Human T-cell lymphotropic virus type 1 Tax mediates enhanced transcription in CD4+ T lymphocytes

Abstract

Human T-cell leukemia virus type 1 (HTLV-1) is the etiologic agent of adult T-cell leukemia/lymphoma and is associated with a variety of immunoregulatory disorders. HTLV-1 has been shown to bind to and infect a variety of hematopoietic and nonhematopoietic cells. However, both in vivo and in vitro, the provirus is mostly detected in and preferentially transforms CD4 ؉ T cells. The molecular mechanism that determines the CD4 ؉ T-cell tropism of HTLV-1 has not been determined. Using cocultures of purified CD4 ؉ and CD8 ؉ T cells with an HTLV-1-producing cell line, we measured viral transcription by using Northern (RNA) blot analysis, protein production by using a p24 antigen capture assay and flow cytometric analysis for viral envelope, and proviral integration by using DNA slot blot analysis. We further measured HTLV-1 long terminal repeatdirected transcription in purified CD4 ؉ and CD8 ؉ T cells by using transient transfection assays and in vitro transcription. We demonstrate a higher rate of viral transcription in primary CD4 ؉ T cells than in CD8 ؉ T cells. HTLV-1 protein production was 5-to 25-fold greater in CD4 ؉ cocultures and mRNA levels were 5-fold greater in these cultures than in the CD8 ؉ cocultures. Transient transfection and in vitro transcription indicated a modest increase in basal transcription in CD4 ؉ T cells, whereas there was a 20-fold increase in reporter gene activity in CD4 ؉ T cells cotransfected with tax. These data suggest that unique or activated transcription factors, particularly Tax-responsive factors in CD4 ؉ T cells, recognize regulatory sequences within the HTLV-1 long terminal repeat, and this mediates the observed enhanced viral transcription and ultimately the cell tropism and leukemogenic potential of the virus.

Repression of tax-mediated human t-lymphotropic virus type 1 transcription by inducible cAMP early repressor (ICER) protein in peripheral blood mononuclear Cells

“…To obtain resting cells, PBMC were cultured for 48 hr in RPMI 1640 supplemented with 10% fetal bovine serum, 0.3 mg/ml Lglutamine, 100 U/ml penicillin and 100 mg/ml streptomycin (complete medium) at 37°C in 7% CO 2 . Cells were stimulated using 5 U/ml rIL2 (Boehringer Mannheim Biochemical, Indianapolis, IN) and 2.5 g/ml phytohemagglutinin P (PHA P) (activation medium) and were maintained in stimulation medium for the indicated times [Newbound et al, 1996].…”

“…For the RNase protection assay a plasmid called pCRJL5, containing the full length human ICER II gene, was a generous gift from Dr. J. Habener and has been previously described [Bodor et al, 1996]. For transient transfections a HTLV-1 LTR and LTR-Tax plasmids have been previously described [Newbound et al, 1996]. A CMV-driven ICER II␥ plasmid was a generous gift from Dr. C. Tinti and has been previously described [Tinti et al, 1996].…”

“…pRC/RSV and pcDNA3 (Invitrogen) were transfected as empty vector controls for RSV and CMV containing plasmids respectively. We have previously published our methods to determine transfection efficiency in primary lymphocytes [Newbound et al, 1996]. Transfections were carried out in triplicate and the results shown are representative of at least four independent trials.…”

Human T cell leukaemia virus type 2 tax protein mediates CC-chemokine expression in peripheral blood mononuclear cells via the nuclear factor kappa B canonical pathway

“…The efficiency of transduction was confirmed by fluorescence microscopy. A luciferase reporter assay was used as described previously [24,32] to confirm the ability of the recombinant Tax2 proteins to regulate viral transcription.…”

The human T-cell leukemia virus type 1 Rex regulatory protein exhibits an impaired functionality in human lymphoblastoid Jurkat T cells

“…Such observations indicate that the viral receptor is present on many cells types, as con-firmed by binding studies and pseudotype interference assays. They in turn indicate that the in vivo CD4 ϩ T-cell tropism of HTLV-1 is determined mainly at a postpenetration step of the retrovirus life cycle (34). In contrast to the wide host range of cells susceptible to HTLV-1 infection, we have repeatedly observed that Jurkat or CEM T cells were unable to sustain a productive infection after coculture with irradiated HTLV-1producing cells.…”