Human T-cell leukemia virus type 2 Tax protein induces interleukin 2-independent growth in a T-cell line
- Rie Kondo†1, 2,
- Masaya Higuchi†1,
- Masahiko Takahashi1,
- Masayasu Oie1,
- Yuetsu Tanaka3,
- Fumitake Gejyo2 and
- Masahiro Fujii1Email author
© Kondo et al; licensee BioMed Central Ltd. 2006
Received: 18 August 2006
Accepted: 02 December 2006
Published: 02 December 2006
While human T-cell leukemia virus type 1 (HTLV-1) is a causative agent of adult T-cell leukemia, HTLV type 2 (HTLV-2) is not associated with this malignancy. Accumulating evidence suggests that Tax, a transforming protein of HTLV-1 or HTLV-2, plays a crucial role in the distinctive pathogenesis of these two infections. We herein examined whether Tax2 by itself has a growth promoting activity in a mouse T-cell line CTLL-2, and compared the activity with that of Tax1.
We found that Tax2 converts the cell growth of CTLL-2 from an interleukin(IL)-2-dependent growth into an independent one. Cyclosporine A, an inhibitor of transcription factor NFAT, inhibited the growth of two out of four Tax2-transformed CTLL-2 cells, but it had little effect on two Tax1-transformed cells. While the HTLV-2-transformed human T-cell lines produce a significant amount of IL-2, Tax2-transformed CTLL-2 cells only produced a minimal amount of IL-2. These results thus suggest that NFAT-inducible gene(s) other than IL-2 play a role in the cell growth of Tax2-transformed CTLL-2 cells.
These results show that HTLV-2 Tax2 by itself has a growth promoting activity toward a T-cell line CTLL-2, and the CTLL-2 assay used in this study may therefore be a useful tool for comparing the activity of Tax2 with that of Tax1 in T-cells, thereby elucidating the mechanism of HTLV-1 specific leukemogenesis.
Human T-cell leukemia virus type 1 (HTLV-1) and HTLV type 2 (HTLV-2) are a family of retroviruses, which share around a 70% nucleotide identity and similar biological properties [1–6]. For instance, both HTLV-1 and HTLV-2 can efficiently transform primary human T-cells in vitro and establish a life-long persistent infection in humans [7–9]. The clinical outcomes of these two infections are, however, significantly distinctive. While HTLV-1 is etiologically associated with adult T-cell leukemia (ATL), HTLV-2 is associated with only a few cases of variant hairy cell leukemia [5, 10–12].
HTLV-1 and HTLV-2 encode a transforming protein Tax1 and Tax2, respectively, which are essential for the transformation of primary human T-cells in vitro [13–16]. Accumulating evidence suggests that Tax1 is a factor responsible for the high-oncogenic activity of HTLV-1 relative to HTLV-2 [4, 5]. Tax1 and Tax2 have more than 75 % amino acid identities, and they also exhibit strikingly similar functions in infected cells [17, 18]. For instance, Tax1 and Tax2 induce the expression of a number of cellular genes through several transcription factor binding sites, such as NF-κB, CREB/ATF, SRF, and AP-1 [4, 19–25]. These Tax-inducible cellular genes play a critical role in the persistent infection in host T-cells, including the transformation of human T-cells [24, 25], but they alone can not explain the pathogenic differences between HTLV-1 and HTLV-2, since the potencies of these functions are equivalent. On the other hand, recent results identified several differences between Tax1 and Tax2, which are likely to be factors that are responsible for the pathogenic difference of two infections [4, 5, 26–35]. Therefore, a comparative analysis of Tax1 and Tax2 is a promising approach to identify a key process responsible for HTLV-1 specific leukemogenesis.
