. "De novo human T-cell leukemia virus type 1 infection of human lymphocytes in NOD-SCID, common γ-chain knockout mice." Kyoto University, 2007. http://hdl.handle.net/2433/135660.
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Mitagami, Yu. "Interferon-γ promotes inflammation and development of T-cell lymphoma in HTLV-1 bZIP factor transgenic mice." Kyoto University, 2016. http://hdl.handle.net/2433/215454.
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Niyogi, Kakoli. "Lipid rafts, exosomes, and human T-cell leukemia virus type 1 biology a new model of viral pathogenesis /." Available to US Hopkins community, 2003. http://wwwlib.umi.com/dissertations/dlnow/3080734.
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Shu, Sherry T. "Pathogenesis and Treatments of Humoral Hypercalcemia of Malignancy in Adult T-Cell Leukemia/Lymphoma Induced by Human T Lymphotropic Virus Type 1." The Ohio State University, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=osu1245283708.
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Adya, Neeraj. "Mechanism of human T cell leukemia virus type-I gene (HTLV-I) regulation as mediated by regulatory protein, Tax." Case Western Reserve University School of Graduate Studies / OhioLINK, 1994. http://rave.ohiolink.edu/etdc/view?acc_num=case1057765609.
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Komurian-Pradel, Florence. "Variabilité génomique du virus HTLV-I (Human T-cell Leukemia Virus type I) en fonction de la géographie et des pathologies associées." Lyon 1, 1992. http://www.theses.fr/1992LYO1T001.
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Oliere, Stéphanie. "Modulation of the innate immune response during Human T-cell Leukemia Virus infection: implication for development of an oncolytic vector." Thesis, McGill University, 2011. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=96869.
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Infection with human T cell Leukemia virus (HTLV-1) can cause Adult T-cell Leukemia (ATL) or 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. The factors that determine HTLV-1 pathogenesis remain elusive, and therefore represent a serious obstacle in the establishment of effective therapies for HTLV-1-associated diseases. Using gene expression profiling of CD4+ T-lymphocytes isolated from HTLV-1-infected individuals, we identified candidate genes differentially regulated in HTLV-1-associated diseases. Of particular interest, SOCS1 was up-regulated in HAM/TSP and AC patients – but not in ATL. SOCS1 positively correlated with HTLV-1 mRNA in HAM/TSP patient samples. SOCS1-mediated degradation of IRF3 - inhibited antiviral signaling during HTLV-1 infection. Our study reveals a novel evasion mechanism utilized by HTLV-1 which leads to increased retroviral replication, without triggering an IRF3-dependent interferon response. Thus, targeting SOCS1 could represent a potential new approach to enhance the therapeutic potency of IFN-α/β treatment in HAM/TSP disease. Although treatment of hematological malignancies has improved considerably, this has not benefited ATL patients, as they are completely refractory to conventional chemotherapeutic regimens. Oncolytic viruses, such as Vesicular stomatitis virus (VSV), have emerged as a potential treatment for cancer. Here we show that in vitro VSV infection induced significant oncolysis in highly proliferating primary ATL cells, but not in primary Chronic Lymphocytic Leukemia (CLL) cells which are arrested in the G0 phase. As chronic activation and proliferation is characteristic of ATL cells, we examined the effect of T-cell activation on VSV permissiveness and lysis. Activation of primary CD4+ T-lymphocytes was sufficient to induce VSV replication and VSV-triggered cell death, suggesting that cellular signaling pathways - ERK, JNK or AKT - that promote VSV replication are engaged during T-cell activation. Similarly, mitogenic activation of primary CLL promotes the exit from G0 and entrance into the cell cycle, rendering them susceptible to VSV-mediated oncolysis. Moreover, a global increase in protein translation mediated by the activation of mTOR and eIF4E was crucial for VSV replication in primary lymphocytes. These findings provide novel molecular targets for ATL and CLL therapeutics.Le rétrovirus T-lymphotropique humain (HTLV-1) est l'agent étiologique de la leucémie à cellule T de l'adulte (ATL) - une leucémie agressive et fatale des lymphocytes T CD4+. HTLV-1 est également associé à une forme de myélopathie chronique appelée Paraparésie Spastique Tropicale ou atteinte neurologique connue sous le nom de HTLV-1 - Associated Myelopathy (HAM/TSP). Bien que la majorité des individus infectés avec HTLV-1 demeurent asymptomatiques (AC) au cours de leur vie, 2 à 5% développent soit une ATL soit une HAM/TSP. Les facteurs qui déterminent la pathogénèse de l'HTLV-1 restent inconnus, et représentent donc un sérieux obstacle à la mise en place de traitements efficaces contre les maladies associées au virus HTLV-1. Les études sur l'expression des gènes des lymphocytes T CD4+ isolés de patients infectés par HTLV-1, ont permis l'identification de gènes d'intérêt exprimés de façon différentielle dans les maladies associées au virus HTLV-1. De façon intéressante, il a été mis en évidence que l'expression de SOCS1 est plus élevé chez les patients asymptomatiques ou HAM/TSP que chez les patients ATL qui expriment généralement très peu d'ARN viral. Chez les patients HAM, il existe une corrélation directe entre le niveau d'expression de SOCS1 et l'activité transcriptionelle provirale. Du point de vue fonctionnel, SOCS1 inhibe la réponse antivirale, entre autre via la dégradation du facteur de transcription IRF3. Cette étude a donc permis d'identifier un nouveau mécanisme utilisé par HTLV-1 afin d'inhiber la réponse antivirale et ainsi d'augmenter sa capacité de réplication.Dans cette étude, nous démontrons également que l'infection in vitro par le VSV induit la lyse oncogénique des cellules ATL, à fort potentiel prolifératif, mais pas celle des cellules primaires de la leucémie lymphocytique chronique (CLL), qui sont quiescentes in vitro. Puisque l'activation et la prolifération chronique est une caractéristique des cellules ATL, nous avons étudié l'effet de l'activation des cellules T sur leur permissivité au VSV ainsi que leur lyse induite par le virus. L'activation des cellules T CD4+ primaires de patients sains est suffisante pour permettre la réplication virale et la mort cellulaire induite par le VSV. L'utilisation d'inhibiteurs de la signalisation cellulaire a montré que l'activation de ERK, JNK ou AKT est suffisante pour permettre la réplication de VSV dans les cellules T CD4+. De façon similaire, la stimulation mitogénique des cellules CLL de patients induisant leur entrée dans le cycle cellulaire, les rend susceptibles à l'oncolyse par le VSV. De plus, une augmentation globale de la traduction protéique induite par l'activation de mTOR et eIF4E est essentielle pour la réplication du VSV dans les lymphocytes primaires.En conclusion, ces résultats permettent d'identifier de nouvelles voies thérapeutiques pour les leucémies de l'ATL et CLL.
