Cell receptors for the mammalian reovirus: reovirus-specific T-cell hybridomas can become persistently infected and undergo autoimmune stimulation.

We have previously described the development of virus-specific helper T cell hybridomas which recognize structural determinants shared by type 1 and type 3 reoviruses that have been exposed to UV radiation. We have found that T-cell hybridomas become persistently infected with live type 3 reovirus used for the immunization. Persistently infected T-hybridoma cells were found to spontaneously produce interleukin 2 (IL-2). To analyze the mechanism of induction of IL-2 secretion of persistently infected T-cell hybridomas, we exposed T-cell hybridomas specific for UV-treated virus to replicating type 3 reovirus. The T-cell hybridomas became infected but did not produce IL-2 unless simultaneously exposed to syngeneic I-A+ antigen-presenting cells. In this situation, the persistently infected T-cell hybridomas produced IL-2 without being reexposed to virus. This process was not a consequence of nonspecific IL-2 gene activation, which occurs in cells persistently infected with reovirus, because reovirus infection did not activate IL-2 secretion in T-cell hybridomas with other antigenic specificities. Reovirus exposure also resulted in persistent infection of certain antigen-presenting B-cell tumor lines. The persistently infected B-cell tumor lines could stimulate reovirus-specific helper T cells but not T-cell hybridomas of other specificities. The data support the thesis that persistent infection of reovirus-specific T cells creates a mechanism in which the virus released from these cells is processed and then reexpressed by I-A+ antigen-presenting cells. The IA antigen and reovirus structures on the antigen-presenting cells then restimulate the T cells through their specific receptors, resulting in IL-2 synthesis and release. These observations may be relevant to mechanisms of autoimmunity induced by virus.     

Novel features of the respiratory tract T-cell response to influenza virus infection: lung T cells increase expression of gamma interferon mRNA in vivo and maintain high levels of mRNA expression for interleukin-5 (IL-5) and IL-10.

Analysis of the respiratory tract before and after primary influenza virus infection revealed a virus-induced preferential accumulation of a CD8+ T-cell population that coexpresses mRNA for interleukin-5 (IL-5) and IL-10 with virus dose-dependent high levels of gamma interferon. However, cytokine production in lung tissues was not restricted to the T-cell population, since CD3- cells were found to express mRNA for various cytokines, including IL-4 and particularly IL-6 and granulocyte-macrophage colony-stimulating factor. These data provide in vivo evidence for a local respiratory tract immune response to influenza virus infection dominated by cytokine-producing CD8+ T cells.     

Differential expression of interleukin-2 and gamma interferon in human immunodeficiency virus disease

Subnormal T-cell production of interleukin-2 (IL-2) in human immunodeficiency virus (HIV) disease has been described; however, it is not clear whether failure to synthesize IL-2 represents a selective or global defect in T-cell cytokine production. We evaluated the intracellular production of gamma interferon (IFN-gamma) and IL-2 in CD4(+) cells that were stimulated with staphylococcal enterotoxin B or cytomegalovirus antigen. Strikingly, IFN-gamma and IL-2 are differentially regulated in T cells of HIV-infected patients such that the numbers of CD69(+) cells or IFN-gamma-positive cells that make IL-2 are proportionally decreased in CD4(+) T cells from HIV-infected patients. These findings demonstrate a selective defect in IL-2 production and suggest that enumeration of IFN-gamma-producing cells in response to T-cell receptor stimulation, while providing some estimate of antigen-reactive cell frequency, may not reflect or predict "normal" T-cell function in HIV-infected patients.     

Polarization of allogeneic T-cell responses by influenza virus-infected dendritic cells

The developing immune response in the lymph nodes of mice infected with influenza virus has both Th1- and Th2-type characteristics. Modulation of the interactions between antigen-presenting cells and T cells is one mechanism that may alter the quality of the immune response. We have previously shown that the ability of dendritic cells (DC) to stimulate the proliferation of alloreactive T cells is changed by influenza virus due to viral neuraminidase (NA) activity. Here we show that DC infected with influenza virus A/PR/8/34 (PR8) stimulate T cells to produce different types of cytokines in a dose-dependent manner. Optimal amounts of the Th1-type cytokines interleukin-2 (IL-2) and gamma interferon (IFN-gamma) were produced from T cells stimulated by DC infected with low doses of PR8, while the Th2-type cytokines IL-4 and IL-10 were produced only in response to DC infected with high doses of PR8. IL-2 and IFN-gamma levels corresponded with T-cell proliferation and were dependent on the activity of viral NA on the DC surface. In contrast, IL-4 secretion required the treatment of T cells with NA. Since viral particles were released only from DC that are infected with high doses of PR8, our results suggest that viral NA on newly formed virus particles desialylates T-cell surface molecules to facilitate a Th2-type response. These results suggest that the activity of NA may contribute to the mixed Th-type response observed during influenza virus infection.     

