A novel membrane protein is a mouse mammary tumor virus receptor

Mouse mammary tumor virus (MMTV) infects a number of different cell types, including mammary gland and lymphoid cells, in vivo. To identify the cellular receptor for this virus, a mouse cDNA expression library was transfected into Cos-7 monkey kidney cells, and those transfected cells able to bind virus were selected by using antibody against the virus's cell surface envelope protein, gp52. One clone isolated from a library prepared from newborn thymus RNA, called MTVR, was able to confer virus binding to both monkey and human cells; this binding was blocked by anti-MTVR antibody. Moreover, transfection of MTVR into CV1 cells rendered them susceptible to infection by a murine leukemia virus-based retrovirus vector pseudotyped with the MMTV envelope protein. An epitope-tagged MTVR cofractionated with cellular membranes. Coimmunoprecipitation of the MMTV envelope protein and a MTVR-rabbit Fc fusion protein showed that these two proteins bound to each other. The MTVR sequence clone is unique, shows no homology to known membrane proteins, and is transcribed in many tissues.     

The amino-terminal region of Tyk2 sustains the level of interferon alpha receptor 1, a component of the interferon alpha/beta receptor

Tyk2 belongs to the Janus kinase (JAK) family of receptor associated tyrosine kinases, characterized by a large N-terminal region, a kinase-like domain and a tyrosine kinase domain. It was previously shown that Tyk2 contributes to interferon-alpha (IFN-alpha) signaling not only catalytically, but also as an essential intracellular component of the receptor complex, being required for high affinity binding of IFN-alpha. For this function the tyrosine kinase domain was found to be dispensable. Here, it is shown that mutant cells lacking Tyk2 have significantly reduced IFN-alpha receptor 1 (IFNAR1) protein level, whereas the mRNA level is unaltered. Expression of the N-terminal region of Tyk2 in these cells reconstituted wild-type IFNAR1 level, but did not restore the binding activity of the receptor. Studies of mutant Tyk2 forms deleted at the N terminus indicated that the integrity of the N-terminal region is required to sustain IFNAR1. These studies also showed that the N-terminal region does not directly modulate the basal autophosphorylation activity of Tyk2, but it is required for efficient in vitro IFNAR1 phosphorylation and for rendering the enzyme activatable by IFN-alpha. Overall, these results indicate that distinct Tyk2 domains provide different functions to the receptor complex: the N-terminal region sustains IFNAR1 level, whereas the kinase-like domain provides a function toward high affinity ligand binding.     

Protective role of gamma interferon during the recall response to influenza virus

During secondary immune responses to influenza virus, virus-specific T memory cells are a major source of gamma interferon (IFN-gamma). We assessed the contribution of IFN-gamma to heterologous protection against the A/WSN/33 (H1N1) virus of wild-type and IFN-gamma-/- mice previously immunized with the A/HK/68 (H3N2) virus. The IFN-gamma-/- mice displayed significantly reduced survival rates subsequent to a challenge with various doses of the A/WSN/33 virus. This was associated with an impaired ability of the IFN-gamma-/- mice to completely clear the pulmonary virus by day 7 after the challenge, although significant reduction of the virus titers was noted. However, the IFN-gamma-/- mice developed type A influenza virus cross-reactive cytotoxic T lymphocytes (CTLs) similar to the wild-type mice, as demonstrated by both cytotoxicity and a limiting-dilution assay for the estimation of CTL precursor frequency. The pulmonary recruitment of T cells in IFN-gamma-/- mice was not dramatically affected, and the percentage of CD4(+) and CD8(+) T cells was similar to that of wild-type mice. The T cells from IFN-gamma-/- mice did not display a significant switch toward a Th2 profile. Furthermore, the IFN-gamma-/- mice retained the ability to mount significant titers of WSN and HK virus-specific hemagglutination-inhibiting antibodies. Together, these results are consistent with a protective role of IFN-gamma during the heterologous response against influenza virus independently of the generation and local recruitment of cross-reactive CTLs.     