Tax2 has been shown to be essential for HTLV-2-mediated transformation of human T-cells . It, however, remains to be elucidated whether Tax2 by itself has a growth promoting activity toward T-cells like Tax1 . We herein showed that Tax2 can reproducibly convert a mouse T-cell line from an IL-2-dependent growth into an independent one. These results demonstrate that Tax2 by itself without any other viral proteins has a growth promoting activity in T-cells, thus suggesting that this growth promoting activity of Tax2 contributes to HTLV-2-mediated T-cell transformation. Since at least two functions, apoptosis inhibition and cell cycle promotion are both required for CTLL-2 to grow in the absence of IL-2, Tax2 can therefore replace these two functions in CTLL-2.
CsA inhibited the growth of two out of four Tax2-transformed CTLL-2 cells (Figure 3), indicating that NFAT-inducible genes are involved in IL-2-independent growth of these Tax2-transformed cells. These results are consistent with the previous results that CsA inhibited cell growth of some but not all HTLV-2-transformed human T-cell lines . There are at least two explanations for the distinct responses of the Tax2-transformed cells to CsA. Tax2 may have two distinctive activities to induce IL-2-independent growth of CTLL-2 cells. Alternatively, some parental CTLL-2 cells may have genetic or epigenetic change(s) conferring resistance to CsA in Tax2-transformed CTLL-2 cells. In contrast to Tax2, the cell growth of Tax1-transformed cells was little affected by CsA. This finding is also consistent with the result that Tax1 minimally activates NFAT, and thus CsA can not inhibit the cell growth of any HTLV-1-transformed T-cell lines .
Unlike the HTLV-2-transformed human T-cell lines sensitive to CsA-mediated growth inhibition, Tax2-transformed CsA-sensitive cells expressed a small amount of IL-2 mRNA (Figure 4). Since there are several NFAT inducible cytokines which promote T-cell growth, such as IL-4 and IL-21, these results indicated that the NFAT-inducible gene(s) other than IL-2 positively regulate the cell growth of the Tax2-transformed cells, thus suggesting that HTLV-2-transformed human T-cells may also utilize multiple NFAT-inducible T-cell growth promoting factors for their growth.
Accumulating evidence suggests that Tax plays a crucial role in the distinctive pathogenesis between HTLV-1 and HTLV-2 [4, 5, 26, 28, 29, 32, 34]. Therefore, further comparative studies of the Tax1 and Tax2 functions in T-cells are expected to advance our understanding of HTLV-1 leukemogenesis. The CTLL-2 assay used in this study is therefore considered to be a useful tool for examining the functions of Tax2 and Tax1 in T-cells, thereby elucidating the mechanism of HTLV-1 specific leukemogenesis.
We thank Dr. Hiroyuki Miyoshi at RIKEN Tsukuba Institute and Dr. William H. Hall for the CSII-EF-RfA plasmid and anti-Tax2 antibody, respectively. We also thank the Takeda Pharmaceutical Company for providing recombinant human IL-2. We would like to express our gratituded to Chika Yamamoto for her excellent technical assistance. This work was supported in part by a Grant-in-Aid for Scientific Research on Priority Areas and for Scientific Research (C) of Japan.
