The Signal Sequence of Lymphocytic Choriomeningitis Virus Contains an Immunodominant Cytotoxic T Cell Epitope That Is Restricted by both H-2Dband H-2KbMolecules

AbstractInfection of H-2bmice with lymphocytic choriomeningitis virus (LCMV) generates three well-characterized H-2Db-restricted immunodominant epitopes delineated in the NP, GP1, and GP2 proteins. Here we report that the H-2Db-restricted GP1 epitope GP33-41/43 (KAVYNFATC/GI) located in the signal sequence of LCMV is also the immunodominant epitope recognized by CTL at the surface of the same infected cells in the context of H-2Kbrestriction. The GP1 epitope bound to H-2Dband H-2Kbmolecules with comparable affinities. The respective binding processes involved different sets of peptide anchoring residues and required dramatically different conformations of the peptide backbone as well as rearrangement of residue side chains. The 10-mer peptide GP34-43 (AVYNFATCGI) was the optimal H-2Kb-binding sequence and the 8-mer peptide GP34-41 (AVYNFATC) the minimal sequence for optimal H-2Kb-restricted CTL recognition. Comparison of lytic activities of primary splenic anti-LCMV CTL from C57BL/6 (Db+/Kb+), B10A.[5R] (Db−/Kb+), and B10A.[2R] (Db+/Kb−) mice against LCMV-infected or peptide-coated target cells expressing either one or the two MHC alleles revealed that the H-2Kb-restricted component of the anti-GP1 CTL response was mounted independently of but as efficiently as its H-2Dbcounterpart. Analysis of the immune response against a GP1 variant that escapes CTL recognition showed that the GP1 epitope: (i) was likely the only immunodominant LCMV epitope in the context of H-2Kb, and (ii) could efficiently evade H-2Dband H-2Kb-restricted CTL mediated lysis

A role for dual viral hits in causation of subacute sclerosing panencephalitis

Subacute sclerosing panencephalitis (SSPE) is a progressive fatal neurodegenerative disease associated with persistent infection of the central nervous system (CNS) by measles virus (MV), biased hypermutations of the viral genome affecting primarily the matrix (M) gene with the conversion of U to C and A to G bases, high titers of antibodies to MV, and infiltration of B cells and T cells into the CNS. Neither the precipitating event nor biology underlying the MV infection is understood, nor is their any satisfactory treatment. We report the creation of a transgenic mouse model that mimics the cardinal features of SSPE. This was achieved by initially infecting mice expressing the MV receptor with lymphocytic choriomeningitis virus Cl 13, a virus that transiently suppressed their immune system. Infection by MV 10 days later resulted in persistent MV infection of neurons. Analysis of brains from infected mice showed the biased U to C hypermutations in the MV M gene and T and B lymphocyte infiltration. These sera contained high titers of antibodies to MV. Thus, a small animal model is now available to both molecularly probe the pathogenesis of SSPE and to test a variety of therapies to treat the disease

Sensitization to self (virus) antigen by in situ expression of murine interferon-gamma.

Autoimmune disease results from inflammatory destruction of tissues by aberrant self-reactive lymphocytes. We studied the autoimmune potential of T lymphocytes immunologically ignorant of viral antigens acting as self antigens and whether the host defense molecule IFN-gamma could stimulate these cells to cytotoxic competency. For this purpose, we produced double transgenic mice expressing pancreatic IFN-gamma as well as lymphocytic choriomeningitis virus (LCMV) nucleoprotein (NP) or glycoprotein (GP) antigen. 100% of the NP+/IFN-gamma+ mice became diabetic before 2 mo of age, while none of the NP single transgenic littermates and only 10% of IFN-gamma single transgenic littermates did. Strikingly, NP+/IFN-gamma+ mice spontaneously developed cytotoxic T lymphocyte activity on LCMV-infected targets and vaccinia virus-NP-infected ones without prior LCMV infection but NP+/IFN-gamma- mice did not, which indicates specific sensitization to the viral antigen by IFN-gamma. These results suggest that lymphocytes ignorant of self antigens can be activated by IFN-gamma released after immunologic stimulation such as viral infection. This mechanism may account for the loss of apparent tolerance to self antigens in autoimmune diseases such as insulin-dependent diabetes mellitus

Adoptive immunotherapy induces CNS dendritic cell recruitment and antigen presentation during clearance of a persistent viral infection

Given the global impact of persistent infections on the human population, it is of the utmost importance to devise strategies to noncytopathically purge tissues of infectious agents. The central nervous system (CNS) poses a unique challenge when considering such strategies, as it is an immunologically specialized compartment that contains a nonreplicative cell population. Administration of exogenously derived pathogen-specific memory T cells (referred to as adoptive immunotherapy) to mice burdened with a persistent lymphocytic choriomeningitis virus (LCMV) infection from birth results in eradication of the pathogen from all tissues, including the CNS. In this study, we sought mechanistic insights into this highly successful therapeutic approach. By monitoring the migration of traceable LCMV-specific memory CD8+ T cells after immunotherapy, it was revealed that cytotoxic T lymphocytes (CTLs) distributed widely throughout the CNS compartment early after immunotherapy, which resulted in a dramatic elevation in the activity of CNS antigen-presenting cells (APCs). Immunotherapy induced microglia activation as well as the recruitment of macrophages and dendritic cells (DCs) into the brain parenchyma. However, DCs emerged as the only CNS APC population capable of inducing memory CTLs to preferentially produce the antiviral cytokine tumor necrosis factor-α, a cytokine demonstrated to be required for successful immunotherapeutic clearance. DCs were also found to be an essential element of the immunotherapeutic process because in their absence, memory T cells failed to undergo secondary expansion, and viral clearance was not attained in the CNS. These experiments underscore the importance of DCs in the immunotherapeutic clearance of a persistent viral infection and suggest that strategies to elevate the activation/migration of DCs (especially within the CNS) may facilitate pathogen clearance