Oligodendroglial response to the immune cytokine interferon gamma

The occurrence and coexistence of peptides of the insulin-like growth factor (IGF)/insulin superfamily were investigated in the ovary and gastro-intestinal tract of the protochordate Ciona intestinalis. Antisera specific for mammalian IGF-I, insulin and relaxin were used in a double-immunofluorescence method on paraffin sections and with an immunogold technique on consecutive semi-thin sections. IGF-I and relaxin immunoreactions but no insulin immunoreactions occurred in the ovary and were confined to medium-sized and mature follicle cells. Two subpopulations of reacting follicular cells were present: those containing only IGF-I immunoreactivity (5%) and those containing IGF-I and relaxin immunoreactivities (95%). In the gastro-intestinal tract, IGF-I and insulin immunoreactions coexisted, whereas no relaxin immunoreactions were obtained. Gel chromatography and radioimmunoassay in Ciona ovary revealed IGF-I immunoreactivity in two peaks with apparent molecular masses of approximately 16 kDa and 3 kDa. The present results indicate that (1) the same IGF-I-related peptide probably occurs in gastro-intestinal tract and ovary, (2) three different members of the insulin/IGF family of peptides are probably present in protochordates, (3) different types of coexistence of these peptides seem to exist in protochordates, i.e. an IGF-I-related peptide and an insulin-related peptide in the digestive tract and, as shown previously, in central nervous system, and the IGF-I-related peptide and relaxin in the ovary, (4) an IGF-I-related peptide and relaxin may be involved in oocyte maturation in the protochordate ovary.     

Immune response in mice that lack the interferon-gamma receptor

Interferon-gamma (IFN-gamma) exerts pleiotropic effects, including antiviral activity, stimulation of macrophages and natural killer cells, and increased expression of major histocompatibility complex antigens. Mice without the IFN-gamma receptor had no overt anomalies, and their immune system appeared to develop normally. However, mutant mice had a defective natural resistance, they had increased susceptibility to infection by Listeria monocytogenes and vaccinia virus despite normal cytotoxic and T helper cell responses. Immunoglobulin isotype analysis revealed that IFN-gamma is necessary for a normal antigen-specific immunoglobulin G2a response. These mutant mice offer the possibility for the further elucidation of IFN-gamma-mediated functions by transgenic cell- or tissue-specific reconstitution of a functional receptor.     

Differences in interferon alpha and beta signaling. Interferon beta selectively induces the interaction of the alpha and betaL subunits of the type I interferon receptor

All Type I interferons (IFNalpha, IFNbeta, IFNomega) bind to the Type I IFN receptor (IFNR) and elicit a common set of signaling events, including activation of the Jak/Stat and IRS pathways. However, IFNbeta selectively induces the association of the alpha subunit of the Type I IFNR with p100, a tyrosyl phosphoprotein, to transduce IFNbeta-specific signals. Using antibodies raised against the different components of the Type I IFNR, we identified p100 as the long form of the beta subunit (betaL subunit) of the Type I IFNR. This was also confirmed in experiments with mouse L-929 cells transfected with truncated forms of betaL. Thus, IFNbeta stimulation of human cells or mouse L-929 transfectants expressing the human alpha and betaL subunits, selectively induces the formation of a signaling complex containing the alpha and betaL subunits of the receptor. The IFNbeta-regulated interaction of the alpha and betaL chains is rapid and transient and follows a similar time course with the tyrosine phosphorylation of these receptor components. These data demonstrate that the signaling specificity for different Type I IFNs is established early in the signaling cascade, at the receptor level, and results from distinct interactions between components of the Type I IFNR.     

