RESUMEN
Infection by chikungunya virus (CHIKV), a mosquito-borne alphavirus, causes severe polyarthralgia and polymyalgia, which can last in some people for months to years. Chronic CHIKV disease signs and symptoms are associated with the persistence of viral nucleic acid and antigen in tissues. Like humans and nonhuman primates, CHIKV infection in mice results in the development of robust adaptive antiviral immune responses. Despite this, joint tissue fibroblasts survive CHIKV infection and can support persistent viral replication, suggesting that they escape immune surveillance. Here, using a recombinant CHIKV strain encoding the fluorescent protein VENUS with an embedded CD8+ T cell epitope, SIINFEKL, we observed a marked loss of both MHC class I (MHC-I) surface expression and antigen presentation by CHIKV-infected joint tissue fibroblasts. Both in vivo and ex vivo infected joint tissue fibroblasts displayed reduced cell surface levels of H2-Kb and H2-Db MHC-I proteins while maintaining similar levels of other cell surface proteins. Mutations within the methyl transferase-like domain of the CHIKV nonstructural protein 2 (nsP2) increased MHC-I cell surface expression and antigen presentation efficiency by CHIKV-infected cells. Moreover, expression of WT nsP2 alone, but not nsP2 with mutations in the methyltransferase-like domain, resulted in decreased MHC-I antigen presentation efficiency. MHC-I surface expression and antigen presentation was rescued by replacing VENUS-SIINFEKL with SIINFEKL tethered to ß2-microglobulin in the CHIKV genome, which bypasses the requirement for peptide processing and TAP-mediated peptide transport into the endoplasmic reticulum. Collectively, this work suggests that CHIKV escapes the surveillance of antiviral CD8+ T cells, in part, by nsP2-mediated disruption of MHC-I antigen presentation.
Asunto(s)
Fiebre Chikungunya , Virus Chikungunya , Humanos , Animales , Ratones , Presentación de Antígeno , Replicación Viral , Proteínas no Estructurales Virales/genética , Proteínas no Estructurales Virales/metabolismo , Epítopos de Linfocito T , Péptidos/metabolismoRESUMEN
Alphaviruses are mosquito-transmitted pathogens that induce high levels of viremia, which facilitates dissemination and vector transmission. One prevailing paradigm is that, after skin inoculation, alphavirus-infected resident dendritic cells migrate to the draining lymph node (DLN), facilitating further rounds of infection and dissemination. Here, we assess the contribution of infiltrating myeloid cells to alphavirus spread. We observe two phases of virus transport to the DLN, one that occurs starting at 1 h post infection and precedes viral replication, and a second that requires replication in the skin, enabling transit to the bloodstream. Depletion of Ly6C+ monocytes reduces local chikungunya (CHIKV) or Ross River virus (RRV) infection in the skin, diminishes the second phase of virus transport to the DLN, and delays spread to distal sites. Our data suggest that infiltrating monocytes facilitate alphavirus infection at the initial infection site, which promotes more rapid spread into circulation.
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Fiebre Chikungunya , Virus Chikungunya , Animales , Monocitos/patología , Mosquitos Vectores , Fiebre Chikungunya/patología , Células Mieloides , Replicación ViralRESUMEN
Infection by chikungunya virus (CHIKV), a mosquito-borne alphavirus, causes severe polyarthralgia and polymyalgia, which can last in some people for months to years. Chronic CHIKV disease signs and symptoms are associated with the persistence of viral nucleic acid and antigen in tissues. Like humans and nonhuman primates, CHIKV infection in mice results in the development of robust adaptive antiviral immune responses. Despite this, joint tissue fibroblasts survive CHIKV infection and can support persistent viral replication, suggesting that they escape immune surveillance. Here, using a recombinant CHIKV strain encoding a chimeric protein of VENUS fused to a CD8+ T cell epitope, SIINFEKL, we observed a marked loss of both MHC class I (MHC-I) surface expression and antigen presentation by CHIKV-infected joint tissue fibroblasts. Both in vivo and ex vivo infected joint tissue fibroblasts displayed reduced cell surface levels of H2-Kb and H2-Db MHC proteins while maintaining similar levels of other cell surface proteins. Mutations within the methyl transferase-like domain of the CHIKV nonstructural protein 2 (nsP2) increased MHC-I cell surface expression and antigen presentation efficiency by CHIKV-infected cells. Moreover, expression of WT nsP2 alone, but not nsP2 with mutations in the methyltransferase-like domain, resulted in decreased MHC-I antigen presentation efficiency. MHC-I surface expression and antigen presentation could be rescued by replacing VENUS-SIINFEKL with SIINFEKL tethered to ß2-microglobulin in the CHIKV genome, which bypasses the need for peptide processing and TAP-mediated peptide transport into the endoplasmic reticulum. Collectively, this work suggests that CHIKV escapes the surveillance of antiviral CD8+ T cells, in part, by nsP2-mediated disruption of MHC-I antigen presentation.
