RESUMEN
BACKGROUND: APOBEC3/Rfv3 restricts acute Friend retrovirus (FV) infection and promotes virus-specific neutralizing antibody (NAb) responses. Classical Rfv3 studies utilized FV stocks containing lactate-dehydrogenase elevating virus (LDV), a potent type I interferon inducer. Previously, we showed that APOBEC3 is required for the anti-FV activity of exogenous IFN-alpha treatment. Thus, type I interferon receptor (IFNAR) signaling may be required for the APOBEC3/Rfv3 response. RESULTS: To test if the APOBEC3/Rfv3 response is dependent on type I IFN signaling, we infected IFNAR knockout versus IFNAR/APOBEC3 double-knockout mice with FV/LDV or LDV-free FV, and evaluated acute FV infection and subsequent NAb titers. We show that LDV co-infection and type I IFN signaling are not required for innate APOBEC3-mediated restriction. By contrast, removal of LDV and/or type I IFN signaling abrogated the APOBEC3-dependent NAb response. CONCLUSIONS: APOBEC3 can restrict retroviruses in a type I IFN-independent manner in vivo. By contrast, the ability of APOBEC3 to promote NAb responses is type I IFN-dependent. These findings reveal novel insights on the interplay between type I IFNs and APOBEC3 in vivo that may have implications for augmenting antiretroviral NAb responses.
Asunto(s)
Anticuerpos Neutralizantes/sangre , Anticuerpos Antivirales/sangre , Citidina Desaminasa/metabolismo , Virus de la Leucemia Murina de Friend/inmunología , Interferón Tipo I/metabolismo , Transducción de Señal , Replicación Viral , Animales , Virus de la Leucemia Murina de Friend/fisiología , Ratones Noqueados , Receptor de Interferón alfa y beta/deficienciaRESUMEN
Somatic hypermutation (SHM) is an integral process in the development of high-affinity antibodies that are important for recovery from viral infections and vaccine-induced protection. Ig SHM occurs predominantly in germinal centers (GC) via the enzymatic activity of activation-induced deaminase (AID). In contrast, the evolutionarily related apolipoprotein B mRNA-editing enzyme, catalytic polypeptide 3 (APOBEC3) proteins are known to restrict retroviruses, including HIV-1. We previously reported that mouse APOBEC3 encodes Recovery from Friend virus 3 (Rfv3), a classical resistance gene in mice that promotes the neutralizing antibody response against retrovirus infection. We now show that APOBEC3/Rfv3 complements AID in driving Ig SHM during retrovirus infection. Analysis of antibody sequences from retrovirus-specific hybridomas and GC B cells from infected mice revealed Ig heavy-chain V genes with significantly increased C-to-T and G-to-A transitions in wild-type as compared with APOBEC3-defective mice. The context of the mutations was consistent with APOBEC3 but not AID mutational activity. These findings help explain the role of APOBEC3/Rfv3 in promoting the neutralizing antibody responses essential for recovery from retroviral infection and highlight APOBEC3-mediated deamination as a previously unidentified mechanism for antibody diversification in vivo.
Asunto(s)
Anticuerpos Neutralizantes/genética , Anticuerpos Antivirales/genética , Citidina Desaminasa/inmunología , Infecciones por Retroviridae/genética , Hipermutación Somática de Inmunoglobulina/inmunología , Animales , Anticuerpos Neutralizantes/inmunología , Linfocitos B/inmunología , Secuencia de Bases , Citidina Desaminasa/genética , Centro Germinal/citología , Ratones , Datos de Secuencia Molecular , Análisis de Secuencia de ADN , Hipermutación Somática de Inmunoglobulina/genética , Bazo/citologíaRESUMEN
Antiretroviral neutralizing antibody (NAb) responses are often evaluated in the absence of Fc-dependent immune effectors. In murine Friend retrovirus infection, Apobec3/Rfv3 promotes a potent polyclonal NAb response. Here, we show that the Apobec3/Rfv3-dependent NAb response correlated with virus-specific IgG2 titers and that the in vivo neutralization potency of Apobec3/Rfv3-resistant antisera was dependent on activating Fcγ receptors but not complement. The data strengthen retroviral vaccine strategies aimed at eliciting NAbs that activate specific Fcγ receptors.
Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Citidina Desaminasa/metabolismo , Virus de la Leucemia Murina de Friend/inmunología , Fragmentos Fc de Inmunoglobulinas/metabolismo , Animales , Inmunoglobulina G/inmunología , Ratones Endogámicos C57BLRESUMEN
Tetherin/BST-2 is a host restriction factor that could directly inhibit retroviral particle release by tethering nascent virions to the plasma membrane. However, the immunological impact of Tetherin during retrovirus infection remains unknown. We now show that Tetherin influences antiretroviral cell-mediated immune responses. In contrast to the direct antiviral effects of Tetherin, which are dependent on cell surface expression, the immunomodulatory effects are linked to the endocytosis of the molecule. Mice encoding endocytosis-competent C57BL/6 Tetherin exhibited lower viremia and pathology at 7 d postinfection with Friend retrovirus (FV) compared with mice encoding endocytosis-defective NZW/LacJ Tetherin. Notably, antiretroviral protection correlated with stronger NK cell responses. In addition, Friend retrovirus infection levels were significantly lower in wild-type C57BL/6 mice than in Tetherin knockout mice at 2 wk postinfection, and antiretroviral protection correlated with stronger NK cell and virus-specific CD8+ T cell responses. The results demonstrate that Tetherin acts as a modulator of the cell-mediated immune response against retrovirus infection in vivo.