Generation of nitric oxide and induction of major histocompatibility complex class II antigen in macrophages from mice lacking the interferon gamma receptor

Availability of mice with a targeted disruption of the interferon gamma (IFN-gamma) receptor gene (IFN-gamma R0/0 mice) made it possible to examine parameters of macrophage activation in the absence of a functional IFN-gamma receptor. We asked to what extent other cytokines could replace IFN-gamma in the induction of nitric oxide or major histocompatibility complex class II antigen (Ia) expression in peritoneal macrophages. In thioglycollate-elicited macrophages from wild-type mice, tumor necrosis factor (TNF) alone was virtually ineffective in inducing release of NO2- (the endproduct of nitric oxide generation), but TNF enhanced NO2- release in the presence of IFN-gamma. In macrophages from IFN-gamma R0/0 mice, which were unresponsive to IFN-gamma, TNF completely failed to stimulate NO2- release. The stimulatory actions of IFN-alpha/beta on NO2- release were indistinguishable in wild-type and IFN-gamma R0/0 macrophages: IFN-alpha/beta was ineffective on its own, showed marginal stimulation of NO2- release in combination with TNF, and was moderately effective in the presence of lipopolysaccharide. The level of constitutive Ia antigen expression was not significantly different in peritoneal macrophages from wild-type and IFN-gamma R0/0 mice. An increased Ia expression was induced by IL-4 and granulocyte-macrophage colony-stimulating factor in both wild-type and IFN-gamma R0/0 macrophages, but the magnitude of this induction was less than with optimal concentrations of IFN-gamma in macrophages from wild-type mice. IFN-alpha/beta showed only a minor stimulatory effect on Ia expression in both wild-type and IFN-gamma R0/0 macrophages. Simultaneous treatment of wild-type macrophages with IFN-alpha/beta and IFN-gamma reduced the IFN-gamma-induced Ia expression in wild-type macrophages, but IFN-alpha/beta did not show an inhibitory effect on IL-4- or granulocyte-macrophage-colony-stimulating factor-induced Ia expression in either wild-type or IFN-gamma R0/0 macrophages. The important role of IFN-gamma in the regulation of the induced expression of major histocompatibility complex class II antigen was confirmed by showing that after systemic infection with the BCG strain of Mycobacterium bovis resident peritoneal macrophages from IFN-gamma R0/0 mice had a lower level of Ia expression than macrophages from wild-type mice. The inability of other cytokines to substitute fully for IFN-gamma in macrophage activation helps to explain the earlier observed decreased resistance of IFN-gamma R0/0 mice to some infections.     

The critical role of interleukin 4 but not interferon gamma in the pathogenesis of colitis in T-cell receptor alpha mutant mice

PURPOSE: To compare keratocyte density determined by using confocal microscopy with keratocyte density determined in the same corneas by histology. METHODS: Digital en face images of central corneas were recorded three times by using confocal microscopy in vivo in six New Zealand White rabbits. Bright objects (keratocyte nuclei) in the images were automatically identified by using a custom algorithm to estimate total and regional stromal keratocyte densities. The corneas were then excised, fixed, and sectioned in a sagittal plane for histology. Keratocyte nuclei were manually counted from digitized images of 50 histologic sections per cornea. Total and regional keratocyte densities were estimated from the histologic sections by using stereologic methods based on nuclei per unit area, mean nuclear diameter, and section thickness. Histologic cell densities were corrected for tissue shrinkage. RESULTS: By confocal microscopy, total keratocyte density was 39,000 +/- 1,200 cells/mm3 (mean +/- SE; n = 6); cell density was 47,100 +/- 1,300 cells/mm3 in the anterior stroma and decreased to 27,900 +/- 2,700 cells/mm3 in the posterior stroma (P = 0.004). Analysis of the three separate confocal images of each cornea produced repeatable total cell densities (mean coefficient of variation = 0.035). By histology, total keratocyte density was 37,800 +/- 1,100 cells/mm3, not significantly different from that estimated by confocal microscopy (P = 0.43); anterior cell density was 48,300 +/- 900 cells/mm3 and decreased to 29,400 +/- 900 cells/mm3 posteriorly (P < 0.001). CONCLUSIONS: Rabbit keratocyte density estimated by automated analysis of confocal microscopy images in vivo is repeatable and agrees with keratocyte density estimated from histologic sections.     