RESUMEN
Chikungunya virus (CHIKV) has recently emerged to cause millions of human infections worldwide. Infection can induce the formation of long intercellular extensions that project from infected cells and form stable non-continuous membrane bridges with neighbouring cells. The mechanistic role of these intercellular extensions in CHIKV infection was unclear. Here we developed a co-culture system and flow cytometry methods to quantitatively evaluate transmission of CHIKV from infected to uninfected cells in the presence of neutralizing antibody. Endocytosis and endosomal acidification were critical for virus cell-to-cell transmission, while the CHIKV receptor MXRA8 was not. By using distinct antibodies to block formation of extensions and by evaluation of transmission in HeLa cells that did not form extensions, we showed that intercellular extensions mediate CHIKV cell-to-cell transmission. In vivo, pre-treatment of mice with a neutralizing antibody blocked infection by direct virus inoculation, while adoptive transfer of infected cells produced antibody-resistant host infection. Together our data suggest a model in which the contact sites of intercellular extensions on target cells shield CHIKV from neutralizing antibodies and promote efficient intercellular virus transmission both in vitro and in vivo.
Asunto(s)
Fiebre Chikungunya , Virus Chikungunya , Humanos , Animales , Ratones , Células HeLa , Anticuerpos Neutralizantes , Técnicas de CocultivoRESUMEN
The microbial-derived metabolite, 3-indolepropionic acid (3-IPA), has been intensely studied since its origins were discovered in 2009; however, 3-IPA's role in immunosuppression has had limited attention. Untargeted metabolomic analyses of T-cell exhaustion and immunosuppression, represented by dysfunctional under-responsive CD8+ T cells, reveal a potential role of 3-IPA in these responses. T-cell exhaustion was examined via infection of two genetically related mouse strains, DBA/1J and DBA/2J, with lymphocytic choriomeningitis virus (LCMV) Clone 13 (Cl13). The different mouse strains produced disparate outcomes driven by their T-cell responses. Infected DBA/2J presented with exhausted T cells and persistent infection, and DBA/1J mice died one week after infection from cytotoxic T lymphocytes (CTLs)-mediated pulmonary failure. Metabolomics revealed over 70 metabolites were altered between the DBA/1J and DBA/2J models over the course of the infection, most of them in mice with a fatal outcome. Cognitive-driven prioritization combined with statistical significance and fold change were used to prioritize the metabolites. 3-IPA, a tryptophan-derived metabolite, was identified as a high-priority candidate for testing. To test its activity 3-IPA was added to the drinking water of the mouse models during LCMV Cl13 infection, with the results showing that 3-IPA allowed the mice to survive longer. This negative immune-modulation effect might be of interest for the modulation of CTL responses in events such as autoimmune diseases, type I diabetes or even COVID-19. Moreover, 3-IPA's bacterial origin raises the possibility of targeting the microbiome to enhance CTL responses in diseases such as cancer and chronic infection.