- Seiki M, Hattori S, Hirayama Y, Yoshida M: Human adult T-cell leukemia virus: complete nucleotide sequence of the provirus genome integrated in leukemia cell DNA. Proc Natl Acad Sci USA. 1983, 80: 3618-3622. 10.1073/pnas.80.12.3618.PubMed CentralView ArticlePubMedGoogle Scholar
- Chen IS, McLaughlin J, Gasson JC, Clark SC, Golde DW: Molecular characterization of genome of a novel human T-cell leukaemia virus. Nature. 1983, 305: 502-505. 10.1038/305502a0.View ArticlePubMedGoogle Scholar
- Azran I, Schavinsky-Khrapunsky Y, Aboud M: Role of Tax protein in human T-cell leukemia virus type-I leukemogenicity. Retrovirology. 2004, 1: 20-10.1186/1742-4690-1-20.PubMed CentralView ArticlePubMedGoogle Scholar
- Hall WW, Fujii M: Deregulation of cell-signaling pathways in HTLV-1 infection. Oncogene. 2005, 24: 5965-5975. 10.1038/sj.onc.1208975.View ArticlePubMedGoogle Scholar
- Feuer G, Green PL: Comparative biology of human T-cell lymphotropic virus type 1 (HTLV-1) and HTLV-2. Oncogene. 2005, 24: 5996-6004. 10.1038/sj.onc.1208971.PubMed CentralView ArticlePubMedGoogle Scholar
- Matsuoka M: Human T-cell leukemia virus type I (HTLV-I) infection and the onset of adult T-cell leukemia (ATL). Retrovirology. 2005, 2: 27-10.1186/1742-4690-2-27.PubMed CentralView ArticlePubMedGoogle Scholar
- Miyoshi I, Kubonishi I, Yoshimoto S, Akagi T, Ohtsuki Y, Shiraishi Y, Nagata K, Hinuma Y: Type C virus particles in a cord T-cell line derived by co-cultivating normal human cord leukocytes and human leukaemic T cells. Nature. 1981, 294: 770-771. 10.1038/294770a0.View ArticlePubMedGoogle Scholar
- Yamamoto N, Okada M, Koyanagi Y, Kannagi M, Hinuma Y: Transformation of human leukocytes by cocultivation with an adult T cell leukemia virus producer cell line. Science. 1982, 217: 737-739. 10.1126/science.6980467.View ArticlePubMedGoogle Scholar
- Chen IS, Quan SG, Golde DW: Human T-cell leukemia virus type II transforms normal human lymphocytes. Proc Natl Acad Sci USA. 1983, 80: 7006-7009. 10.1073/pnas.80.22.7006.PubMed CentralView ArticlePubMedGoogle Scholar
- Uchiyama T, Yodoi J, Sagawa K, Takatsuki K, Uchino H: Adult T-cell leukemia: clinical and hematologic features of 16 cases. Blood. 1977, 50: 481-492.PubMedGoogle Scholar
- Takatsuki K: Discovery of adult T-cell leukemia. Retrovirology. 2005, 2: 16-10.1186/1742-4690-2-16.PubMed CentralView ArticlePubMedGoogle Scholar
- Gallo RC: The discovery of the first human retrovirus: HTLV-1 and HTLV-2. Retrovirology. 2005, 2: 17-10.1186/1742-4690-2-17.PubMed CentralView ArticlePubMedGoogle Scholar
- Grassmann R, Berchtold S, Radant I, Alt M, Fleckenstein B, Sodroski JG, Haseltine WA, Ramstedt U: Role of human T-cell leukemia virus type 1 X region proteins in immortalization of primary human lymphocytes in culture. J Virol. 1992, 66: 4570-4575.PubMed CentralPubMedGoogle Scholar
- Akagi T, Shimotohno K: Proliferative response of Tax1-transduced primary human T cells to anti-CD3 antibody stimulation by an interleukin-2-independent pathway. J Virol. 1993, 67: 1211-1217.PubMed CentralPubMedGoogle Scholar
- Ross TM, Pettiford SM, Green PL: The tax gene of human T-cell leukemia virus type 2 is essential for transformation of human T lymphocytes. J Virol. 1996, 70: 5194-5202.PubMed CentralPubMedGoogle Scholar