The neonatal Fc receptor is not required for mucosal infection by mouse mammary tumor virus

The milk-borne mouse mammary tumor virus (MMTV) infects newborn mice via the intestine. Infection is initially restricted to Peyer's patches and later spreads to the epithelial cells of the mammary gland. The receptor that mediates uptake and transport of MMTV across the intestinal barrier has not yet been identified, The neonatal Fc receptor (nFcR), which is expressed by enterocytes during the first two weeks of life, is downregulated at weaning, and its disappearance correlates with the onset of intestinal resistance to MMTV. To test whether the nFcR mediates transport and allows infection, we foster nursed on infected MMTV mothers beta2 microglobulin-deficient (beta2m-deficient) newborn mice that are unable to express the nFcR at the surface of their enterocytes. Exposure of beta2m-deficient mice to milk-borne virus resulted in the deletion of peripheral blood T cells reactive to the superantigen encoded by MMTV. Since beta2m-deficient newborn mice are susceptible to MMTV infection despite the lack of the nFcR, we conclude that the nFcR is not required for MMTV transport.     

Interferon beta, a cofactor in the interferon gamma production induced by gram-negative bacteria in mice

The interferon (IFN) gamma production of splenocytes from closely related C57BL/10ScSn (Sn) and C57BL/10ScCr (Cr) mice was compared. Concanavalin A and CD3 monoclonal antibodies induced high levels of IFN-gamma in both Sn and Cr splenocytes. By contrast, treatment with gram-negative bacteria induced IFN-gamma only in Sn splenocytes; in Cr splenocytes, the IFN-gamma response was heavily impaired. The IFN-gamma induction by bacteria requires the cooperation of IFN-gamma-producing cells with macrophages. Depletion of macrophages from Sn splenocytes resulted in the loss of ability to produce IFN-gamma after bacterial stimulation. Reconstitution with new Sn macrophages restored the IFN-gamma responsiveness, whereas reconstitution with Cr macrophages failed to do so. Normal function of IFN-gamma-producing cells and a defective function of macrophages of Cr mice was demonstrated by evidence showing that whole or macrophage-depleted Cr splenocytes, when supplemented with Sn macrophages, acquire the ability to produce IFN-gamma in response to bacteria. A similar effect was achieved by supplementing Cr splenocytes with supernatants of bacteria-stimulated Sn macrophages or with recombinant murine IFN-beta or IFN-alpha. Preincubation of active macrophage supernatants with antibodies to IFN-beta suppressed the helper activity for Cr splenocytes. Moreover, the bacteria-induced production of IFN-gamma by Sn splenocytes could be inhibited by antibodies to murine IFN-beta. The results provide evidence that IFN-beta is an important cofactor of IFN-gamma induction, which is not induced in Cr mice by gram-negative bacteria.     

A novel V beta 2-specific endogenous mouse mammary tumor virus which is capable of producing a milk-borne exogenous virus

We have previously reported new Mtv loci, Mtv-48 and -51, in the Japanese laboratory mouse strains CS and NC. Here we show by backcross analysis that both Mtv-48 and -51 cosegregate with very slow deletion of T cells bearing V beta 2. The nucleotide sequences of the open reading frames in the 3' long terminal repeats of Mtv-48 and -51 were very similar to those of Mtv-DDO, mouse mammary tumor virus C4 [MMTV(C4)], and MMTV(BALB/cV), which encode V beta 2-specific superantigens. Furthermore, backcross female mice carrying Mtv-48 but not Mtv-51 were found to be able to produce milk-borne MMTV(CS), which can vigorously stimulate V beta 2-expressing T cells after local injection in vivo in an I-E-dependent manner. On the other hand, mice carrying Mtv-51 but not Mtv-48 could not produce such an MMTV in milk. The nucleotide sequences of MMTV(CS) open reading frame were completely matched with those of Mtv-48. These results indicate that the provirus Mtv-48 but not Mtv-51 is capable of producing a milk-borne virus of which the superantigen stimulates V beta 2-expressing T cells.     