RESUMEN
Mature male mice produce a particularly high concentration of major urinary proteins (MUPs) in their scent marks that provide identity and status information to conspecifics. Darcin (MUP20) is inherently attractive to females and, by inducing rapid associative learning, leads to specific attraction to the individual male's odour and location. Other polymorphic central MUPs, produced at much higher abundance, bind volatile ligands that are slowly released from a male's scent marks, forming the male's individual odour that females learn. Here, we show that infection of C57BL/6 males with LCMV WE variants (v2.2 or v54) alters MUP expression according to a male's infection status and ability to clear the virus. MUP output is substantially reduced during acute adult infection with LCMV WE v2.2 and when males are persistently infected with LCMV WE v2.2 or v54. Infection differentially alters expression of darcin and, particularly, suppresses expression of a male's central MUP signature. However, following clearance of acute v2.2 infection through a robust virus-specific CD8 cytotoxic T cell response that leads to immunity to the virus, males regain their normal mature male MUP pattern and exhibit enhanced MUP output by 30 days post-infection relative to uninfected controls. We discuss the likely impact of these changes in male MUP signals on female attraction and mate selection. As LCMV infection during pregnancy can substantially reduce embryo survival and lead to lifelong infection in surviving offspring, we speculate that females use LCMV-induced changes in MUP expression both to avoid direct infection from a male and to select mates able to develop immunity to local variants that will be inherited by their offspring.
Asunto(s)
Coriomeningitis Linfocítica/complicaciones , Virus de la Coriomeningitis Linfocítica/patogenicidad , Proteínas/metabolismo , Animales , Femenino , Péptidos y Proteínas de Señalización Intercelular/genética , Coriomeningitis Linfocítica/inmunología , Virus de la Coriomeningitis Linfocítica/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Odorantes , Feromonas/metabolismo , Proteínas/análisis , Proteínas/genéticaRESUMEN
The detection of foreign antigens in vivo has relied on fluorescent conjugation or indirect read-outs such as antigen presentation. In our studies, we found that these widely used techniques had several technical limitations that have precluded a complete picture of antigen trafficking or retention across lymph node cell types. To address these limitations, we developed a 'molecular tracking device' to follow the distribution, acquisition, and retention of antigen in the lymph node. Utilizing an antigen conjugated to a nuclease-resistant DNA tag, acting as a combined antigen-adjuvant conjugate, and single-cell mRNA sequencing, we quantified antigen abundance in the lymph node. Variable antigen levels enabled the identification of caveolar endocytosis as a mechanism of antigen acquisition or retention in lymphatic endothelial cells. Thus, these molecular tracking devices enable new approaches to study dynamic tissue dissemination of antigen-adjuvant conjugates and identify new mechanisms of antigen acquisition and retention at cellular resolution in vivo.
The lymphatic system is a network of ducts that transports fluid, proteins, and immune cells from different organs around the body. Lymph nodes provide pit stops at hundreds of points along this network where immune cells reside, and lymph fluid can be filtered and cleaned. When pathogens, such as viruses or bacteria, enter the body during an infection, fragments of their proteins can get swept into the lymph nodes. These pathogenic proteins or protein fragments activate resident immune cells and kickstart the immune response. Vaccines are designed to mimic this process by introducing isolated pathogenic proteins in a controlled way to stimulate similar immune reactions in lymph nodes. Once an infection has been cleared by the immune system, or a vaccination has triggered the immune system, most pathogenic proteins get cleared away. However, a small number of pathogenic proteins remain in the lymph nodes to enable immune cells to respond more strongly and quickly the next time they see the same pathogen. Yet it is largely unclear how much protein remains for training and how or where it is all stored. Current techniques are not sensitive or long-lived enough to accurately detect and track these small protein deposits over time. Walsh, Sheridan, Lucas, et al. have addressed this problem by developing biological tags that can be attached to the pathogenic proteins so they can be traced. These tags were designed so the body cannot easily break them down, helping them last as long as the proteins they are attached to. Walsh, Sheridan, Lucas et al. tested whether vaccinating mice with the tagged proteins allowed the proteins to be tracked. The method they used was designed to identify individual cell types based on their genetic information along with the tag. This allowed them to accurately map the complex network of cells involved in storing and retrieving archived protein fragments, as well as those involved in training new immune cells to recognize them. These results provide important insights into the protein archiving system that is involved in enhancing immune memory. This may help guide the development of new vaccination strategies that can manipulate how proteins are archived to establish more durable immune protection. The biological tags developed could also be used to track therapeutic proteins, allowing scientists to determine how long cancer drugs, antibody therapies or COVID19 anti-viral agents remain in the body. This information could then be used by doctors to plan specific and personalized treatment timetables for patients.