Immunization with vaccinia virus recombinants that express the surface glycoproteins of human parainfluenza virus type 3 (PIV3) protects patas monkeys against PIV3 infection.

Patas monkeys (Eryphrocebus patas) were immunized intradermally with two vaccinia virus recombinants that individually express the hemagglutinin-neuraminidase glycoprotein or the fusion glycoprotein of human parainfluenza virus type 3 (PIV3). These immunizations induced a high titer of PIV3 serum-neutralizing antibodies. At 1 month after immunization, monkeys were challenged intratracheally with PIV3. Subsequent virus replication was reduced in these monkeys by 3.2 log10 and 1.9 log10 (mean peak virus titers) in the upper and lower respiratory tracts, respectively, compared with control animals. The average duration of virus shedding was also reduced from 9.0 to 3.4 days in the upper respiratory tract and from 5.3 to 1.2 days in the lower respiratory tract. These findings demonstrate that a single intradermal dose of live recombinant vaccinia viruses can significantly restrict the replication of a virus which primarily infects the epithelial cells of the respiratory tract.     

Caspase-dependent and -independent T-cell death pathways in pathogenic simian immunodeficiency virus infection: relationship to disease progression

Studies of human immunodeficiency virus (HIV) and nonhuman primate models of pathogenic and nonpathogenic simian immunodeficiency virus (SIV) infections have suggested that enhanced ex vivo CD4 T-cell death is a feature of pathogenic infection in vivo. However, the relative contributions of the extrinsic and intrinsic pathways to programmed T-cell death in SIV infection have not been studied. We report here that the spontaneous death rate of CD4+ T cells from pathogenic SIVmac251-infected rhesus macaques ex vivo is correlated with CD4 T-cell depletion and plasma viral load in vivo. CD4+ T cells from SIVmac251-infected macaques showed upregulation of the death ligand (CD95L) and of the proapoptotic proteins Bim and Bak, but not of Bax. Both CD4+ and CD8+ T cells from SIVmac251-infected macaques underwent caspase-dependent death following CD95 ligation. The spontaneous death of CD4+ and CD8+ T cells was not prevented by a decoy CD95 receptor or by a broad-spectrum caspase inhibitor (zVAD-fmk), suggesting that this form of cell death is independent of CD95/CD95L interaction and caspase activation. IL-2 and IL-15 prevented the spontaneous death of CD4+ and CD8+ T cells, whereas IL-10 prevented only CD8 T-cell death and IL-7 had no effect on T-cell death. Our results indicate that caspase-dependent and caspase-independent pathways are involved in the death of T cells in pathogenic SIVmac251-infected primates.     

Induction of CD4(+)CD25(+)Foxp3(-) regulatory T cells by Thy-1 stimulation of CD4(+) T cells

Thy-1 (CD90) on mouse T cells has been reported to have both T-cell activating and regulatory roles. In this study, we show that monoclonal antibody (mAb)-mediated crosslinking of Thy-1 on CD4(+) mouse T-cells-induced regulatory T (T(reg)) cells that expressed CD25, CD39 and glucocorticoid-induced tumor necrosis factor receptor family-related gene, but not CD73, CD122 or Foxp3. The proliferation of CD4(+) T(responder) cells in response to anti-CD3/anti-CD28mAb-coated T-cell expander beads or syngeneic dendritic cells and soluble anti-CD3mAb was inhibited by Thy-1-induced T(reg) cells, in spite of elevated IL-2 levels in the co-cultures. Interestingly, stimulation with T-cell expander beads caused Thy-1-induced T(reg) cells to synthesize large amounts of interleukin-2 (IL-2). IL-10 was also elevated in co-cultures of activated T(responder) cells and Thy-1-induced T(reg) cells. However, mAb-mediated neutralization of IL-10 did not restore T(responder)-cell proliferation to control levels, which excluded IL-10 as a potential mediator of Thy-1-induced T(reg)-cell suppressor function. In addition, Thy-1-induced T(reg) cells did not inhibit IL-2-dependent proliferation of CTLL-2 cells, suggesting that IL-2 receptor signaling remained intact in the presence of Thy-1-induced T(reg) cells. We suggest that T(reg) cells induced by Thy-1 ligation in vivo may contribute to the maintenance of T-cell homeostasis.     

