Increased baseline RASGRP1 signals Enhance Stem Cell Fitness during Native Hematopoiesis

AbstractOncogenic mutations in RAS genes, like KRASG12D or NRASG12D, trap Ras in the active state and cause myeloproliferative disorder and T cell leukemia (T-ALL) when induced in the bone marrow via Mx1CRE. The RAS exchange factor RASGRP1 is frequently overexpressed in T-ALL patients. In T-ALL cell lines overexpression of RASGRP1 increases flux through the RASGTP/RasGDP cycle. Here we expanded RASGRP1 expression surveys in pediatric T-ALL and generated a RoLoRiG mouse model crossed to Mx1CRE to determine the consequences of induced RASGRP1 overexpression in primary hematopoietic cells. RASGRP1-overexpressing, GFP-positive cells outcompeted wild type cells and dominated the peripheral blood compartment over time. RASGRP1 overexpression bestows gain-of-function colony formation properties to bone marrow progenitors in medium containing limited growth factors. RASGRP1 overexpression enhances baseline mTOR-S6 signaling in the bone marrow, but not in vitro cytokine-induced signals. In agreement with these mechanistic findings, hRASGRP1-ires-EGFP enhances fitness of stem- and progenitor- cells, but only in the context of native hematopoiesis. RASGRP1 overexpression is distinct from KRASG12D or NRASG12D, does not cause acute leukemia on its own, and leukemia virus insertion frequencies predict that RASGRP1 overexpression can effectively cooperate with lesions in many other genes to cause acute T cell leukemia.

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Biochemical Characterization, Specificity and Inhibition Studies of HTLV-1, HTLV-2, and HTLV-3 Proteases

Life ◽

10.3390/life11020127 ◽

2021 ◽

Vol 11 (2) ◽

pp. 127

Author(s):

Norbert Kassay ◽

János András Mótyán ◽

Krisztina Matúz ◽

Mária Golda ◽

József Tőzsér

Keyword(s):

T Cell ◽

Biochemical Characterization ◽

Leukemia Virus ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Cell Leukemia Virus Type ◽

Adult T Cell Leukemia ◽

Human T Cell ◽

T Cell Leukemia Virus

The human T-lymphotropic viruses (HTLVs) are causative agents of severe diseases including adult T-cell leukemia. Similar to human immunodeficiency viruses (HIVs), the viral protease (PR) plays a crucial role in the viral life-cycle via the processing of the viral polyproteins. Thus, it is a potential target of anti-retroviral therapies. In this study, we performed in vitro comparative analysis of human T-cell leukemia virus type 1, 2, and 3 (HTLV-1, -2, and -3) proteases. Amino acid preferences of S4 to S1′ subsites were studied by using a series of synthetic oligopeptide substrates representing the natural and modified cleavage site sequences of the proteases. Biochemical characteristics of the different PRs were also determined, including catalytic efficiencies and dependence of activity on pH, temperature, and ionic strength. We investigated the effects of different HIV-1 PR inhibitors (atazanavir, darunavir, DMP-323, indinavir, ritonavir, and saquinavir) on enzyme activities, and inhibitory potentials of IB-268 and IB-269 inhibitors that were previously designed against HTLV-1 PR. Comparative biochemical analysis of HTLV-1, -2, and -3 PRs may help understand the characteristic similarities and differences between these enzymes in order to estimate the potential of the appearance of drug-resistance against specific HTLV-1 PR inhibitors.

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Overexpression of hTLX3 in Association with Mutant IL7Rα Is Sufficient to Generate T-ALL In Vivo and to Transform Thymocytes in Vitro

Blood ◽

10.1182/blood-2020-140060 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 21-21

Author(s):

Gisele Olinto Libanio Rodrigues ◽

Julie Hixon ◽

Hila Winer ◽

Erica Matich ◽

Caroline Andrews ◽

...