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Sharma, Varun Kumar. "The role of small non-coding RNAs in human T-cell leukemia virus type 1 (HTLV-1) infection and transformation." Doctoral thesis, Università degli studi di Padova, 2014. http://hdl.handle.net/11577/3423709.
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Human T-cell leukemia virus type 1 (HTLV-1) is the causative agent of two distinct pathologies, adult T-cell leukemia/lymphoma (ATLL), an aggressive neoplasm of mature CD4+ T-cells, and tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM), a demyelinating neurodegenerative disease. The emerging importance of small noncoding RNAs in normal cell physiology and disease has prompted studies of their role in T-cell activation and transformation. The work described in the present thesis was aimed at understanding the role of small noncoding RNAs, in particular microRNAs and tRNA fragments (tRFs), in HTLV-1 infection and ATLL pathogenesis.
The laboratory generated small RNA libraries to identify the repertoire of small noncoding RNAs expressed in two HTLV-1-infected T-cell lines (C91PL and MT-2) compared to normal CD4+ T-cells. Results revealed upregulation of miR-34a in the cell lines. Many tRFs were identified in both uninfected and infected cells. One of the most abundant tRFs (tRF-3019) was derived from the 3’ end of tRNA-proline, which is considered to be the primer for HTLV-1 reverse transcriptase. Results of an in vitro reverse transcriptase assay verified that tRF-3019 was capable of priming HTLV-1 reverse transcriptase. Both tRNA-proline and tRF-3019 were detected in HTLV-1 virus particles. tRF-3019 may thus play an important role in HTLV-1 reverse transcription and could represent a target to control HTLV-1 infection.
Data from a microarray-based analysis of microRNA expression in ATLL samples compared to normal CD4+ T-cells revealed 21 downregulated microRNAs and 6 upregulated microRNAs. Upregulated microRNAs included miR-34a, which is a member of the highly conserved miR-34 family that acts as a tumor suppressor induced by p53 in other cell types. However, p53 is known to be functionally inactivated or mutated in ATLL cells and HTLV-1-infected cell lines. Treatment of infected cell lines with nutlin-3a, a drug that restores p53 activity by interfering with MDM2, resulted in an upregulation of miR-34a and strong downregulation of several of its predicted targets. These findings indicate that unblocking the p53 pathway in HTLV-1-infected cells promotes engagement of the miR-34a/mRNA regulatory network.
The final aim of the project was to identify microRNAs regulated by the viral regulatory protein Tax. To this end the HTLV-1-negative T-cell line Jurkat was transfected with a Tax expression plasmid and assayed for changes in mRNA and microRNA expression by quantitative RT-PCR. Results revealed significant alterations in the levels of 7 microRNAs in the presence of Tax. These included let-7g, whose levels were reduced in the Tax-expressing cells. Let-7g was also found to be downregulated in ATLL samples compared to normal CD4 cells analysed by microarrays, suggesting that this microRNA might play a tumor suppressor role in HTLV-1-mediated transformation. Experiments are currently underway to identify targets of let-7g in infected cells using as a starting point 14 genes identified by integrating results from microRNA target prediction programs with expression profiles for microRNAs and mRNAs in ATLL cells vs. CD4 controls.Il virus T-linfotropico umano di tipo 1 (HTLV-1) è l’agente eziologico della leucemia/linfoma a cellule T dell’adulto (ATLL, Adult T-cell leukemia/lymphoma), un’aggressiva neoplasia a carico dei linfociti T CD4+ maturi, e della paraparesi spastica tropicale/mielopatia associata ad HTLV (TSP/HAM, Tropical spastic paraparesis/HTLV-associated myelopathy), una patologia degenerativa del sistema nervoso centrale. L’interesse crescente nello studio e nella comprensione della funzione degli “small non-coding RNA” in cellule normali e tumorali ci ha spinto ad uno studio del loro ruolo nell’ attivazione e nella trasformazione delle cellule T. Il lavoro descritto nella presente tesi mira a comprendere il ruolo degli “small non-coding RNA” (sncRNA), in particolare microRNA e frammenti tRNA (tRFs), nell’ infezione da HTLV-1 e nella patogenesi dell’ATLL. Nel nostro laboratorio sono state generate librerie di “small RNA” per identificare il repertorio di sncRNA espressi in due linee cellulari infettate con HTLV-1 (C91PL e MT-2) rispetto alle cellule T CD4 + normali. I risultati hanno rivelato un’aumentata espressione del miR-34a nelle linee cellulari infettate. Molti frammenti di tRNA (tRFs) sono stati identificati sia nelle cellule infettate che non infettate. Uno dei tRFs più abbondanti (tRF-3019) è derivato dall’ estremità 3’ del tRNA-prolina, che è considerato il primer per la trascrittasi inversa dell’HTLV-1. I risultati ottenuti da un saggio di trascrittasi inversa in vitro hanno dimostrato che il tRF-3019 è in grado di funzionare da primer nella trascrizione inversa di HTLV-1. La presenza sia del tRNA-prolina che del tRF-3019 è stata evidenziata nelle particelle virali. Il tRF-3019 potrebbe quindi svolgere un ruolo importante nella retrotrascrizione del virus e potrebbe rappresentare un “target” terapeutico nell’infezione da HTLV-1. I dati ottenuti dall’ analisi con microarray sull’ espressione di microRNA in campioni di ATLL e in campioni di cellule T-CD4 + normali ha rivelato una diminuzione nell’espressione di 21 microRNA e un’aumentata espressione di 6 microRNA. I microRNA sovraespressi comprendono anche il miR-34a, che è un membro della famiglia dei miR-34, altamente conservati, che agiscono come oncosoppressori indotti da p53 in diversi tipi cellulari. Tuttavia, p53 è inattiva o mutata in cellule ATLL e in linee cellulari HTLV-1-infettate. Il trattamento di linee cellulari infettate con Nutlin-3a, un farmaco che ripristina l'attività di p53 legandosi a MDM2, ha rivelato un aumeto di espressione di miR-34a e una forte riduzione dell’espressione di alcuni dei suoi target. Questi risultati suggeriscono che attivando il pathway di p53 in cellule HTLV-1-infettate si potrebbe promuovere l’ingaggio del network regolatorio del miR-34a. Infine, ci siamo proposti di identificare i microRNA regolati dalla proteina virale Tax. A tal fine la linea cellulare T non infetta, Jurkat, è stata transfettata con un plasmide di espressione per Tax e sono state testate le variazioni di espressione di mRNA e microRNA mediante RT-PCR. I risultati hanno rivelato che in presenza di Tax ci sono alterazioni significative nei livelli di espressione di 7 microRNA. Queste variazioni includono il microRNA let-7g, i cui livelli sono ridotti nelle cellule che esprimono Tax. Da studi effettuati su microrrays, let-7g risulta sottoespresso in campioni ATLL rispetto alle cellule CD4 normali, suggerendo che questo microRNA potrebbe svolgere un ruolo di oncosoppressore nella trasformazione mediata da HTLV-1. Gli esperimenti, attualmente in corso, permetteranno di identificare i target di let-7g in cellule infettate utilizzando come punto di partenza 14 geni ottenuti dall’integrazione dei risultati dei programmi di predizione dei target dei microRNA con i profili di espressione di microRNA e mRNA in cellule ATLL rispetto ai controlli CD4.