Asunto(s)
Antígenos/metabolismo , Ganglios Linfáticos/metabolismo , Análisis de la Célula Individual , Animales , Presentación de Antígeno , Antígenos/genética , Antígenos/inmunología , Caveolas/inmunología , Caveolas/metabolismo , Células Cultivadas , ADN/genética , ADN/metabolismo , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Endocitosis , Células Endoteliales/inmunología , Células Endoteliales/metabolismo , Ganglios Linfáticos/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones Endogámicos C57BL , Ratones Transgénicos , Ovalbúmina/genética , Ovalbúmina/inmunología , Ovalbúmina/metabolismo , Fragmentos de Péptidos/genética , Fragmentos de Péptidos/inmunología , Fragmentos de Péptidos/metabolismo , Oligonucleótidos Fosforotioatos/genética , Oligonucleótidos Fosforotioatos/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Análisis de Secuencia de ARN , Factores de Tiempo , Distribución Tisular , TranscriptomaRESUMEN
Early and robust T cell responses have been associated with survival from Lassa fever (LF), but the Lassa virus-specific memory responses have not been well characterized. Regions within the virus surface glycoprotein (GPC) and nucleoprotein (NP) are the main targets of the Lassa virus-specific T cell responses, but, to date, only a few T cell epitopes within these proteins have been identified. We identified GPC and NP regions containing T cell epitopes and HLA haplotypes from LF survivors and used predictive HLA-binding algorithms to identify putative epitopes, which were then experimentally tested using autologous survivor samples. We identified 12 CD8-positive (CD8+) T cell epitopes, including epitopes common to both Nigerian and Sierra Leonean survivors. These data should be useful for the identification of dominant Lassa virus-specific T cell responses in Lassa fever survivors and vaccinated individuals as well as for designing vaccines that elicit cell-mediated immunity.IMPORTANCE The high morbidity and mortality associated with clinical cases of Lassa fever, together with the lack of licensed vaccines and limited and partially effective interventions, make Lassa virus (LASV) an important health concern in its regions of endemicity in West Africa. Previous infection with LASV protects from disease after subsequent exposure, providing a framework for designing vaccines to elicit similar protective immunity. Multiple major lineages of LASV circulate in West Africa, and therefore, ideal vaccine candidates should elicit immunity to all lineages. We therefore sought to identify common T cell epitopes between Lassa fever survivors from Sierra Leone and Nigeria, where distinct lineages circulate. We identified three such epitopes derived from highly conserved regions within LASV proteins. In this process, we also identified nine other T cell epitopes. These data should help in the design of an effective pan-LASV vaccine.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Epítopos de Linfocito T/química , Fiebre de Lassa/inmunología , Virus Lassa/inmunología , Nucleoproteínas/inmunología , Proteínas del Envoltorio Viral/inmunología , Adolescente , Secuencia de Aminoácidos , Animales , Anticuerpos Antivirales/biosíntesis , Antígenos Virales/química , Antígenos Virales/genética , Antígenos Virales/inmunología , Linfocitos T CD8-positivos/virología , Niño , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/inmunología , Femenino , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/inmunología , Antígenos HLA-DQ/genética , Antígenos HLA-DQ/inmunología , Haplotipos , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/inmunología , Humanos , Sueros Inmunes/análisis , Memoria Inmunológica , Fiebre de Lassa/genética , Fiebre de Lassa/patología , Virus Lassa/patogenicidad , Masculino , Nigeria , Nucleoproteínas/genética , Sierra Leona , Sobrevivientes , Proteínas del Envoltorio Viral/genética , Adulto JovenRESUMEN