- Robek MD, Ratner L: Immortalization of CD4(+) and CD8(+) T lymphocytes by human T-cell leukemia virus type 1 Tax mutants expressed in a functional molecular clone. J Virol. 1999, 73: 4856-4865.PubMed CentralPubMedGoogle Scholar
- Slamon DJ, Shimotohno K, Cline MJ, Golde DW, Chen IS: Identification of the putative transforming protein of the human T-cell leukemia viruses HTLV-I and HTLV-II. Science. 1984, 226: 61-65. 10.1126/science.6089351.View ArticlePubMedGoogle Scholar
- Seiki M, Inoue J, Takeda T, Yoshida M: Direct evidence that p40x of human T-cell leukemia virus type I is a trans-acting transcriptional activator. Embo J. 1986, 5: 561-565.PubMed CentralPubMedGoogle Scholar
- Cross SL, Feinberg MB, Wolf JB, Holbrook NJ, Wong-Staal F, Leonard WJ: Regulation of the human interleukin-2 receptor alpha chain promoter: activation of a nonfunctional promoter by the transactivator gene of HTLV-I. Cell. 1987, 49: 47-56. 10.1016/0092-8674(87)90754-9.View ArticlePubMedGoogle Scholar
- Maruyama M, Shibuya H, Harada H, Hatakeyama M, Seiki M, Fujita T, Inoue J, Yoshida M, Taniguchi T: Evidence for aberrant activation of the interleukin-2 autocrine loop by HTLV-1-encoded p40x and T3/Ti complex triggering. Cell. 1987, 48: 343-350. 10.1016/0092-8674(87)90437-5.View ArticlePubMedGoogle Scholar
- Zhao LJ, Giam CZ: Human T-cell lymphotropic virus type I (HTLV-I) transcriptional activator, Tax, enhances CREB binding to HTLV-I 21-base-pair repeats by protein-protein interaction. Proc Natl Acad Sci USA. 1992, 89: 7070-7074. 10.1073/pnas.89.15.7070.PubMed CentralView ArticlePubMedGoogle Scholar
- Fujii M, Tsuchiya H, Chuhjo T, Akizawa T, Seiki M: Interaction of HTLV-1 Tax1 with p67SRF causes the aberrant induction of cellular immediate early genes through CArG boxes. Genes Dev. 1992, 6: 2066-2076.View ArticlePubMedGoogle Scholar
- Iwai K, Mori N, Oie M, Yamamoto N, Fujii M: Human T-cell leukemia virus type 1 tax protein activates transcription through AP-1 site by inducing DNA binding activity in T cells. Virology. 2001, 279: 38-46. 10.1006/viro.2000.0669.View ArticlePubMedGoogle Scholar
- Sun SC, Yamaoka S: Activation of NF-kappaB by HTLV-I and implications for cell transformation. Oncogene. 2005, 24: 5952-5964. 10.1038/sj.onc.1208969.View ArticlePubMedGoogle Scholar
- Grassmann R, Aboud M, Jeang KT: Molecular mechanisms of cellular transformation by HTLV-1 Tax. Oncogene. 2005, 24: 5976-5985. 10.1038/sj.onc.1208978.View ArticlePubMedGoogle Scholar
- Semmes OJ, Majone F, Cantemir C, Turchetto L, Hjelle B, Jeang KT: HTLV-I and HTLV-II Tax: differences in induction of micronuclei in cells and transcriptional activation of viral LTRs. Virology. 1996, 217: 373-379. 10.1006/viro.1996.0126.View ArticlePubMedGoogle Scholar
- Tanaka Y, Hayashi M, Takagi S, Yoshie O: Differential transactivation of the intercellular adhesion molecule 1 gene promoter by Tax1 and Tax2 of human T-cell leukemia viruses. J Virol. 1996, 70: 8508-8517.PubMed CentralPubMedGoogle Scholar
- Endo K, Hirata A, Iwai K, Sakurai M, Fukushi M, Oie M, Higuchi M, Hall WW, Gejyo F, Fujii M: Human T-cell leukemia virus type 2 (HTLV-2) Tax protein transforms a rat fibroblast cell line but less efficiently than HTLV-1 Tax. J Virol. 2002, 76: 2648-2653. 10.1128/JVI.76.6.2648-2653.2002.PubMed CentralView ArticlePubMedGoogle Scholar