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Oliere, Stéphanie. "Modulation of the innate immune response during human T-cell leukemia virus infection implications for development of an oncolytic virotherapy for ATL." Thesis, McGill University, 2010. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=111920.
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Infection with human T cell Leukemia virus (HTLV-1) can cause Adult T-cell Leukemia (ATL) or 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. The factors that determine HTLV-1 pathogenesis remain elusive, and therefore represent a serious obstacle in the establishment of effective therapies for HTLV-1-associated diseases.Using gene expression profiling of CD4+ T-lymphocytes isolated from HTLV-1-infected individuals, we identified candidate genes differentially regulated in HTLV-1-associated diseases. Of particular interest, SOCS1 was up-regulated in HAM/TSP and AC patients -- but not in ATL. SOCS1 positively correlated with HTLV-1 mRNA in HAM/TSP patient samples. SOCS1-mediated degradation of IRF3 - inhibited antiviral signaling during HTLV-1 infection. Our study reveals a novel evasion mechanism utilized by HTLV-1 which leads to increased retroviral replication, without triggering an IRF3-dependent interferon response. Thus, targeting SOCS1 could represent a potential new approach to enhance the therapeutic potency of IFN-alpha/beta treatment in HAM/TSP disease.
Although treatment of hematological malignancies has improved considerably, this has not benefited ATL patients, as they are completely refractory to conventional chemotherapeutic regimens. Oncolytic viruses, such as Vesicular stomatitis virus (VSV), have emerged as a potential treatment for cancer. Here we show that in vitro VSV infection induced significant oncolysis in highly proliferating primary ATL cells, but not in primary Chronic Lymphocytic Leukemia (CLL) cells which are arrested in the G0 phase. As chronic activation and proliferation is characteristic of ATL cells, we examined the effect of T-cell activation on VSV permissiveness and lysis. Activation of primary CD4+ T-lymphocytes was sufficient to induce VSV replication and VSV-triggered cell death, suggesting that cellular signaling pathways - ERK, JNK or AKT - that promote VSV replication are engaged during T-cell activation. Similarly, mitogenic activation of primary CLL promotes the exit from G0 and entrance into the cell cycle, rendering them susceptible to VSV-mediated oncolysis. Moreover, a global increase in protein translation mediated by the activation of mTOR and eIF4E was crucial for VSV replication in primary lymphocytes. These findings provide novel molecular targets for ATL and CLL therapeutics.
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Shoji(kawata), Sanae. "p21Waf1/Cip1/Sdi1 functions to prevent apoptosis as well as stimulate growth in cells transformed or immortalized by human T-cell leukemia virus type 1-encoded Tax." Kyoto University, 2003. http://hdl.handle.net/2433/148476.
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Nasr, Al-Ghadban Rihab Raif. "Ciblage thérapeutique de Tax et de la voie NF-kB dans la leucémie T de l' adulte liée au rétrovirus HTLV-I." Paris 7, 2005. http://www.theses.fr/2005PA077074.
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Macaire, Héloïse. "Régulation de l’expression des protéines anti-apoptotiques Bfl-1 et Bcl-xL par les protéines virales Tax et HBZ du virus HTLV-1 et identification de petites molécules anti-Bfl-1 à visée thérapeutique." Thesis, Lyon 1, 2011. http://www.theses.fr/2011LYO10357/document.
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Le virus humain T lymphotrope de type 1 (HTLV-1) est l’agent étiologique de la leucémie/lymphome T de l’adulte (ATLL) qui se développe après plusieurs décennies et pour laquelle il n’existe à ce jour pas de traitement efficace. Parmi les protéines virales de HTLV-1, Tax et HBZ jouent un rôle déterminant dans le développement de l’ATLL. Si Tax participe au processus leucémogène dès les étapes précoces, HBZ jouerait plutôt un rôle dans le maintien du phénotype tumoral dans les étapes tardives. Dans ce contexte, là nous nous sommes intéressés à la régulation de l’expression des protéines anti-apoptotiques Bfl-1 et Bcl-xL, par les protéines virales Tax et HBZ. Nous avons montré que Tax induit l’expression des protéines anti-apoptotiques Bfl-1 et Bcl-xL de la famille Bcl-2 via la voie NF-κB, alors que HBZ n’a aucun effet sur leur expression. De plus, Tax coopère avec les facteurs de transcription c-Jun et JunD de la voie AP-1 pour augmenter l’expression de ces gènes anti-apoptotiques. En revanche, HBZ module uniquement la trans-activation de bfl-1 induite par Tax. L’ensemble de nos résultats indique donc que Tax joue un rôle prépondérant dans l’activation de l’expression de Bfl-1 et de Bcl-xL et suggère que Bfl-1 et Bcl-xL sont exprimées au cours des étapes précoces et tardives du développement de l’ATLL. Par une stratégie d’ARN interférence, nous avons ensuite montré que Bfl-1 et/ou Bcl-xL sont impliquées dans la survie de lignées cellulaires T infectées par HTLV-1, suggérant que Bfl-1 et Bcl-xL représentent des cibles thérapeutiques potentielles pour traiter l’ATLL. Actuellement, il existe des petites molécules ciblant les membres anti-apoptotiques de la famille Bcl-2, mais aucune ne cible spécifiquement Bfl-1. En collaboration avec la société IMAXIO, nous avons identifié par deux cribles à haut débit 83 molécules capables d’inhiber l’activité anti-apoptotique de Bfl-1. L’une de ces molécules induit spécifiquement la mort de lignées cellulaires T infectées par HTLV-1 pour lesquelles Bfl-1 représente un gène de survie. Ainsi, ce travail doit permettre à terme de développer de futurs médicaments dirigés contre Bfl-1 et de proposer une nouvelle stratégie thérapeutique ciblée contre l’ATLLHuman T lymphotropic virus type 1 (HTLV-1) is the etiological agent of adult T-cell leukemia/lymphoma (ATLL) that develops after several decades and for which there is no effective treatment. Among the viral proteins of HTLV-1, Tax and HBZ play a major role in the development of ATLL. If Tax participates in the initiation of leukemogenesis from the early stages, HBZ rather plays a role in maintaining the tumor phenotype in the late stages. The aims of our study were to better understand the regulation of Bfl-1 and Bcl-xL anti-apoptotic protein expression by Tax and HBZ viral proteins, as well as their role in the survival of HTLV-1-infected T-cells to propose new therapeutic strategies. We showed that Tax induces Bfl-1 and Bcl-xL expression via the NF-κB pathway, whereas HBZ has no effect on their expression. Tax also cooperates with c-Jun and JunD transcription factors of AP-1 family to increase the expression of these anti-apoptotic genes. By contrast, HBZ modulates the Tax-induced bfl-1 trans-activation. Altogether, our data indicate that Tax plays a key role in activating Bfl-1 and Bcl-xL expression and suggests that Bfl-1 and Bcl-xL are potentially expressed during the early and the late stages of ATLL development. Using short hairpin RNA strategy, we then showed that Bfl-1 and/or Bcl-xL are involved in HTLV-1-infected T-cell line survival, indicating that Bfl-1 and Bcl-xL represent potential therapeutic targets in the case of ATLL. One approach currently being developed in anti-cancer drug discovery is to search for small inhibitory compounds targeting anti-apoptotic proteins of the Bcl-2 family. But so far, no drug specifically targeting Bfl-1 is available. In collaboration with the IMAXIO Company, we have identified 83 molecules able to inhibit Bfl-1 anti-apoptotic activity using two high-throughput screening. One of these molecules specifically induced the death of HTLV-1-infected T-cell for which Bfl-1 represents a survival gene. This work provides new insight for long-term development of future drugs directed against Bfl-1 and should allow us to propose new therapeutic strategy for ATLL treatment
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Ferreira, Mari Cleia Martins Rodrigues. "Estudo da expressão do gene hSecurina e quantificação do índice de DNA em portadores assintomáticos do vírus linfotrópico T humano tipo 1 e pacientes com leucemia/linfoma de células T do adulto." Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/5/5167/tde-10012017-095656/.