T-cell activation is required for efficient replication of human herpesvirus 6.

We have investigated whether T-cell activation is required for the replication of the T-lymphotropic human herpesvirus 6. The virus did not replicate in quiescent peripheral blood lymphocytes but replicated efficiently following exposure of the cells to the polyclonal mitogen phytohemagglutinin (PHA). When purified T cells were treated with PHA in the absence of accessory cells, no virus replication was observed unless exogenous interleukin-2 (IL-2) was added to the medium, promoting cell division. Incubation of peripheral blood lymphocytes in the absence of PHA but in the presence of IL-2 resulted in delayed cell blastogenesis and virus replication. Cell blastogenesis and virus replication did not occur in the purified T-cell cultures incubated with IL-2 alone. Taken together, the results show that human herpesvirus 6 replication requires full progression of the cell cycle. This finding might have implications for the pathogenicity of the virus in the human host.     

Epithelial cells infected with respiratory syncytial virus are resistant to the anti-inflammatory effects of hydrocortisone

In this work we continue our study of the biochemical responses of respiratory epithelial cells to infection with human paramyxovirus pathogens. In our earlier studies, we detected elevated concentrations of the proinflammatory chemokines MIP-1alpha and IL-8 in upper and lower respiratory tract secretions from patients infected with respiratory syncytial virus (RSV). Here we demonstrate the same trend for individuals infected with parainfluenza virus (PIV), with elevated concentrations of MIP-1alpha and IL-8 (means of 309 +/- 51 and 2280 +/- 440 pg/ml/mg protein, respectively) detected in nasal wash samples from 17 patients with culture-positive PIV. Similar to our findings with RSV, cells of the HEp-2 epithelial line and primary cultures of human bronchial epithelial cells respond to PIV infection with production and release of both MIP-1alpha and IL-8. Addition of the glucocorticoid anti-inflammatory agent hydrocortisone (200-1000 ng/ml) attenuated the production of MIP-1alpha and IL-8 in PIV-infected cells while having minimal to no effect on the production of these mediators from cells infected with RSV. Neither virus infection resulted in a change in the total cellular concentration of glucocorticoid receptors, nor did hydrocortisone exert any differential effect on viral replication. As repression of chemokine production by epithelial cells is likely to result in diminished recruitment of proinflammatory leukocytes, these results may explain in part why glucocorticoid therapy reduces the symptoms associated with acute PIV infection, but have little to no effect in the overall outcome in the case of RSV.     

Prostaglandin E2 inhibits human T-cell proliferation after crosslinking of the CD3-Ti complex by directly affecting T cells at an early step of the activation process

We have studied the effects of prostaglandin E2 (PGE2) on in vitro human T-cell activation induced by crosslinking of the CD3-Ti complex with the monoclonal anti-CD3 antibodies OKT3 and UCHT-1. PGE2 (greater than or equal to 3 X 10(-9) M) when added simultaneously with anti-CD3 to cultures of peripheral blood mononuclear cells (PBMC), significantly suppressed, in a dose-dependent way, T-cell proliferation (P less than 0.002). However, when T cells were first preactivated with OKT3 for 3 days, subsequent proliferation driven by recombinant interleukin 2 (IL-2) was not inhibited by addition of PGE2. This indicates that PGE2 affects the activation step resulting from crosslinking of CD3-Ti, but not the IL-2-driven proliferative phase. Other manifestations of T-cell activation were therefore examined. Both IL-2 production and the expression of receptors for IL-2 (as detected with the anti-Tac monoclonal antibody) were inhibited by PGE2. The addition of purified interleukin 1 (IL-1) or recombinant IL-2 to the cultures did not reverse the inhibiting effect of PGE2 on IL-2-receptor expression. PGE2, added at the time of culture initiation, also inhibited T-cell proliferation in cultures which were supplemented with exogenous IL-1 or IL-2. Proof for a direct effect of PGE2 on T cells was obtained in experiments in which monocyte-depleted T cells were stimulated, in the presence of IL-1, with solid-phase-bound anti-CD3 antibody. Proliferation of T cells in this system is accessory cell independent and still was strongly inhibited by PGE2. Finally, preincubation of PBMC with PGE2 (3 X 10(-6) M) for 48 hr did not result in the generation of suppressor cells for anti-CD3-induced T-cell proliferation or for IL-2 production. Our results demonstrate that PGE2 has a direct inhibitory effect on an early step of T-cell activation, resulting in decreased IL-2 production, decreased IL-2-receptor expression, decreased responsiveness to exogenous IL-2, and decreased proliferation. However, PGE2 does not affect IL-2-driven proliferation of activated T cells. The inhibitory effect on T-cell activation is not mediated through suppressor T cells, nor through inhibition of accessory cell function.     