Keyword(s):

Bone Marrow ◽

T Cells ◽

T Cell ◽

Cell Line ◽

Lymphoblastic Leukemia ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Cell Counts

Mutations of the IL-7Rα chain occur in approximately 10% of pediatric T-cell acute lymphoblastic leukemia cases. While we have shown that mutant IL7Ra is sufficient to transform an immortalized thymocyte cell line, mutation of IL7Ra alone was insufficient to cause transformation of primary T cells, suggesting that additional genetic lesions may be present contributing to initiate leukemia. Studies addressing the combinations of mutant IL7Ra plus TLX3 overexpression indicates in vitro growth advantage, suggesting this gene as potential collaborative candidate. Furthermore, patients with mutated IL7R were more likely to have TLX3 or HOXA subgroup leukemia. We sought to determine whether combination of mutant hIL7Ra plus TLX3 overexpression is sufficient to generate T-cell leukemia in vivo. Double negative thymocytes were isolated from C57BL/6J mice and transduced with retroviral vectors containing mutant hIL7R plus hTLX3, or the genes alone. The combination mutant hIL7R wild type and hTLX3 was also tested. Transduced thymocytes were cultured on the OP9-DL4 bone marrow stromal cell line for 5-13 days and accessed for expression of transduced constructs and then injected into sublethally irradiated Rag-/- mice. Mice were euthanized at onset of clinical signs, and cells were immunophenotyped by flow cytometry. Thymocytes transduced with muthIL-7R-hTLX3 transformed to cytokine-independent growth and expanded over 30 days in the absence of all cytokines. Mice injected with muthIL7R-hTLX3 cells, but not the controls (wthIL7R-hTLX3or mutIL7R alone) developed leukemia approximately 3 weeks post injection, characterized by GFP expressing T-cells in blood, spleen, liver, lymph nodes and bone marrow. Furthermore, leukemic mice had increased white blood cell counts and presented with splenomegaly. Phenotypic analysis revealed a higher CD4-CD8- T cell population in the blood, bone marrow, liver and spleen compared in the mutant hIL7R + hTLX3 mice compared with mice injected with mutant IL7R alone indicating that the resulting leukemia from the combination mutant hIL7R plus hTLX3 shows early arrest in T-cell development. Taken together, these data show that oncogenic IL7R activation is sufficient for cooperation with hTLX3 in ex vivo thymocyte cell transformation, and that cells expressing the combination muthIL7R-hTLX3 is sufficient to trigger T-cell leukemia in vivo. Figure Disclosures No relevant conflicts of interest to declare.

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Tax and Overlapping Rex Sequences Do Not Confer the Distinct Transformation Tropisms of Human T-Cell Leukemia Virus Types 1 and 2

Journal of Virology ◽

10.1128/jvi.77.14.7728-7735.2003 ◽

2003 ◽

Vol 77 (14) ◽

pp. 7728-7735 ◽

Cited By ~ 19

Author(s):

Jianxin Ye ◽

Li Xie ◽

Patrick L. Green

Keyword(s):

T Cells ◽

T Cell ◽

Leukemia Virus ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Cell Leukemia Virus Type ◽