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INTRODUÇÃO: A Leucemia/linfoma de células T do Adulto (ATL) é uma doença maligna de fenótipo T CD3+/CD4+/CD25+/CD7- e, geneticamente, apresenta cariótipo complexo e aneuploidia. Clinicamente muito agressiva e ainda incurável, está associada ao vírus linfotrópico T humano do tipo-1 que, preferencialmente, infecta linfócitos T CD4+. Dos indivíduos portadores do HTLV-1, somente 3-5% irão evoluir para ATL e após longo período de latência. Entretanto, os fatores virais ou do hospedeiro que estão associados com a progressão para ATL permanecem desconhecidos. O proto-oncogene hSecurina é um regulador mitótico importante para o processo de segregação cromossômica durante a separação das cromátides irmãs e está envolvido na patogênese de vários tumores. Com o objetivo de avaliar o conteúdo de DNA, o ciclo celular e a expressão do gene hSecurina em células T CD4+ e CD8+ dos portadores assintomáticos do HTLV-1 em comparação com ATL e indivíduos saudáveis, nos propusemos a realizar o presente estudo. MÉTODOS: Foram avaliados 38 portadores assintomáticos do HTLV-1, 20 casos de ATL pareados por sexo e idade com 35 indivíduos saudáveis. Foram estudados, individualmente, os subtipos linfocitários T CD4+ e CD8+, sendo o ciclo celular avaliado por citometria de fluxo e a expressão do gene hSecurina pela reação em cadeia da polimerase quantitativa em Tempo Real. RESULTADOS: Neste estudo, observamos parada de maturação de linfócitos T CD4+ na fase G0/G1 em portadores assintomáticos do HTLV-1 com diferença estatisticamente significante em comparação aos grupos-controle (p=0,041) e ATL (p=0,023). No grupo de portadores assintomáticos, observamos correlação inversa entre a porcentagem de células em G0/G1 e expressão de hSecurina (p=0,018) em linfócitos T CD4+. Porém, neste mesmo grupo, houve correlação direta entre porcentagem de células em fase S e expressão de hSecurina em linfócitos T CD4+ (p=0,001). Como esperado, observou-se maior fase S em ATL em comparação aos grupos-controle (p=0,020) e portador do HTLV-1 (p < 0,001). CONCLUSÃO: Neste estudo, demonstramos que linfócitos T CD4+ de portadores assintomáticos do vírus HTLV-1 apresentam atraso no ciclo celular com aumento de células na fase G0/G1. Este retardo da progressão do ciclo celular correlacionou-se de forma inversamente proporcional à expressão do gene hSecurinaINTRODUCTION: Adult T-Cell Leukemia (ATL) is a malignant disease of the CD3+/CD4+/CD25+/CD7- T-lymphocytes and genetically features complex karyotypes and aneuploidy. It is a clinically aggressive disease, which is yet incurable. It is associated with the human T-cell leukemia virus type 1 (HTLV-1) that preferentially infects CD4+ T-lymphocytes. Among all individuals that carry HTLV-1, only 3-5% will develop ATL and that too after a long latency period. However, the viral or host factors that are associated with the progression of ATL remain unknown. The proto-oncogene hSecurin is an important mitotic regulator for the process of chromosome segregation during sister chromatid separation and is involved in the pathogenesis of various tumors. We decided to conduct this study in order to analyze the DNA content, cell cycle, and expression of the hSecurin gene in CD4+ and CD8+ T cells of asymptomatic HTLV-1 carriers compared with that in ATL and healthy individuals. METHODS: We evaluated 38 asymptomatic HTLV-1 carriers, 20 patients with ATL, and 35 healthy subjects paired by sex and age. We individually studied the lymphocyte subtypes T CD4+ and CD8+; their cell cycles were evaluated by flow cytometry, and the expression of the hSecurin gene was analyzed using quantitative real time polymerase chain reaction. RESULTS: We observed lymphocyte maturation arrest in CD4+ T cells in the G0/G1 phase of asymptomatic HTLV-1 carriers with a statistically significant difference compared to that in the control (p = 0.041) and ATL (p = 0.023) groups. In the asymptomatic HTLV-1 carrier group, we also found an inverse correlation between the percentage of cells in G0/G1 phase and the hSecurin expression (p = 0.018) in TCD4+ lymphocytes. However, in this same group, there was also a direct correlation between the percentage of S phase cells and hSecurin expression in TCD4+ lymphocytes (p = 0.001). As expected, there was a higher number of S phase cells in the ATL group compared to that in the control (p = 0.020) and asymptomatic HTLV-1 carrier (p < 0.001) groups. CONCLUSION: In this study, we demonstrated that CD4 + T lymphocytes from asymptomatic HTLV-1 virus carriers present cell cycle arrest with increased G0/G1 phase cells. This delay in cell cycle progression correlated inversely with the expression of the hSecurin gene
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Meireles, Ana Luísa Langanke Pedroso. "Quantificação de células endoteliais circulantes em portadores assintomáticos do vírus linfotrópico humano de células T do tipo 1 (HTLV1) por citometria de fluxo." Universidade de São Paulo, 2009. http://www.teses.usp.br/teses/disponiveis/5/5167/tde-09062009-170516/.