Leishmania amazonensis: participation of regulatory T and B cells in the in vitro priming (PIV) of CBA/J spleen cells susceptible response

CBA/J mice are resistant to Leishmania major and susceptible to Leishmania amazonensis. Early events determine infection outcome. Until now, PIV (in vitro priming) immune response to L. amazonensis has not been assessed. Herein, we have shown that compared to L. major, L. amazonensis induced higher parasite burden associated to similar IL-4, IFN-gamma, and TNF-alpha mRNA expressions and IFN-gamma and IL-10 levels. Although similar amounts of IL-10 were detected, the frequency of intracellular IL-10 positive B cells was enhanced in spleen cells stimulated with anti-CD3/anti-CD28, or anti-CD3/anti-CD28 and L. amazonensis, compared to L. major-stimulation. Interestingly, IL-10- producing B cells were reduced in response to anti-CD3/anti-CD28 stimulation combined with L. major compared to the other groups. L. amazonensis may favor T regulatory cell development, since 40% of all the CD4+CD25+ were CD25(high) cells. These data suggest that in PIV, susceptibility to L. amazonensis is not related to Th cell polarization, but to the presence and activity of regulatory T and B cells.     

Alpha and lambda interferon together mediate suppression of CD4 T cells induced by respiratory syncytial virus

The mechanism by which respiratory syncytial virus (RSV) suppresses T-cell proliferation to itself and other antigens is poorly understood. We used monocyte-derived dendritic cells (MDDC) and CD4 T cells and measured [(3)H]thymidine incorporation to determine the factors responsible for RSV-induced T-cell suppression. These two cell types were sufficient for RSV-induced suppression of T-cell proliferation in response to cytomegalovirus or Staphylococcus enterotoxin B. Suppressive activity was transferable with supernatants from RSV-infected MDDC and was not due to transfer of live virus or RSV F (fusion) protein. Supernatants from RSV-infected MDDC, but not MDDC exposed to UV-killed RSV or mock conditions, contained alpha interferon (IFN-alpha; median, 43 pg/ml) and IFN-lambda (approximately 1 to 20 ng/ml). Neutralization of IFN-alpha with monoclonal antibody (MAb) against one of its receptor chains, IFNAR2, or of IFN-lambda with MAb against either of its receptor chains, IFN-lambdaR1 (interleukin 28R [IL-28R]) or IL-10R2, had a modest effect. In contrast, blocking the two receptors together markedly reduced or completely blocked the RSV-induced suppression of CD4 T-cell proliferation. Defining the mechanism of RSV-induced suppression may guide vaccine design and provide insight into previously uncharacterized human T-cell responses and activities of interferons.     

Interleukin-10 determines viral clearance or persistence in vivo

Persistent viral infections are a major health concern. One obstacle inhibiting the clearance of persistent infections is functional inactivation of antiviral T cells. Although such immunosuppression occurs rapidly after infection, the mechanisms that induce the loss of T-cell activity and promote viral persistence are unknown. Herein we document that persistent viral infection in mice results in a significant upregulation of interleukin (IL)-10 by antigen-presenting cells, leading to impaired T-cell responses. Genetic removal of Il10 resulted in the maintenance of robust effector T-cell responses, the rapid elimination of virus and the development of antiviral memory T-cell responses. Therapeutic administration of an antibody that blocks the IL-10 receptor restored T-cell function and eliminated viral infection. Thus, we identify a single molecule that directly induces immunosuppression leading to viral persistence and demonstrate that a therapy to neutralize IL-10 results in T-cell recovery and the prevention of viral persistence.     