Human T Cell ◽

T Cell Leukemia Virus

ABSTRACT Human T-cell leukemia virus type 1 (HTLV-1) and HTLV-2 are distinct oncogenic retroviruses that infect several cell types but display their biological and pathogenic activity only in T cells. Previous studies have indicated that in vivo HTLV-1 has a preferential tropism for CD4+ T cells, whereas HTLV-2 in vivo tropism is less clear but appears to favor CD8+ T cells. Both CD4+ and CD8+ T cells are susceptible to HTLV-1 and HTLV-2 infection in vitro, and HTLV-1 has a preferential immortalization and transformation tropism of CD4+ T cells, whereas HTLV-2 immortalizes and transforms primarily CD8+ T cells. The molecular mechanism that determines this tropism of HTLV-1 and HTLV-2 has not been determined. HTLV-1 and HTLV-2 carry the tax and rex transregulatory genes in separate but partially overlapping reading frames. Since Tax has been shown to be critical for cellular transformation in vitro and interacts with numerous cellular processes, we hypothesized that the viral determinant of transformation tropism is encoded by tax. Using molecular clones of HTLV-1 (Ach) and HTLV-2 (pH6neo), we constructed recombinants in which tax and overlapping rex genes of the two viruses were exchanged. p19 Gag expression from proviral clones transfected into 293T cells indicated that both recombinants contained functional Tax and Rex but with significantly altered activity compared to the wild-type clones. Stable transfectants expressing recombinant viruses were established, irradiated, and cocultured with peripheral blood mononuclear cells. Both recombinants were competent to transform T lymphocytes with an efficiency similar to that of the parental viruses. Flow cytometry analysis indicated that HTLV-1 and HTLV-1/TR2 had a preferential tropism for CD4+ T cells and that HTLV-2 and HTLV-2/TR1 had a preferential tropism for CD8+ T cells. Our results indicate that tax/rex in different genetic backgrounds display altered functional activity but ultimately do not contribute to the different in vitro transformation tropisms. This first study with recombinants between HTLV-1 and HTLV-2 is the initial step in elucidating the different pathobiologies of HTLV-1 and HTLV-2.

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Allogeneic Bone Marrow Transplantation from Unrelated Human T-Cell Leukemia Virus-I–negative Donors for Adult T-Cell Leukemia/Lymphoma: Retrospective Analysis of Data from the Japan Marrow Donor Program

Biology of Blood and Marrow Transplantation ◽

10.1016/j.bbmt.2006.09.002 ◽

2007 ◽

Vol 13 (1) ◽

pp. 90-99 ◽

Cited By ~ 47

Author(s):

Koji Kato ◽

Yoshinobu Kanda ◽

Tetsuya Eto ◽

Tsuyoshi Muta ◽

Hisashi Gondo ◽

...

Keyword(s):

Bone Marrow ◽

T Cell ◽

Leukemia Virus ◽

Allogeneic Bone Marrow Transplantation ◽

Allogeneic Bone ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Adult T Cell Leukemia ◽

Human T Cell ◽

T Cell Leukemia Virus

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Inhibition of human T-cell leukemia virus type I by the short oligoguanylic acids in vitro

Materials Science and Engineering C ◽

10.1016/j.msec.2004.07.003 ◽

2004 ◽

Vol 24 (6-8) ◽

pp. 887-892

Author(s):

Da-Wei Yang ◽

Masahiro Shuda ◽

Hideko Kaji

Keyword(s):

T Cell ◽

Virus Type ◽

Leukemia Virus ◽

Type I ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Cell Leukemia Virus Type ◽

Human T Cell ◽

T Cell Leukemia Virus

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Comparison of the Genetic Organization, Expression Strategies and Oncogenic Potential of HTLV-1 and HTLV-2

Leukemia Research and Treatment ◽

10.1155/2012/876153 ◽

2012 ◽

Vol 2012 ◽

pp. 1-14 ◽

Cited By ~ 5

Author(s):

Francesca Rende ◽

Ilaria Cavallari ◽

Maria Grazia Romanelli ◽

Erica Diani ◽

Umberto Bertazzoni ◽

...

Keyword(s):

T Cell ◽

Leukemia Virus ◽

Spastic Paraparesis ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Pathogenic Potential ◽

Adult T Cell Leukemia ◽

Human T Cell

Human T cell leukemia virus types 1 and 2 (HTLV-1 and HTLV-2) are genetically related complex retroviruses that are capable of immortalizing human T-cells in vitro and establish life-long persistent infections in vivo. In spite of these apparent similarities, HTLV-1 and HTLV-2 exhibit a significantly different pathogenic potential. HTLV-1 is recognized as the causative agent of adult T-cell leukemia/lymphoma (ATLL) and tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM). In contrast, HTLV-2 has not been causally linked to human malignancy, although it may increase the risk of developing inflammatory neuropathies and infectious diseases. The present paper is focused on the studies aimed at defining the viral genetic determinants of the pathobiology of HTLV-1 and HTLV-2 through a comparison of the expression strategies and functional properties of the different gene products of the two viruses.