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Células endoteliais provenientes da medula óssea (MO) participam da fisiopatologia de várias doenças que possuem dano vascular como fator em comum. Apesar de consideradas evento raro, encontram-se em quantidade aumentada na circulação periférica de pacientes oncológicos. Evidências sugerem que células endoteliais progenitoras (CEPs) contribuem para a angiogênese tumoral. Com esta descoberta, CEPs e células endoteliais maduras (CEMs) vêm sendo estudadas como potenciais alvos terapêuticos com o uso de drogas anti-angiogênicas. Portadores do vírus linfotrópico humano de células T do tipo 1 (HTLV1) têm possibilidade de desenvolver doenças causadas pelo vírus com elevada taxa de mortalidade, com destaque para a Leucemia/Linfoma de células T do Adulto (ATL). O tratamento para a forma sintomática da doença permanece desapontador. Este foi um estudo transversal desenvolvido com o objetivo de quantificar células endoteliais circulantes no sangue de portadores assintomáticos do HTLV1 em comparação a indivíduos saudáveis, por citometria de fluxo. Foram estudados 30 indivíduos portadores do vírus HTLV1, pareados por idade e sexo com o grupo controle. Três pacientes tiveram o diagnóstico de ATL sendo retirados da pesquisa. Foi utilizada como critério de inclusão a sorologia para HTLV1+, e negativa para as demais doenças transmissíveis por transfusão. Em nosso estudo os valores de CEPs encontrados foram maiores na população portadora assintomática (mediana: 0,8288 células / mm 3 ) em relação à população controle (mediana: 0,4905 células / mm 3 ; p = 0,035). Não houve diferença estatística entre a quantificação de CEMs e células endoteliais ativadas entre os portadores assintomáticos e o grupo controle saudável. Nossos achados sugerem que exista atividade angiogênica mesmo na ausência de transformação neoplásica, e que o valor de CEPs pode ser utilizado como marcador de atividade de doença e aplicado para monitorar a eficácia antitumoral da terapia antiangiogênicaEndothelial cells originated from the bone marrow (BM) take part in the physiopathology of several diseases which have vascular damage as a common factor. In spite of being a rare event, they are found in augmented quantity in the peripheral circulation of cancer patients. Evidence indicates that bone marrow-derived endothelial progenitor cells (CEPs) can contribute to tumor angiogenesis. Upon such a finding, circulating CEPs and mature endothelial cells (CEMs) have been researched as potential therapeutic targets and antiangiogenic drugs can be an option in anti-tumor therapy. Human T Cell Lymphotropic Virus Type 1 (HTLV1) carriers may develop diseases caused by the virus with high mortality rate, especially adult T-cell leukemia/lymphoma (ATL). The treatment for the symptomatic form of the disease remains disappointing. This cross-sectional study aimed at quantifying circulating endothelial cells in the blood of HTLV1 asymptomatic carriers in comparison to healthy individuals by flow cytometry. A sample of 30 individuals, HTLV1 carriers, age and sex paired, has been compared to the control group. Three patients were diagnosed with ATL, and deleted. HTLV1+ serology has been utilized as inclusion criteria, and negative for the remaining transfusion-transmittable diseases. CEPs values were greater in the asymptomatic carrier population (median: 0,8288 cells/mm 3 ) in relation to the control population (median: 0,4905 cells/mm 3 ; p = 0,035). There was no statistically significant difference in the quantification of CEMs and activated endothelial cells between asymptomatic carriers and the control group. This evidence suggest that there is angiogenic activity without neoplasic transformation, and the level of circulating endothelial progenitor cells can be used as biologic marker of disease activity and can reflect the antitumor efficacy of angiogenesis inhibitors
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Brocardo, Graciela Aparecida. "Avaliação do comprimento dos telômeros em células infectadas pelo vírus HTLV-I utilizando a técnica hibridização in situ fluorescente e citometria de fluxo (Flow-FISH)." Universidade de São Paulo, 2008. http://www.teses.usp.br/teses/disponiveis/5/5167/tde-25032009-174020/.
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INTRODUÇÃO: A Leucemia/Linfoma de células T do adulto (ATL) é uma doença linfoproliferativa crônica com transformação clonal predominantemente de linfócitos TCD4+, causada pelo vírus linfotrópico T humano do tipo I (HTLV-I). A ATL se desenvolve em 3-5% dos portadores do vírus HTLV-I, após longo período de latência clínica, acompanhado de expansão clonal dos linfócitos infectados. As células da ATL apresentam várias anormalidades cromossômicas, semelhantes àquelas resultantes de disfunção telomérica e a instabilidade genômica contribui para o desenvolvimento da ATL. Para entender o papel do encurtamento telomérico na oncogênese da ATL, avaliamos o comprimento dos telômeros de linfócitos TCD4 e TCD8 em portadores do vírus HTLV-I e em portadores de ATL. RESULTADOS: Não foi evidenciada diferença significativa no comprimento de telômero dos subtipos linfocitários TCD4+ e TCD8+ entre portadores do vírus HTLV-I e indivíduos saudáveis, assim como, entre portadores de ATL e indivíduos saudáveis. Entretanto, quando incluímos na análise a variável idade, evidenciamos redução significativa do comprimento do telômero com a idade em portadores do vírus HTLV-I e maior perda telomérica nos portadores do vírus HTLV-I e portadores de ATL em relação aos indivíduos saudáveis de mesma idade, embora a diferença entre os grupos não atinja o nível de significância estatística. Estes resultados podem ser explicados pelo fato de que as células dos indivíduos infectados pelo vírus HTLV-I apresentam maior taxa proliferativa devido à ação viral, mesmo em estado de latência clínica. A perda telomérica em função da idade nos portadores de ATL não demostrou-se significativa devido ao pequeno número de casos analisados em decorrência da raridade da doença. Entretanto, quando analisamos o comprimento telomérico nos subtipos linfocitários de portadores de ATL, evidenciamos acentuada perda telomérica na célula maligna e valores próximos ao limite superior esperado para a idade no subtipo linfocitário não transformado, demonstrando que a disfunção telomérica deve estar associada à transformação celular. Estabelecemos valores de referência de comprimento telomérico dos subtipos linfocitários TCD4+ e TCD8+ de indivíduos saudáveis, definidos por faixa etária. CONCLUSÃO: Nossos resultados demonstram que portadores do vírus HTLV-I apresentam maior perda telomérica em função da idade que indivíduos saudáveis, mas, sem refletir significância estatística e clínica. Entretanto, portadores de ATL apresentam perda acentuada de comprimento de telômero na célula maligna, demonstrando que a determinação do comprimento de telômero pode auxiliar futuramente o monitoramento dos indivíduos infectados pelo HTLV-I, indicando conversão à doençaINTRODUCTION: Adult T-cell Leukemia/Lymphoma (ATL) is a chronic lymphproliferative disease with clonal transformation predominantly of the TCD4+ lymphocytes, caused by the Human T lymphotropic virus type-I (HTLV-I). ATL develops itself in 3-5% of HTLV-I carriers after a long period of clinical latency accompanied by clonal expansion of the infected lymphocytes. The ATL cells present several chromosomic abnormalities, similar to those resulting from telomere dysfunction and the genomic instability contributes to the development of ATL. In order to understanding the role of telomeric shortening in the ATL oncogenesis, we assessed the length of telomeres of lymphocytes TCD4 and TCD8 in HTLV-I carriers and