IL-10 blockade facilitates DNA vaccine-induced T cell responses and enhances clearance of persistent virus infection

Therapeutic vaccination is a potentially powerful strategy to establish immune control and eradicate persistent viral infections. Large and multifunctional antiviral T cell responses are associated with control of viral persistence; however, for reasons that were mostly unclear, current therapeutic vaccination approaches to restore T cell immunity and control viral infection have been ineffective. Herein, we confirmed that neutralization of the immunosuppressive factor interleukin (IL)-10 stimulated T cell responses and improved control of established persistent lymphocytic choriomeningitis virus (LCMV) infection. Importantly, blockade of IL-10 also allowed an otherwise ineffective therapeutic DNA vaccine to further stimulate antiviral immunity, thereby increasing T cell responses and enhancing clearance of persistent LCMV replication. We therefore propose that a reason that current therapeutic vaccination strategies fail to resurrect/sustain T cell responses is because they do not alleviate the immunosuppressive environment. Consequently, blocking key suppressive factors could render ineffective vaccines more efficient at improving T cell immunity, and thereby allow immune-mediated control of persistent viral infection

c-Jun NH2-Terminal Kinase (JNK)1 and JNK2 Signaling Pathways Have Divergent Roles in CD8+ T Cell–mediated Antiviral Immunity

c-Jun NH2-terminal kinases (JNK) play important roles in T helper cell (Th) proliferation, differentiation, and maintenance of Th1/Th2 polarization. To determine whether JNKs are involved in antiviral T cell immunity, and whether JNK1 and JNK2 bear biological differences, we investigated the immune responses of JNK1-deficient and JNK2-deficient mice to lymphocytic choriomeningitis virus (LCMV). After LCMV infection, wild-type (JNK+/+) mice had a 5- to 10-fold increase in splenic CD8+ T cells. In contrast, infected JNK1−/− mice showed a significantly lower virus-specific CD8+ T cell expansion. However, JNK1−/− mice cleared LCMV infection with similar kinetics as JNK+/+ mice. Splenic T cells from LCMV-infected JNK1−/− animals produced interferon γ after stimulation with viral peptides. However, fewer JNK1−/− T cells acquired an activated phenotype (CD44hi) and more JNK1−/−CD8+CD44hi cells underwent apoptosis than JNK+/+ cells at the peak of the primary response. In contrast, LCMV-infected JNK2−/− mice generated more virus-specific CD8+ T cells than JNK+/+ mice. These results indicate that JNK1 and JNK2 signal pathways have distinct roles in T cell responses during a viral infection. JNK1 is involved in survival of activated T cells during immune responses, and JNK2 plays a role in control of CD8+ T cell expansion in vivo

Measles Virus Recognizes Its Receptor, CD46, via Two Distinct Binding Domains within SCR1-2

AbstractMeasles virus (MV) enters cells by attachment of the viral hemagglutinin to the major cell surface receptor CD46 (membrane cofactor protein). CD46 is a transmembrane glycoprotein whose ectodomain is largely composed of four conserved modules called short consensus repeats (SCRs). We have previously shown that MV interacts with SCR1 and SCR2 of CD46. (M. Manchesteret al.(1995)Proc. Natl. Acad. Sci. USA92, 2303–2307) Here we report mapping the MV interaction with SCR1 and SCR2 of CD46 using a combination of peptide inhibition and mutagenesis studies. By testing a series of overlapping peptides corresponding to the 126 amino acid SCR1-2 region for inhibition of MV infection, two domains were identified that interacted with MV. One domain was found within SCR1 (amino acids 37–56) and another within SCR2 (amino acids 85–104). These results were confirmed by constructing chimeras with complementary regions from structurally similar, but non-MV-binding, SCRs of decay accelerating factor (DAF; CD55). These results indicate that MV contacts at least two distinct sites within SCR1-2

Measles Virus Infection in a Transgenic Model Virus-Induced Immunosuppression and Central Nervous System Disease

AbstractMeasles virus (MV) infects 40 million persons and kills one million per year primarily by suppressing the immune system and afflicting the central nervous system (CNS). The lack of a suitable small animal model has impeded progress of understanding how MV causes disease and the development of novel therapies and improved vaccines. We tested a transgenic mouse line in which expression of the MV receptor CD46 closely mimicked the location and amount of CD46 found in humans. Virus replicated in and was recovered from these animals' immune systems and was associated with suppression of humoral and cellular immune responses. Infectious virus was recovered from the CNS, replicated primarily in neurons, and spread to distal sites presumably by fast axonal transport. Thus, a small animal model is available for analysis of MV pathogenesis