Changes in paracrine interleukin-2 requirement, CCR7 expression, frequency, and cytokine secretion of human immunodeficiency virus-specific CD4+ T cells are a consequence of antigen load

Virus-specific CD4+ T-cell responses are thought to be required for the induction and maintenance of many effective CD8+ T-cell and B-cell immune responses in experimental animals and humans. Although the presence of human immunodeficiency virus (HIV)-specific CD4+ T cells has been documented in patients at all stages of HIV infection, many fundamental questions regarding their frequency and function remain. A 10-color, 12-parameter flow cytometric panel was utilized to examine the frequency, memory phenotype (CD27, CCR7, and CD45RA), and cytokine production (interleukin-2 [IL-2], gamma interferon, and tumor necrosis factor alpha) of CD4+ T cells specific for HIV antigens as well as for adenovirus, Epstein-Barr virus (EBV), influenza H1N1 virus, influenza H3N2 virus, cytomegalovirus, varicella-zoster virus (VZV), and tetanus toxoid in normal controls, long-term nonprogressors (LTNP), and HIV-infected patients with progressive disease on or off therapy. The HIV-specific CD4+ T-cell responses in LTNP and patients on therapy were similar in frequency, phenotype, and cytokine production to responses directed against adenovirus, EBV, influenza virus, and VZV. HIV-specific CD4+ T cells from patients off antiretroviral therapy demonstrated a shift towards a CCR7(-) CD45RA(-) phenotype and a reduced percentage of IL-2-producing cells. The alterations in cytokine production during HIV viremia were found to be intrinsic to the HIV-specific CD4+ T cells and caused a requirement for IL-2 supplied exogenously for proliferation to occur. These observations suggest that many previously described changes in HIV-specific CD4+ T-cell function and phenotype are a consequence of high levels of antigen in viremic patients. In addition, defects in function and phenotype of HIV-specific CD4+ T cells are not readily discernible in the context of antiretroviral therapy but rather are similar to responses to other viruses.     

IL-15 maintains T-cell survival via S-nitrosylation-mediated inhibition of caspase-3

Caspase activity is critical for both T-cell survival and death. However, little is known regarding what determines caspase activity in cycling T cells. Interleukin (IL)-2 and IL-15 confer very different susceptibilities to T-cell death. We therefore considered that IL-2 and IL-15 differentially regulate caspase activity to influence T-cell survival. We observed that IL-2-cultured primary murine effector T cells manifested elevated levels of caspase-3 activity compared with IL-15-cultured T cells. T cell receptor (TCR) restimulation further increased caspase activity and induced considerable cell death in IL-2-cultured T cells, but provoked only a minimal increase of caspase activity and cell death in IL-15-cultured T cells. IL-2 sensitization to cell death was caspase-3 mediated. Interestingly, increased active caspase-3 levels with IL-2 were independent of active initiator caspase-8 and caspase-9 that were similar with IL-2 and IL-15. Rather, caspase-3 activity was inhibited by posttranslational S-nitrosylation in IL-15-cultured T cells, but not in the presence of IL-2. This paralleled increased reactive nitrogen and oxygen species with IL-15 and reduced glycolysis. Taken together, these data suggest that the metabolic state conferred by IL-15 inhibits T-cell apoptosis in part by maintaining low levels of active caspase-3 via S-nitrosylation.     

Programmed cell death of T lymphocytes during acute viral infection: a mechanism for virus-induced immune deficiency.

Acute viral infections induce immune deficiencies, as shown by unresponsiveness to mitogens and unrelated antigens. T lymphocytes isolated from mice acutely infected with lymphocytic choriomeningitis virus (LCMV) were found in this study to undergo activation-induced apoptosis upon signalling through the T-cell receptor (TcR)-CD3 complex. Kinetic studies demonstrated that this sensitivity to apoptosis directly correlated with the induction of immune deficiency, as measured by impaired proliferation in response to anti-CD3 antibody or to concanavalin A. Cell cycling in interleukin-2 (IL-2) alone stimulated proliferation of LCMV-induced T cells without inducing apoptosis, but preculturing of T cells from acutely infected mice in IL-2 accelerated apoptosis upon subsequent TcR-CD3 cross-linking. T lymphocytes isolated from mice after the acute infection were less responsive to IL-2, but those T cells, presumably memory T cells, responding to IL-2 were primed in each case to die a rapid apoptotic death upon TcR-CD3 cross-linking. These results indicate that virus infection-induced unresponsiveness to T-cell mitogens is due to apoptosis of the activated lymphocytes and suggest that the sensitization of memory cells by IL-2 induced during infection will cause them to die upon antigen recognition, thereby impairing specific responses to nonviral antigens.     