in ATL carriers. RESULTS: No significant difference was evidentiated in the telomere length of lymphocytary subtypes TCD4+ and TCD8+ between HTLV-I carriers and healthy subjects, as well as, between ATL carriers and healthy subjects. However, when the age variable was included in the analysis, we observed significant decrease of telomeric length with age progression in HTLV-I carriers and higher telomeric loss in HTLV-I carriers and ATL carriers when compared to healthy subjects of the same age, although the difference between groups does not reach the level of statistic relevance. These results may be explained by the fact that the cells of HTLV-I infected subjects present higher proliferative rate due to the viral action, even during clinical latency. Age-related telomeric loss in ATL carriers did not manifest itself as significant due to the small number of analyzed cases as a consequence of the diseases rareness. However, when the telomere length on the lymphocytary subtypes of ATL carriers was analyzed, we evidentiated accentuated telomeric loss in the malignant cell and values close to the age-expected upper limit in the nontransformed lymphocytary subtype, demonstrating that the telomere dysfunction may be associated to the cellular transformation. We have determined reference values of telomere length for lymphocytary subtypes TCD4+ and TCD8+ on healthy subjects, defined by age range. CONCLUSION: Our results demonstrate that HTLV-I carriers present higher telomeric loss due to age than healthy subjects, however, with no reflection in clinical and statistical significance. Nevertheless, ATL carriers present accentuated loss of telomere length in the malignant cell, demonstrating that the telomere length determination may, in the future, assist in the monitoring of HTLV-I infected subjects, indicating conversion to the disease
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Heidari, Mansour. "Investigation of the molecular function of the nuclear oncoprotein HOX11 in human t-cell leukaemia." Thesis, Heidari, Mansour (2003) Investigation of the molecular function of the nuclear oncoprotein HOX11 in human t-cell leukaemia. PhD thesis, Murdoch University, 2003. https://researchrepository.murdoch.edu.au/id/eprint/71/.
Full textAbstract:
HOXll, the prototypical member of the HOXll family (HOX11, HOXllLl and HOXllL2) was originally discovered as a transcriptional regulator aberrantly expressed in tumours with an immature T-cell phenotype (T-ALL) as a result of specific chromosomal translocations involving T-cell receptor loci. Subsequently, it was revealed that HOXll is required for normal spleen development since newborn Hoxll-/- mice exhibit asplenia. In both its normal and abnormal roles, HOXll has been postulated to function by binding regulatory elements within specific target genes to control gene transcription. However, very few genomic targets of HOX11 have been identified and little is known about its mode of action. In this study, we sought to further understand the role of HOX11 in controlling differentiation and cell growth by 1) determining the identity of genomic sequences that are directly bound by HOXll and 2) determining the identity of proteins which exist within HOXll-containing nuclear complexes.
To identify direct HOXll target sequences, a whole genome PCR-based screening method was employed using immobilised recombinant HOXll that had first been expressed as a biologically active GST fusion protein. Using this approach, restriction enzyme-cleaved human genomic DNA was selected for high-affinity HOXll binding sites. Unexpectedly, almost all clones isolated contained sequences derived from satellite 2 DNA that, together with related satellite 3 DNA, is found on most chromosomes at transcriptionally inactive pericentromeric heterochromatin. The specific binding of HOXl1 to satellite 2 DNA was verified by bandshift assays using both recombinant HOXll protein and nuclear extract derived from the T-ALL cell line, ALL-SIL. DNA-protein complexes containing HOX11 were identified by their ablation upon addition of HOXl1 antibody.
To confirm that HOXll associates with pericentromeric heterochromatin in vivo, HOXll was characterised in terms of its nuclear localisation during interphase in unsynchronised leukaemic T-cells (ALL-SIL) harbouring a translocation involving the HOXll locus. Using indirect immunofluorescence and confocal microscopy, HOXll antibody produced a punctate pattern of staining in the nucleus with discrete areas of dense staining superimposed on a diffuse distribution of HOXll protein. By dual staining, the bright HOXll foci correlated with centromeres since they overlapped with signals detected by an antibody specific for the centromeric protein CENP-B. Further evidence for a direct interaction of HOXll with satellite 2 DNA was provided by chromatin immunoprecipitation assay. In the presence of HOXll antibody, DNA fragments containing satellite 2 sequences were irnmunoprecipitated from sheared, cross-linked ALL-SIL chromatin but not from chromatin isolated from the HOXll-negative T-cell line PER-1 17. Finally, using a combination of immunoprecipitation with HOXll antibody, gel electrophoresis and mass peptide fingerprinting, a set of nuclear proteins were identified as potential HOXll interactors which are known to either localise to centromeric regions or act as regulators of gene expression. Together, these results implicate HOXl 1 in a functional interaction with centromeric heterochromatin, which may be a key feature of this oncoprotein in terms of both its T-cell transformation and transcriptional regulatory functions.
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Heidari, Mansour. "Investigation of the molecular function of the nuclear oncoprotein HOX11 in human t-cell leukaemia." Murdoch University, 2003. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20060815.123509.
Full textAbstract:
HOXll, the prototypical member of the HOXll family (HOX11, HOXllLl and HOXllL2) was originally discovered as a transcriptional regulator aberrantly expressed in tumours with an immature T-cell phenotype (T-ALL) as a result of
specific chromosomal translocations involving T-cell receptor loci. Subsequently, it was revealed that HOXll is required for normal spleen development since newborn
Hoxll-/- mice exhibit asplenia. In both its normal and abnormal roles, HOXll has been postulated to function by binding regulatory elements within specific target genes
to control gene transcription. However, very few genomic targets of HOX11 have been identified and little is known about its mode of action. In this study, we sought to
further understand the role of HOX11 in controlling differentiation and cell growth by 1) determining the identity of genomic sequences that are directly bound by HOXll and 2) determining the identity of proteins which exist within HOXll-containing nuclear complexes.
To identify direct HOXll target sequences, a whole genome PCR-based screening method was employed using immobilised recombinant HOXll that had first been expressed as a biologically active GST fusion protein. Using this approach, restriction enzyme-cleaved human genomic DNA was selected for high-affinity HOXll binding sites. Unexpectedly, almost all clones isolated contained sequences derived from satellite 2 DNA that, together with related satellite 3 DNA, is found on most chromosomes at transcriptionally inactive pericentromeric heterochromatin. The specific binding of HOXl1 to satellite 2 DNA was verified by bandshift assays using
both recombinant HOXll protein and nuclear extract derived from the T-ALL cell line, ALL-SIL. DNA-protein complexes containing HOX11 were identified by their ablation upon addition of HOXl1 antibody.