Mouse mammary tumor virus/v-Ha-ras transgene-induced mammary tumors exhibit strain-specific allelic loss on mouse chromosome 4

Hybrid mice carrying oncogenic transgenes afford powerful systems for investigating loss of heterozygosity (LOH) in tumors. Here, we apply this approach to a neoplasm of key importance in human medicine: mammary carcinoma. We performed a whole genome search for LOH using the mouse mammary tumor virus/v-Ha-ras mammary carcinoma model in female (FVB/N x Mus musculus castaneus)F1 mice. Mammary tumors developed as expected, as well as a few tumors of a second type (uterine leiomyosarcoma) not previously associated with this transgene. Genotyping of 94 anatomically independent tumors revealed high-frequency LOH ( approximately 38%) for markers on chromosome 4. A marked allelic bias was observed, with M. musculus castaneus alleles almost exclusively being lost. No evidence of genomic imprinting effects was noted. These data point to the presence of a tumor suppressor gene(s) on mouse chromosome 4 involved in mammary carcinogenesis induced by mutant H-ras expression, and for which a significant functional difference may exist between the M. musculus castaneus and FVB/N alleles. Provisional subchromosomal localization of this gene, designated Loh-3, can be made to a distal segment having syntenic correspondence to human chromosome 1p; LOH in this latter region is observed in several human malignancies, including breast cancers. Evidence was also obtained for a possible second locus associated with LOH with less marked allele bias on proximal chromosome 4.     

Impaired febrile response in mice lacking the prostaglandin E receptor subtype EP3

Fever, a hallmark of disease, is elicited by exogenous pyrogens, that is, cellular components, such as lipopolysaccharide (LPS), of infectious organisms, as well as by non-infectious inflammatory insults. Both stimulate the production of cytokines, such as interleukin (IL)-1beta, that act on the brain as endogenous pyrogens. Fever can be suppressed by aspirin-like anti-inflammatory drugs. As these drugs share the ability to inhibit prostaglandin biosynthesis, it is thought that a prostaglandin is important in fever generation. Prostaglandin E2 (PGE2) may be a neural mediator of fever, but this has been much debated. PGE2 acts by interacting with four subtypes of PGE receptor, the EP1, EP2, EP3 and EP4 receptors. Here we generate mice lacking each of these receptors by homologous recombination. Only mice lacking the EP3 receptor fail to show a febrile response to PGE2 and to either IL-1beta or LPS. Our results establish that PGE2 mediates fever generation in response to both exogenous and endogenous pyrogens by acting at the EP3 receptor.     

Retroviral transformation in vitro of chicken T cells expressing either alpha/beta or gamma/delta T cell receptors by reticuloendotheliosis virus strain T

Exposure of normal juvenile chicken bone marrow cells to the replication defective avian reticuloendotheliosis virus strain T (REV-T) (chicken syncytial virus [CSV]) in vitro resulted in the generation of transformed cell lines containing T cells. The transformed T cells derived from bone marrow included cells expressing either alpha/beta or gamma/delta T cell receptors (TCRs) in proportions roughly equivalent to the proportions of TCR-alpha/beta and TCR-gamma/delta T cells found in the normal bone marrow in vivo. Essentially all TCR-alpha/beta-expressing transformed bone marrow-derived T cells expressed CD8, whereas few, if any, expressed CD4. In contrast, among TCR-gamma/delta T cells, both CD8+ and CD8- cells were derived, all of which were CD4-. Exposure of ex vivo spleen cells to REV-T(CSV) yielded transformed polyclonal cell lines containing > 99% B cells. However, REV-T(CSV) infection of mitogen-activated spleen cells in vitro resulted in transformed populations containing predominantly T cells. This may be explained at least in part by in vitro activation resulting in dramatically increased levels of T cell REV-T(CSV) receptor expression. In contrast to REV-T(CSV)-transformed lines derived from normal bone marrow, transformed lines derived from activated spleen cells contained substantial numbers of CD4+ cells, all of which expressed TCR-alpha/beta. While transformed T cells derived from bone marrow were stable for extended periods of in vitro culture and were cloned from single cells, transformed T cells from activated spleen were not stable and could not be cloned. We have therefore dissociated the initial transformation of T cells with REV-T(CSV) from the requirements for long-term growth. These results provide the first demonstration of efficient in vitro transformation of chicken T lineage cells by REV-T(CSV). Since productive infection with REV-T(CSV) is not sufficient to promote long-term growth of transformed cells, these results further suggest that immortalization depends not only upon expression of the v-rel oncogene but also on intracellular factor(s) whose expression varies according to the state of T cell physiology and/or activation.     