Lassa virus infects hundreds of thousands of people each year across rural West Africa, resulting in a high number of cases of Lassa fever (LF), a febrile disease associated with high morbidity and significant mortality. The lack of approved treatments or interventions underscores the need for an effective vaccine. At least four viral lineages circulate in defined regions throughout West Africa with substantial interlineage nucleotide and amino acid diversity. An effective vaccine should be designed to elicit Lassa virus specific humoral and cell mediated immunity across all lineages. Most current vaccine candidates use only lineage IV antigens encoded by Lassa viruses circulating around Sierra Leone, Liberia, and Guinea but not Nigeria where lineages I-III are found. As previous infection is known to protect against disease from subsequent exposure, we sought to determine whether LF survivors from Nigeria and Sierra Leone harbor memory T cells that respond to lineage IV antigens. Our results indicate a high degree of cross-reactivity of CD8+ T cells from Nigerian LF survivors to lineage IV antigens. In addition, we identified regions within the Lassa virus glycoprotein complex and nucleoprotein that contributed to these responses while T cell epitopes were not widely conserved across our study group. These data are important for current efforts to design effective and efficient vaccine candidates that can elicit protective immunity across all Lassa virus lineages.
Asunto(s)
Antígenos Virales/inmunología , Linfocitos T CD8-positivos/inmunología , Epítopos de Linfocito T/inmunología , Virus Lassa/inmunología , África Occidental , Reacciones Cruzadas , Femenino , Humanos , Masculino , Especificidad de la EspecieRESUMEN
Lymphocytic choriomeningitis virus (LCMV) WE variant 2.2 (v2.2) generated a high level of the major mouse urinary protein: MUP. Mice infected with LCMV WE v54, which differed from v2.2 by a single amino acid in the viral glycoprotein, failed to generate MUP above baseline levels found in uninfected controls. Variant 54 bound at 2.5 logs higher affinity to the LCMV receptor α-dystroglycan (α-DG) than v2.2 and entered α-DG-expressing but not α-DG-null cells. Variant 2.2 infected both α-DG-null or -expressing cells. Variant 54 infected more dendritic cells, generated a negligible CD8 T cell response, and caused a persistent infection, while v2.2 generated cytotoxic T lymphocytes (CTLs) and cleared virus within 10 days. By 20 days postinfection and through the 80-day observation period, significantly higher amounts of MUP were found in v2.2-infected mice. Production of MUP was dependent on virus-specific CTL as deletion of such cells aborted MUP production. Furthermore, MUP production was not elevated in v2.2 persistently infected mice unless virus was cleared following transfer of virus-specific CTL.
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Linfocitos T CD8-positivos/inmunología , Regulación de la Expresión Génica/inmunología , Coriomeningitis Linfocítica/inmunología , Virus de la Coriomeningitis Linfocítica/inmunología , Proteínas/inmunología , Animales , Distroglicanos/inmunología , Coriomeningitis Linfocítica/patología , RatonesRESUMEN
Here, we report a pathogenic role for type I IFN (IFN-I) signaling in macrophages, and not ß cells in the islets, for the development of type 1 diabetes (T1D). Following lymphocytic choriomeningitis (LCMV) infection in the Rip-LCMV-GP T1D model, macrophages accumulated near islets and in close contact to islet-infiltrating GP-specific (autoimmune) CD8+ T cells. Depletion of macrophages with clodronate liposomes or genetic ablation of Ifnar in macrophages aborted T1D, despite proliferation of GP-specific (autoimmune) CD8+ T cells. Histopathologically, disrupted IFNα/ß receptor (IFNAR) signaling in macrophages resulted in restriction of CD8+ T cells entering into the islets with significant lymphoid accumulation around the islet. Collectively, these results provide evidence that macrophages via IFN-I signaling, while not entering the islets, are directly involved in interacting, directing, or restricting trafficking of autoreactive-specific T cells into the islets as an important component in causing T1D.