To confirm that HOXll associates with pericentromeric heterochromatin in vivo, HOXll was characterised in terms of its nuclear localisation during interphase in
unsynchronised leukaemic T-cells (ALL-SIL) harbouring a translocation involving the HOXll locus. Using indirect immunofluorescence and confocal microscopy, HOXll
antibody produced a punctate pattern of staining in the nucleus with discrete areas of dense staining superimposed on a diffuse distribution of HOXll protein. By dual staining, the bright HOXll foci correlated with centromeres since they overlapped with signals detected by an antibody specific for the centromeric protein CENP-B. Further evidence for a direct interaction of HOXll with satellite 2 DNA was provided by chromatin immunoprecipitation assay. In the presence of HOXll antibody, DNA fragments containing satellite 2 sequences were irnmunoprecipitated from sheared, cross-linked ALL-SIL chromatin but not from chromatin isolated from the HOXll-negative T-cell line PER-1 17. Finally, using a combination of immunoprecipitation with HOXll antibody, gel electrophoresis and mass peptide fingerprinting, a set of nuclear proteins were identified as potential HOXll interactors which are known to either localise to centromeric regions or act as regulators of gene expression. Together, these results implicate HOXl 1 in a functional interaction with centromeric heterochromatin,
which may be a key feature of this oncoprotein in terms of both its T-cell transformation and transcriptional regulatory functions.
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Mayakonda, Thippeswamy Anand [Verfasser], and Christoph [Akademischer Betreuer] Plass. "Epigenetic blueprint of human thymopoiesis and adult T-cell Acute Lymphoblastic Leukemia / Anand Mayakonda Thippeswamy ; Betreuer: Christoph Plass." Heidelberg : Universitätsbibliothek Heidelberg, 2021. http://d-nb.info/1237415071/34.
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Ibach, Tabea [Verfasser]. "Adoptive T-cell Therapy via Chimeric Antigen Receptors (CARs) against Leukemia in Combination with a human Suicide Gene / Tabea Ibach." Düsseldorf : Universitäts- und Landesbibliothek der Heinrich-Heine-Universität Düsseldorf, 2020. http://d-nb.info/1209354136/34.
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MURATE, TAKASHI, MASANORI DAIBATA, KAZUNORI OHNISHI, YOSUKE OSAWA, MOTOSHI SUZUKI, TETSUHITO KOJIMA, AKIRA TAKAGI, et al. "INVOLVEMENT OF KRAS G12A MUTATION IN THE IL-2-INDEPENDENT GROWTH OF A HUMAN T-LGL LEUKEMIA CELL LINE, PLT-2." Nagoya University School of Medicine, 2012. http://hdl.handle.net/2237/16737.
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Waskow, Claudia, Bonin Malte von, Martin Wermke, Cosgun Kadriye Nehir, Christian Thiede, Martin Bornhauser, and Gerard Wagemaker. "In Vivo Expansion of Co-Transplanted T Cells Impacts on Tumor Re-Initiating Activity of Human Acute Myeloid Leukemia in NSG Mice." Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2016. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-191633.
Full textAbstract:
Human cells from acute myeloid leukemia (AML) patients are frequently transplanted into immune-compromised mouse strains to provide an in vivo environment for studies on the biology of the disease. Since frequencies of leukemia re-initiating cells are low and a unique cell surface phenotype that includes all tumor re-initiating activity remains unknown, the underlying mechanisms leading to limitations in the xenotransplantation assay need to be understood and overcome to obtain robust engraftment of AML-containing samples. We report here that in the NSG xenotransplantation assay, the large majority of mononucleated cells from patients with AML fail to establish a reproducible myeloid engraftment despite high donor chimerism. Instead, donor-derived cells mainly consist of polyclonal disease-unrelated expanded co-transplanted human T lymphocytes that induce xenogeneic graft versus host disease and mask the engraftment of human AML in mice. Engraftment of mainly myeloid cell types can be enforced by the prevention of T cell expansion through the depletion of lymphocytes from the graft prior transplantation.
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Waskow, Claudia, Bonin Malte von, Martin Wermke, Cosgun Kadriye Nehir, Christian Thiede, Martin Bornhauser, and Gerard Wagemaker. "In Vivo Expansion of Co-Transplanted T Cells Impacts on Tumor Re-Initiating Activity of Human Acute Myeloid Leukemia in NSG Mice." Public Library of Science, 2013. https://tud.qucosa.de/id/qucosa%3A28097.
Full textAbstract:
Human cells from acute myeloid leukemia (AML) patients are frequently transplanted into immune-compromised mouse strains to provide an in vivo environment for studies on the biology of the disease. Since frequencies of leukemia re-initiating cells are low and a unique cell surface phenotype that includes all tumor re-initiating activity remains unknown, the underlying mechanisms leading to limitations in the xenotransplantation assay need to be understood and overcome to obtain robust engraftment of AML-containing samples. We report here that in the NSG xenotransplantation assay, the large majority of mononucleated cells from patients with AML fail to establish a reproducible myeloid engraftment despite high donor chimerism. Instead, donor-derived cells mainly consist of polyclonal disease-unrelated expanded co-transplanted human T lymphocytes that induce xenogeneic graft versus host disease and mask the engraftment of human AML in mice. Engraftment of mainly myeloid cell types can be enforced by the prevention of T cell expansion through the depletion of lymphocytes from the graft prior transplantation.
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Abozaid, Suhair Mohamed. "Studies on the interaction of surfactant protein SP-D with Inflenza A virus, Aspergillus fumigatus and dendritic cells." Thesis, Brunel University, 2016. http://bura.brunel.ac.uk/handle/2438/13595.
Full textAbstract:
Surfactant proteins, SP-A and SP-D, are collagen-containing calcium-dependent (C-type) lectins, called, collectins. Their primary structure has four regions: a cysteine-linked N- terminal region involved in multimerization, a collagen region composed of Gly-X-Y repeats, coiled-coil neck region, and the C-terminal carbohydrate recognition domains (CRD) or C-type lectin domain. SP-A looks like a bouquet, while SP-D is a cruciform- like structure, with four arms of equal length. SP-A and SP-D have been shown to act as innate immune molecules at pulmonary as well as extra-pulmonary sites by binding to pathogens, allergens and apoptotic/necrotic cells via their CRD region. SP-A and SP-D can induce pathogen neutralization and enhanced phagocytosis. In addition, SP-A and SP-D can interact via CRDs with allergens and dampen allergic reaction in vitro and in vivo. This thesis examines in vitro interaction of a recombinant fragment of human SP-D containing neck and CRD regions (rhSP-D) with IAV and Aspergillus fumigatus, in addition to characterizing a dichotomy of the effects of SP-A and SP-D on dendritic cells in an attempt to explain how SP-A and SP-D modulate DC functions differentially. Experiments involving interaction of rhSP-D with IAV pandemic strain show that it can be a restrictive factor against the virus, in addition to modulating immune response by a macrophage cell line. The rhSP-D can have anti-A. fumigatus effect directly and indirectly in the context of pathogen as well as allergen. A comparison has been made between two recombinant fragments of SP-D that have been expressed with and without 8 Gly-X-Y repeats for their fungistatic properties. The effects of SP-A and SP-D on cultured DC maturation, and effector cytokine and proliferative response of co-cultured cells have also been examined in vitro.