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In vitro infection of human B lymphocytes with adult T-cell leukemia virus

Cancer Letters ◽

10.1016/0304-3835(84)90038-7 ◽

1984 ◽

Vol 22 (1) ◽

pp. 11-21 ◽

Cited By ~ 7

Author(s):

Morihisa Okada ◽

Yoshio Koyanagi ◽

Nobuyuki Kobayashi ◽

Yuetsu Tanaka ◽

Masuyo Nakai ◽

...

Keyword(s):

T Cell ◽

B Lymphocytes ◽

Leukemia Virus ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Adult T Cell Leukemia ◽

T Cell Leukemia Virus

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Human T-Cell Leukemia Virus Type 1 Tax Oncoprotein Suppression of Multilineage Hematopoiesis of CD34+ Cells In Vitro

Journal of Virology ◽

10.1128/jvi.77.22.12152-12164.2003 ◽

2003 ◽

Vol 77 (22) ◽

pp. 12152-12164 ◽

Cited By ~ 23

Author(s):

Adam Tripp ◽

Yingxian Liu ◽

Michelle Sieburg ◽

Joanne Montalbano ◽

Stephen Wrzesinski ◽

...

Keyword(s):

T Cell ◽

Leukemia Virus ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Hematopoietic Progenitor ◽

Cell Leukemia Virus Type ◽

Human T Cell ◽

T Cell Lines

ABSTRACT Human T-cell leukemia virus type 1 (HTLV-1) and HTLV-2 are highly related viruses that differ in disease manifestation. HTLV-1 is the etiologic agent of adult T-cell leukemia and lymphoma, an aggressive clonal malignancy of human CD4-bearing T lymphocytes. Infection with HTLV-2 has not been conclusively linked to lymphoproliferative disorders. We previously showed that human hematopoietic progenitor (CD34+) cells can be infected by HTLV-1 and that proviral sequences were maintained after differentiation of infected CD34+ cells in vitro and in vivo. To investigate the role of the Tax oncoprotein of HTLV on hematopoiesis, bicistronic lentiviral vectors were constructed encoding the HTLV-1 or HTLV-2 tax genes (Tax1 and Tax2, respectively) and the green fluorescent protein marker gene. Human hematopoietic progenitor (CD34+) cells were infected with lentivirus vectors, and transduced cells were cultured in a semisolid medium permissive for the development of erythroid, myeloid, and primitive progenitor colonies. Tax1-transduced CD34+ cells displayed a two- to fivefold reduction in the total number of hematopoietic clonogenic colonies that arose in vitro, in contrast to Tax2-transduced cells, which showed no perturbation of hematopoiesis. The ratio of colony types that developed from Tax1-transduced CD34+ cells remained unaffected, suggesting that Tax1 inhibited the maturation of relatively early, uncommitted hematopoietic stem cells. Since previous reports have linked Tax1 expression with initiation of apoptosis, lentiviral vector-mediated transduction of Tax1 or Tax2 was investigated in CEM and Jurkat T-cell lines. Ectopic expression of either Tax1 or Tax2 failed to induce apoptosis in T-cell lines. These data demonstrate that Tax1 expression perturbs development and maturation of pluripotent hematopoietic progenitor cells, an activity that is not displayed by Tax2, and that the suppression of hematopoiesis is not attributable to induction of apoptosis. Since hematopoietic progenitor cells may serve as a latently infected reservoir for HTLV infection in vivo, the different abilities of HTLV-1 and -2 Tax to suppress hematopoiesis may play a role in the respective clinical outcomes after infection with HTLV-1 or -2.

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