RESUMEN
Understanding of T cell exhaustion and successful therapy to restore T cell function was first described using Clone (Cl) 13 variant selected from the lymphocytic choriomeningitis virus (LCMV) Armstrong (ARM) 53b parental strain. T cell exhaustion plays a pivotal role in both persistent infections and cancers of mice and humans. C57BL/6, BALB, SWR/J, A/J, 129, C3H, and all but one collaborative cross (CC) mouse strain following Cl 13 infection have immunosuppressed T cell responses, high PD-1, and viral titers leading to persistent infection and normal life spans. In contrast, the profile of FVB/N, NZB, PL/J, SL/J, and CC NZO mice challenged with Cl 13 is a robust T cell response, high titers of virus, PD-1, and Lag3 markers on T cells. These mice all die 7 to 9 d after Cl 13 infection. Death is due to enhanced pulmonary endothelial vascular permeability, pulmonary edema, collapse of alveolar air spaces, and respiratory failure. Pathogenesis involves abundant levels of Cl 13 receptor alpha-dystroglycan on endothelial cells, with high viral replication in such cells leading to immunopathologic injury. Death is aborted by blockade of interferon-1 (IFN-1) signaling or deletion of CD8 T cells.
Asunto(s)
Linfocitos T CD8-positivos , Interferón Tipo I , Coriomeningitis Linfocítica , Virus de la Coriomeningitis Linfocítica/fisiología , Replicación Viral/genética , Animales , Antígenos CD/genética , Antígenos CD/metabolismo , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/patología , Humanos , Interferón Tipo I/genética , Interferón Tipo I/metabolismo , Coriomeningitis Linfocítica/genética , Coriomeningitis Linfocítica/metabolismo , Coriomeningitis Linfocítica/patología , Ratones , Receptor de Muerte Celular Programada 1/genética , Receptor de Muerte Celular Programada 1/metabolismo , Proteína del Gen 3 de Activación de LinfocitosRESUMEN
The recent Ebola epidemic exemplified the importance of understanding and controlling emerging infections. Despite the importance of T cells in clearing virus during acute infection, little is known about Ebola-specific CD8+ T cell responses. We investigated immune responses of individuals infected with Ebola virus (EBOV) during the 2013-2016 West Africa epidemic in Sierra Leone, where the majority of the >28,000 EBOV disease (EVD) cases occurred. We examined T cell memory responses to seven of the eight Ebola proteins (GP, sGP, NP, VP24, VP30, VP35, and VP40) and associated HLA expression in survivors. Of the 30 subjects included in our analysis, CD8+ T cells from 26 survivors responded to at least one EBOV antigen. A minority, 10 of 26 responders (38%), made CD8+ T cell responses to the viral GP or sGP. In contrast, 25 of the 26 responders (96%) made response to viral NP, 77% to VP24 (20 of 26), 69% to VP40 (18 of 26), 42% (11 of 26) to VP35, with no response to VP30. Individuals making CD8+ T cells to EBOV VP24, VP35, and VP40 also made CD8+ T cells to NP, but rarely to GP. We identified 34 CD8+ T cell epitopes for Ebola. Our data indicate the immunodominance of the EBOV NP-specific T cell response and suggest that its inclusion in a vaccine along with the EBOV GP would best mimic survivor responses and help boost cell-mediated immunity during vaccination.