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Mathieu-Mahul, Danièle. "Analyse moleculaire d'anomalies chromosomiques specifiques d'hemopathies malignes humaines." Paris 7, 1987. http://www.theses.fr/1987PA077133.
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Mutambu, Susan L. "Seroepidemiology of Plasmodium falciparum, human immunodeficiency virus and human T-cell leukemia virus infections in mothers and their infants in Zimbabwe." Thesis, 1995. http://hdl.handle.net/10125/9443.
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Siddon, Nicole Ann. "The role of chromatin in the transcriptional regulation of the human T-cell leukemia virus type 1." Phd thesis, 2000. http://hdl.handle.net/1885/147385.
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Chen, Cheng-Tao, and 陳成桃. "The Seroepidemiology and Molecular Epidemiology of Human T Cell Leukemia Virus Type I in Taiwan and Kinmen." Thesis, 1996. http://ndltd.ncl.edu.tw/handle/33389335961572459723.
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Chuang, Pei-Chun, and 莊珮君. "The Development of Human T Cell Leukemia Virus Type I and Type II Recombinant Protein Vaccine and DNA Vaccine." Thesis, 1996. http://ndltd.ncl.edu.tw/handle/23010738750262170428.
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Lin, Ming-Tseh, and 林敏哲. "Molecular Epidemiological Studies of Human T-lymphotropic Virus TypeⅠ and TypeⅡ Infection in Taiwan and Analyses of Provirus Integration in Adult T-cell Leukemia." Thesis, 1996. http://ndltd.ncl.edu.tw/handle/58577730785158270795.
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DEMIR, AHU. "KINETIC CHARACTERIZATION AND NEWLY DISCOVERED INHIBITORS FOR VARIOUS CONSTRUCTS OF HUMAN T-CELL LEUKEMIA VIRUS-I PROTEASE AND INHIBITION EFFECT OF DISCOVERED MOLECULES ON HTLV-1 INFECTED CELLS." Diss., 2010. http://hdl.handle.net/10919/19164.
Full textAbstract:
Discovered in 1980, HTLV-1 (Human T-cell Leukemia Virus-1), was the first identified human retrovirus and is shown to be associated with a variety of diseases including: adult T-cell leukemia lymphoma (ATLL), tropical spastic paraparesis/HTLV-1 associated myelopathy (TSP/HAM), chronic arthropathy, uveitis, infective dermatitis, and polymyositis. The mechanism by which the virus causes disease is still unknown. HTLV- 1 infection has been reported in many regions of the world but is most prevalent in Southern Japan, the Caribbean basin, Central and West Africa, the Southeastern United States, Melanesia, parts of South Africa, the Middle East and India. Approximately 30 million people are infected by HTLV-1 worldwide, although only 3-5% of the infected individuals evolve Adult T-cell Leukemia (ATL) during their life and the prognosis for those infected is still poor.
The retroviral proteases (PRs) are essential for viral replication because they process viral Gag and Gag-(Pro)-Pol polyproteins during maturation, much like the PR from Human Immunodeficiency Virus-1 (HIV-1). Various antiviral inhibitors are in clinical use and one of the most significant classes is HIV-1 PR inhibitors, which have used for antiretroviral therapy in the treatment of AIDS. HTLV-1 PR and HIV-1 PR are homodimeric aspartic proteases with 125 and 99 residues, respectively. Even though substrate specificities of these two enzymes are different, HTLV-1 PR shares 28% similarity with HIV-1 PR overall and the substrate binding sites have 45% similarity.
In addition to the 125-residue full length HTLV-1 PR, constructs with various C- terminal deletions (giving proteases with lengths of 116, 121, or 122 amino acids) were made in order to elucidate the effect of the residues in the C-terminal region. It was suggested that five amino acids in the C-terminal region are not necessary for the enzymatic activity in Hayakawa et al. 1992. In 2004 Herger et al. had suggested that 10 amino acids at the C-terminal region are not necessary for catalytic activity. A recent paper suggested that C-terminal residues are essential; and that catalytic activity lowers upon truncation, with even the last 5 amino acids necessary for full catalytic activity (1).
The mutation L40I has been made to prevent autoproteolysis and the W98V mutation was made to make the active site of HTLV-1 PR similar to HIV-1 PR. We have characterized C-terminal amino acids of HTLV-1 PR as not being essential for full catalytic activity. We have discovered potential new inhibitors by in silico screening of 116-HTLV-1 PR. These small molecules were tested kinetically for various constructs including the 116, 121 and 122-amino acid forms of HTLV-1 PR. Inhibitors with the best inhibition constants were used in HTLV-1 infected cells and one of the inhibitors seems to inhibit gag processing.
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Wu, Min-Huan, and 吳明寰. "Characterization of Human T-cell Leukemia/Lymphoma Virus Type I ( HTLV-I ) Envelope Proteins produced in Insect Cells by using GP46 and GP21 Monoclonal Antibodies." Thesis, 1994. http://ndltd.ncl.edu.tw/handle/30645389659407713218.
Full textAbstract:
碩士國立臺灣大學
動物學系
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The entire envelope gene of human T-cell leukemia/lymphoma virus type I ( HTLV-I ) has been successfully expressed in insect cell by using two baculovirus vectors, under the control of the polyhedrin and basic protein promoters. Even with the radio- immunoprecipitation analysis, the recombinant envelope proteins were still not detectable in the culture media of recombinant baculovirus infected Sf 21-AE cells. The glycosylation of expressed HTLV-I envelope proteins driven by basic protein promoter were more completely glycosylated. Treatment of glycosylation inhibitors ( tunicamycin and deoxynojirimycin ) has been shown to result in the absence of 21kDa HTLV-I envelope protein in recombinant baculovirus infected Sf 21-AE cells. The evidences suggest that the HTLV-I envelpoe protein precursor can be cleaved only after glycosylation. EndoH and PNGase F treatment revealed a high mannose type glycosylation having occurred in 46-62kDa proteins produced by polyhedrin promoter-driven expression and the 27, 30 and 46-62kDa proteins by basic protein promoter-driven expression.