- Hirata A, Higuchi M, Niinuma A, Ohashi M, Fukushi M, Oie M, Akiyama T, Tanaka Y, Gejyo F, Fujii M: PDZ domain-binding motif of human T-cell leukemia virus type 1 Tax oncoprotein augments the transforming activity in a rat fibroblast cell line. Virology. 2004, 318: 327-336. 10.1016/j.virol.2003.10.006.View ArticlePubMedGoogle Scholar
- Meertens L, Chevalier S, Weil R, Gessain A, Mahieux R: A 10-amino acid domain within human T-cell leukemia virus type 1 and type 2 tax protein sequences is responsible for their divergent subcellular distribution. J Biol Chem. 2004, 279: 43307-43320. 10.1074/jbc.M400497200.View ArticlePubMedGoogle Scholar
- Sieburg M, Tripp A, Ma JW, Feuer G: Human T-cell leukemia virus type 1 (HTLV-1) and HTLV-2 tax oncoproteins modulate cell cycle progression and apoptosis. J Virol. 2004, 78: 10399-10409. 10.1128/JVI.78.19.10399-10409.2004.PubMed CentralView ArticlePubMedGoogle Scholar
- Tsubata C, Higuchi M, Takahashi M, Oie M, Tanaka Y, Gejyo F, Fujii M: PDZ domain-binding motif of human T-cell leukemia virus type 1 Tax oncoprotein is essential for the interleukin 2 independent growth induction of a T-cell line. Retrovirology. 2005, 2: 46-10.1186/1742-4690-2-46.PubMed CentralView ArticlePubMedGoogle Scholar
- Tripp A, Banerjee P, Sieburg M, Planelles V, Li F, Feuer G: Induction of cell cycle arrest by human T-cell lymphotropic virus type 1 Tax in hematopoietic progenitor (CD34+) cells: modulation of p21cip1/waf1 and p27kip1 expression. J Virol. 2005, 79: 14069-14078. 10.1128/JVI.79.22.14069-14078.2005.PubMed CentralView ArticlePubMedGoogle Scholar
- Xie L, Yamamoto B, Haoudi A, Semmes OJ, Green PL: PDZ binding motif of HTLV-1 Tax promotes virus-mediated T-cell proliferation in vitro and persistence in vivo. Blood. 2006, 107: 1980-1988. 10.1182/blood-2005-03-1333.PubMed CentralView ArticlePubMedGoogle Scholar
- Sheehy N, Lillis L, Watters K, Lewis M, Gautier V, Hall W: Functional analysis of human T lymphotropic virus type 2 Tax proteins. Retrovirology. 2006, 3: 20-10.1186/1742-4690-3-20.PubMed CentralView ArticlePubMedGoogle Scholar
- Iwanaga Y, Tsukahara T, Ohashi T, Tanaka Y, Arai M, Nakamura M, Ohtani K, Koya Y, Kannagi M, Yamamoto N, Fujii M: Human T-cell leukemia virus type 1 tax protein abrogates interleukin-2 dependence in a mouse T-cell line. J Virol. 1999, 73: 1271-1277.PubMed CentralPubMedGoogle Scholar
- Niinuma A, Higuchi M, Takahashi M, Oie M, Tanaka Y, Gejyo F, Tanaka N, Sugamura K, Xie L, Green PL, Fujii M: Aberrant activation of the interleukin-2 autocrine loop through the nuclear factor of activated T cells by nonleukemogenic human T-cell leukemia virus type 2 but not by leukemogenic type 1 virus. J Virol. 2005, 79: 11925-11934. 10.1128/JVI.79.18.11925-11934.2005.PubMed CentralView ArticlePubMedGoogle Scholar
- Tanaka Y, Yoshida A, Tozawa H, Shida H, Nyunoya H, Shimotohno K: Production of a recombinant human T-cell leukemia virus type-I trans-activator (tax1) antigen and its utilization for generation of monoclonal antibodies against various epitopes on the tax1 antigen. Int J Cancer. 1991, 48: 623-630.View ArticlePubMedGoogle Scholar
- Lewis MJ, Novoa P, Ishak R, Ishak M, Salemi M, Vandamme AM, Kaplan MH, Hall WW: Isolation, cloning, and complete nucleotide sequence of a phenotypically distinct Brazilian isolate of human T-lymphotropic virus type II (HTLV-II). Virology. 2000, 271: 142-154. 10.1006/viro.2000.0284.View ArticlePubMedGoogle Scholar
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