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Anticuerpos Antivirales/inmunología , Linfocitos T CD8-positivos/inmunología , Ebolavirus/inmunología , Epidemias , Antígenos HLA/inmunología , Fiebre Hemorrágica Ebola/inmunología , Adolescente , Adulto , Anticuerpos Antivirales/sangre , Antígenos Virales/inmunología , Epítopos de Linfocito T/inmunología , Femenino , Antígenos HLA/sangre , Fiebre Hemorrágica Ebola/sangre , Fiebre Hemorrágica Ebola/epidemiología , Fiebre Hemorrágica Ebola/prevención & control , Humanos , Masculino , Nucleoproteínas/inmunología , Sierra Leona , Sobrevivientes , Vacunación/métodos , Proteínas Virales/inmunología , Adulto JovenRESUMEN
The human cytomegalovirus opening reading frame UL144 is an ortholog of the TNF receptor superfamily member, herpesvirus entry mediator (HVEM; TNFRSF14). HVEM binds the TNF ligands, LIGHT and LTa; the immunoglobulin inhibitory receptor, B and T lymphocyte attenuator (BTLA); and the natural killer cell-activating receptor CD160. However, UL144 selectively binds BTLA, avoiding activation of inflammatory signaling initiated by CD160 in natural killer cells. BTLA and CD160 cross-compete for binding HVEM, but the structural basis for the ligand selectivity by UL144 and how it acts as an anti-inflammatory agonist remains unclear. Here, we modeled the UL144 structure and characterized its binding with BTLA. The UL144 structure was predicted to closely mimic the surface of HVEM, and we also found that both HVEM and UL144 bind a common epitope of BTLA, whether engaged in trans or in cis, that is shared with a BTLA antibody agonist. On the basis of the UL144 selectivity, we engineered a BTLA-selective HVEM protein to understand the basis for ligand selectivity and BTLA agonism to develop novel anti-inflammatory agonists. This HVEM mutein did not bind CD160 or TNF ligands but did bind BTLA with 10-fold stronger affinity than wild-type HVEM and retained potent inhibitory activity that reduced T-cell receptor, B-cell receptor, and interferon signaling in B cells. In conclusion, using a viral immune evasion strategy that shows broad immune-ablating activity, we have identified a novel anti-inflammatory BTLA-selective agonist.
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Linfocitos B/metabolismo , Células Asesinas Naturales/metabolismo , Glicoproteínas de Membrana/metabolismo , Modelos Moleculares , Receptores Inmunológicos/agonistas , Miembro 14 de Receptores del Factor de Necrosis Tumoral/metabolismo , Linfocitos T/metabolismo , Proteínas Virales/metabolismo , Sustitución de Aminoácidos , Antiinflamatorios no Esteroideos/química , Antiinflamatorios no Esteroideos/metabolismo , Antiinflamatorios no Esteroideos/farmacología , Antígenos CD/química , Antígenos CD/genética , Antígenos CD/metabolismo , Linfocitos B/citología , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Sitios de Unión , Línea Celular Tumoral , Diseño de Fármacos , Proteínas Ligadas a GPI/química , Proteínas Ligadas a GPI/genética , Proteínas Ligadas a GPI/metabolismo , Células HEK293 , Humanos , Células Asesinas Naturales/citología , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/inmunología , Cinética , Ligandos , Glicoproteínas de Membrana/química , Glicoproteínas de Membrana/genética , Mutación , Conformación Proteica , Ingeniería de Proteínas , Dominios y Motivos de Interacción de Proteínas , Receptores Inmunológicos/química , Receptores Inmunológicos/genética , Receptores Inmunológicos/metabolismo , Miembro 14 de Receptores del Factor de Necrosis Tumoral/química , Miembro 14 de Receptores del Factor de Necrosis Tumoral/genética , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Recombinantes de Fusión/farmacología , Linfocitos T/citología , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Proteínas Virales/química , Proteínas Virales/genéticaRESUMEN
Blockade of IFN-α but not IFN-ß signaling using either an antibody or a selective S1PR1 agonist, CYM-5442, prevented type 1 diabetes (T1D) in the mouse Rip-LCMV T1D model. First, treatment with antibody or CYM-5442 limited the migration of autoimmune "antiself" T cells to the external boundaries around the islets and prevented their entry into the islets so they could not be positioned to engage, kill, and thus remove insulin-producing ß cells. Second, CYM-5442 induced an exhaustion signature in antiself T cells by up-regulating the negative immune regulator receptor genes Pdcd1, Lag3, Ctla4, Tigit, and Btla, thereby limiting their killing ability. By such means, insulin production was preserved and glucose regulation maintained, and a mechanism for S1PR1 immunomodulation described.