HTLV-1 bZIP Factor Impairs Anti-viral Immunity by Inducing Co-inhibitory Molecule,T Cell Immunoglobulin and ITIM Domain (TIGIT)

Human T-cell leukemia virus type 1 (HTLV-1) infects CD4⁺ T cells and induces proliferation of infected cells in vivo, which leads to the onset of adult T-cell leukemia (ATL) in some infected individuals. The HTLV-1 bZIP factor (HBZ) gene, which is encoded in the minus strand of HTLV-1, plays critical roles in pathogenesis. In this study, RNA-seq and ChIP-seq analyses using HBZ transduced T cells revealed that HBZ upregulates the expression and promoter acetylation levels of a co-inhibitory molecule, T cell immunoglobulin and ITIM domain (TIGIT), in addition to those of regulatory T cells related genes, Foxp3 and Ccr4. TIGIT was expressed on CD4⁺ T cells from HBZ-transgenic (HBZ-Tg) mice, and on ATL cells and HTLV-1 infected CD4⁺ T cells of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) in vivo. Expression of Blimp1 and IL-10 was upregulated in TIGIT⁺CD4⁺ cells of HBZ-Tg mice compared with TIGIT^-CD4⁺ T cells, suggesting the correlation between TIGIT expression and IL-10 production. When CD4⁺ T cells from HBZ-Tg mice were stimulated with TIGIT’s ligand, CD155, their production of the inhibitory cytokine IL-10 was enhanced. Furthermore, dendritic cells from HBZ-Tg mice produced high levels of IL-10 after stimulation. These data suggest that HBZ alters immune system to suppressive state via TIGIT and IL-10. Importantly, TIGIT suppressed T-cell responses to another HTLV-1 virus protein, Tax, in vitro. Blocking of TIGIT and PD-1 slightly increased anti-Tax T-cell activity in some HAM/TSP patients. These results suggest that HBZ-induced TIGIT on HTLV-1 infected cells impairs T-cell responses to viral antigens. This study shows that HBZ-induced TIGIT plays a pivotal role in attenuating host immune responses and shaping a microenvironment favorable to HTLV-1.     

Induction of self-reactive T cells after murine coronavirus infection.

We studied the mechanism of in vitro spontaneous lymphokine production by spleen cells from mice injected intraperitoneally with murine coronavirus stain JHM 1 month after infection, when infectious virus had already been cleared from the spleens. Removal of either CD4+ T cells or Ia+ antigen-presenting cells (APC) from the spleen cells abrogated interleukin-2 (IL-2) production. Addition of anti-CD4 or anti-Iad monoclonal antibodies to the culture suppressed IL-2 production. These results suggest that the response involved typical receptor-mediated activation of T cells. Surprisingly, reciprocal mixing experiments with a coculture of T cells from infected mice and APC from either infected or naive mice resulted in the production of IL-2. The absence of viral antigens in spleen cells 1 month after infection, as indicated by their inability to induce the proliferation of T-cell clones specific for the viral antigens, suggest that the T cells from mice 1 month after infection were not responding to the viral antigens. The inoculum components other than the virus did not induce this immune response. We also found that the frequency of self-reactive but not alloreactive IL-2-producing T cells in the spleens of infected mice was 3- to 10-fold higher than that in naive mice. These findings suggest that an increased frequency of self-reactive T cells which secrete IL-2 occurs following murine coronavirus infection. This may have important implications in the development of autoimmunelike phenomena following murine coronavirus infection.     

Proliferation Response to Interleukin-2 and Jak/Stat Activation of T Cells Immortalized by Human T-Cell Lymphotropic Virus Type 1 Is Independent of Open Reading Frame I Expression

Human T-cell lymphotropic virus type 1 (HTLV-1), a complex retrovirus, encodes a hydrophobic 12-kD protein from pX open reading frame (ORF) I that localizes to cellular endomembranes and contains four minimal SH3 binding motifs (PXXP). We have demonstrated the importance of ORF I expression in the establishment of infection and hypothesize that p12(I) has a role in T-cell activation. In this study, we tested interleukin-2 (IL-2) receptor expression, IL-2-mediated proliferation, and Jak/Stat activation in T-cell lines immortalized with either wild-type or ORF I mutant clones of HTLV-1. All cell lines exhibited typical patterns of T-cell markers and maintained mutation fidelity. No significant differences between cell lines were observed in IL-2 receptor chain (alpha, beta, or gamma(c)) expression, in IL-2-mediated proliferation, or in IL-2-induced phosphorylated forms of Stat3, Stat5, Jak1, or Jak3. The expression of ORF I is more likely to play a role in early HTLV-1 infection, such as in the activation of quiescent T cells in vivo.