Efficient, repeated adenovirus-mediated gene transfer in mice lacking both tumor necrosis factor alpha and lymphotoxin alpha

The efficiency of adenovirus-mediated gene transfer is now well established. However, the cellular and the humoral immune responses triggered by vector injection lead to the rapid elimination of the transduced cells and preclude any efficient readministration. The present investigation focuses on the role of tumor necrosis factor alpha (TNF-alpha), a proinflammatory cytokine, and the related cytokine lymphotoxin alpha (LTalpha), in mounting an immune reaction against recombinant adenovirus vectors. After gene transfer in the liver, mice genetically deficient for both cytokines (TNF-alpha/LTalpha-/-), in comparison with normal mice, presented a weak acute-phase inflammatory reaction, a reduction in cellular infiltrates in the liver, and a severely impaired T-cell proliferative response to both Adenoviral and transgene product antigens. Moreover, we observed a strong reduction in the humoral response to the vector and the transgene product, with a drastic reduction of anti-adenovirus immunoglobulin A and G antibody isotypes. In addition, the reduction in antibody response observed in TNF-alpha/LTalpha-/- and TNF-alpha/LTalpha+/- mice versus TNF-alpha/LTalpha+/+ mice links antibody levels to TNF-alpha/LTalpha gene dosage. Due to the absence of neutralizing antibodies, the TNF-alpha/LTalpha knockout mice successfully express a second gene transduced by a second vector injection. The discovery of the pivotal role played by TNF-alpha in controlling the antibody response against adenovirus will allow more efficient adenovirus-based strategies for gene therapy to be proposed.     

Specific contribution of Tyk2 JH regions to the binding and the expression of the interferon alpha/beta receptor component IFNAR1

Cytokine signaling involves the activation of the Janus kinase (JAK) family of tyrosine kinases. These enzymes are physically associated with cytokine receptor components. Here, we sought to define the molecular basis of the interaction between Tyk2 and IFNAR1, a component of the interferon alpha/beta receptor, by delimiting a minimal IFNAR1 binding region in the Tyk2 protein. Using an in vitro assay system, we narrowed down the interaction domain to a region comprising the JH7 and part of the JH6 homology boxes (amino acids 22-221). When expressed in Tyk2-negative cells, the JH7-6 region was unable to stabilize IFNAR1 protein levels, a critical function that we previously attributed to the N region (amino acids 1-591) of Tyk2. Moreover, substitution of the JH7-JH6 domain in JAK1 with that of Tyk2 did not restore IFNAR1 level nor interferon alpha signaling in Tyk2-negative cells. Thus, the major interaction surface lies within JH7-6, but additional JH regions (JH5-4-3) contribute in a specific manner to the in vivo assembly of Tyk2 and IFNAR1. Evidence is also provided of the lack of specificity of the Tyk2 kinase-like and tyrosine kinase domains in interferon alpha/beta receptor signaling.     

B and T cells are required for mouse mammary tumor virus spread within the mammary gland

Mouse mammary tumor virus (MMTV) is an infectious retrovirus transmitted through milk from mother to newborns. MMTV encodes a superantigen (SAg) whose activity is indispensable for the virus life cycle, since a genetically engineered virus with a mutation in the sag gene neither amplified in cells of the immune system of suckling pups nor infected their mammary glands. When wild-type MMTV was injected directly into the mammary glands of uninfected pubescent mice, their lymphoid as well as mammary gland cells became virus infected. To test whether this infection of lymphoid cells was dependent on SAg activity and required for virus spread within the mammary gland, we performed mammary gland injections of wild-type MMTV(C3H) into two strains of transgenic mice that lacked SAg-cognate, V beta 14+ T cells. Neither the MTV-ORF or LEL strains showed infection of their mammary glands. Moreover, no MMTV infection of their peripheral lymphocytes was detected. Similar experiments with mice lacking B cells (mu-chain knockouts) showed no detectable virus spread in the mammary glands or lymphoid tissues. These data suggest that SAg activity and MMTV-infected lymphocytes are required, not only for initial steps of viral infection, but also for virus spread within the mammary gland. Virus spread at late times in infection determines whether MMTV induces mammary tumors.     