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Diabetes Mellitus Tipo 1/prevención & control , Indanos/administración & dosificación , Interferón-alfa/metabolismo , Oxadiazoles/administración & dosificación , Estado Prediabético/tratamiento farmacológico , Receptores de Lisoesfingolípidos/agonistas , Linfocitos T/efectos de los fármacos , Animales , Diabetes Mellitus Tipo 1/inmunología , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Indanos/farmacología , Insulina/metabolismo , Células Secretoras de Insulina/inmunología , Islotes Pancreáticos/inmunología , Ratones , Oxadiazoles/farmacología , Estado Prediabético/inmunología , Receptores Inmunológicos/metabolismo , Transducción de Señal/efectos de los fármacos , Receptores de Esfingosina-1-Fosfato , Linfocitos T/inmunologíaRESUMEN
Innate lymphoid cells encompass a diverse array of lymphocyte subsets with unique phenotype that initiate inflammation and provide host defenses in specific microenvironments. In this study, we identify a rare human CD4(+)CD3(-) innate-like lymphoid population with high TNF expression that is enriched in blood from patients with rheumatoid arthritis. These CD4(+)CD3(-) cells belong to the T cell lineage, but the lack of AgR at the cell surface renders them nonresponsive to TCR-directed stimuli. By developing a culture system that sustains survival, we show that CD4(+)CD3(-) innate-like T cells display IL-7-dependent induction of surface lymphotoxin-αß, demonstrating their potential to modify tissue microenvironments. Furthermore, expression of CCR6 on the CD4(+)CD3(-) population defines a CD127(high) subset that is highly responsive to IL-7. This CD4(+)CD3(-) population is enriched in the peripheral blood from rheumatoid arthritis patients, suggesting a link to their involvement in chronic inflammatory disease.
Asunto(s)
Artritis Reumatoide/metabolismo , Complejo CD3/metabolismo , Linfocitos T CD4-Positivos/metabolismo , Heterotrímero de Linfotoxina alfa1 y beta2/metabolismo , Factores de Necrosis Tumoral/metabolismo , Adulto , Anciano , Animales , Artritis Reumatoide/inmunología , Linfocitos T CD4-Positivos/inmunología , Línea Celular , Femenino , Células HEK293 , Proteínas de Homeodominio/genética , Humanos , Inflamación , Interleucina-7/metabolismo , Subunidad alfa del Receptor de Interleucina-7/metabolismo , Activación de Linfocitos/inmunología , Recuento de Linfocitos , Heterotrímero de Linfotoxina alfa1 y beta2/inmunología , Masculino , Ratones , Ratones Noqueados , Persona de Mediana Edad , Receptores CCR6/metabolismoRESUMEN
Lymphocyte activation is regulated by costimulatory and inhibitory receptors, of which both B and T lymphocyte attenuator (BTLA) and CD160 engage herpesvirus entry mediator (HVEM). Notably, it remains unclear how HVEM functions with each of its ligands during immune responses. In this study, we show that HVEM specifically activates CD160 on effector NK cells challenged with virus-infected cells. Human CD56(dim) NK cells were costimulated specifically by HVEM but not by other receptors that share the HVEM ligands LIGHT, Lymphotoxin-α, or BTLA. HVEM enhanced human NK cell activation by type I IFN and IL-2, resulting in increased IFN-γ and TNF-α secretion, and tumor cell-expressed HVEM activated CD160 in a human NK cell line, causing rapid hyperphosphorylation of serine kinases ERK1/2 and AKT and enhanced cytolysis of target cells. In contrast, HVEM activation of BTLA reduced cytolysis of target cells. Together, our results demonstrate that HVEM functions as a regulator of immune function that activates NK cells via CD160 and limits lymphocyte-induced inflammation via association with BTLA.