Defective nitric oxide effector functions lead to extreme susceptibility of Trypanosoma cruzi-infected mice deficient in gamma interferon receptor or inducible nitric oxide synthase

Trypanosoma cruzi, the causative agent of Chagas' disease, induces an innate and adaptive host immune response during the acute phase of infection. These responses were analyzed by comparing mouse lines deficient for the gamma interferon (IFN-gamma) receptor (IFN-gammaR(-/-)) or deficient for inducible nitric oxide synthase (iNOS(-/-)). Both lines were highly susceptible, with similar and dramatically increased parasite burdens and severe histopathology and were incapable of surviving even very low doses, exhibiting similar mortality kinetics. This pathophysiological correlation has a common cause, since both mutant mouse strains were unable to respond to infection by producing nitric oxide (NO) with the consequence that mutant macrophages had impaired trypanocidal activities. These in vivo and subsequent in vitro studies further demonstrated that an IFN-gamma-dependent pathway of iNOS induction is crucial for efficient NO production and mandatory for resisting acute infection with T. cruzi. Despite this defect, both mutant mouse strains had a rather normal proinflammatory cytokine response (interleukin-12 [IL-12], IFN-gamma, IL-6), with the exception of an impaired tumor necrosis factor alpha and IL-1alpha response in IFN-gammaR(-/-) mice, demonstrating that only the latter two cytokines are dependent on IFN-gamma activation. Moreover, polarization of T cells in type 1 and type 2 T-helper (Th1/Th2) and cytotoxic T (Tc1/Tc2) cells as well as T. cruzi-specific antibody responses were normal in IFN-gammaR(-/-) mice, demonstrating that IFN-gamma is not necessary for the promotion of T-cell differentiation and T. cruzi-specific antibody responses.     

Differential reactivity of TCR Vbeta10 alleles to a mouse mammary tumor virus superantigen

Mouse mammary tumor virus (MMTV) expresses a superantigen (SAg) which plays a critical role in the viral life cycle. We have recently described the new infectious MMTV (SIM) encoding a Vbeta4-specific SAg in mice with a TCR-Vbeta(b) haplotype. We have now compared the SAg activity of this virus in BALB/c mice harboring the TCR-Vbeta(a), TCR-Vbeta(b) or TCR-Vbeta(c) haplotypes which differ by a central deletion in the TCR-Vbeta(a) and TCR-Vbeta(c) locus and by mutations in some of the remaining Vbeta elements. Injection of MMTV (SIM) led to a strong stimulation of Vbeta4+ CD4+ T cells in TCR-Vbeta(b) mice, but only to a weak stimulation of these cells in TCR-Vbeta(a) or TCR-Vbeta(c) mice. A large increase in the percentage of Vbeta10+ cells was observed among CD4+ T cells in mice with the Vbeta(a) or Vbeta(c), but not the Vbeta(b) TCR-Vbeta haplotype. Vbeta10+ cells dominated the response when Vbeta10(a/c) and Vbeta4 subsets were present together. This is the first report of a viral SAg interacting with murine Vbeta10+ cells. Six amino acid differences between Vbeta10(a/c) and Vbeta10(b) could account for the gain of reactivity of Vbeta10(a/c) to the MMTV(SIM) SAg. No mutations were found in the hypervariable region 4 (HV4) of the TCR. Mutations at positions 22 and 28 introduce into Vbeta10(a/c) the same amino acids which are found at these positions in the MMTV(SIM)-reactive Vbeta4. Tridimensional models indicated that these amino acids lie close to HV4 and are likely to be important for the interaction of the SAg with the TCR.