Asunto(s)
Antígenos CD/inmunología , Linfocitos T CD8-positivos/inmunología , Virus de la Leucemia Murina de Friend/inmunología , Inmunidad Celular , Células Asesinas Naturales/inmunología , Glicoproteínas de Membrana/inmunología , Infecciones por Retroviridae/inmunología , Infecciones Tumorales por Virus/inmunología , Animales , Antígenos CD/genética , Linfocitos T CD8-positivos/patología , Virus de la Leucemia Murina de Friend/genética , Células Asesinas Naturales/patología , Glicoproteínas de Membrana/genética , Ratones , Ratones Noqueados , Infecciones por Retroviridae/genética , Infecciones por Retroviridae/patología , Factores de Tiempo , Infecciones Tumorales por Virus/genética , Infecciones Tumorales por Virus/patología , Viremia/genética , Viremia/inmunología , Viremia/patologíaRESUMEN
B cell functional defects are associated with delayed neutralizing antibody development in pathogenic lentivirus infections. However, the timeframe for alterations in the antibody repertoire and somatic hypermutation (SHM) remains unclear. Here, we utilized the SIV/rhesus macaque (RM) model to investigate the dynamics of immunoglobulin V(H) gene diversity and SHM following infection. Three RMs were infected with SIVmac239, and V(H)1, V(H)3, and V(H)4 genes were amplified from peripheral blood at 0, 2, 6, 24, and 36 weeks postinfection for next-generation sequencing. Analysis of over 3.8 million sequences against currently available RM germline V(H) genes revealed a highly biased V(H) gene repertoire in outbred RMs. SIV infection did not significantly perturb the predominant IgG1 response, but overall immunoglobulin SHM declined during the course of SIV infection. Moreover, SHM at the AID deamination hotspot, WRC, rapidly decreased and was suppressed throughout SIV infection. In contrast, a transient increase in mutations at the APOBEC3G deamination hotspot, CCC, coincided with a spike in APOBEC3G expression during acute SIV infection. The results outline a timetable for altered V(H) gene repertoire and IgG SHM in the SIV/RM model and suggest a burst of APOBEC3G-mediated antibody SHM during acute SIV infection.
Asunto(s)
Linfocitos B/inmunología , Cadenas Pesadas de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/genética , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Hipermutación Somática de Inmunoglobulina/genética , Animales , Diversidad de Anticuerpos , Secuencia de Bases , Citidina Desaminasa/genética , Frecuencia de los Genes , Variación Genética/inmunología , Secuenciación de Nucleótidos de Alto Rendimiento , Inmunoglobulina G/genética , Inmunoglobulina G/inmunología , Inmunoglobulina M/genética , Inmunoglobulina M/inmunología , Macaca mulatta , Análisis de Secuencia de ADN , Virus de la Inmunodeficiencia de los Simios/inmunologíaRESUMEN
Members of the APOBEC3 family of deoxycytidine deaminases counteract a broad range of retroviruses in vitro through an indirect mechanism that requires virion incorporation and inhibition of reverse transcription and/or hypermutation of minus strand transcripts in the next target cell. The selective advantage to the host of this indirect restriction mechanism remains unclear, but valuable insights may be gained by studying APOBEC3 function in vivo. Apobec3 was previously shown to encode Rfv3, a classical resistance gene that controls the recovery of mice from pathogenic Friend retrovirus (FV) infection by promoting a more potent neutralizing antibody (NAb) response. The underlying mechanism does not involve a direct effect of Apobec3 on B cell function. Here we show that while Apobec3 decreased titers of infectious virus during acute FV infection, plasma viral RNA loads were maintained, indicating substantial release of noninfectious particles in vivo. The lack of plasma virion infectivity was associated with a significant post-entry block during early reverse transcription rather than G-to-A hypermutation. The Apobec3-dependent NAb response correlated with IgG binding titers against native, but not detergent-lysed virions. These findings indicate that innate Apobec3 restriction promotes NAb responses by maintaining high concentrations of virions with native B cell epitopes, but in the context of low virion infectivity. Finally, Apobec3 restriction was found to be saturable in vivo, since increasing FV inoculum doses resulted in decreased Apobec3 inhibition. By analogy, maximizing the release of noninfectious particles by modulating APOBEC3 expression may improve humoral immunity against pathogenic human retroviral infections.
Asunto(s)
Anticuerpos Neutralizantes/inmunología , Formación de Anticuerpos , Citidina Desaminasa/metabolismo , Virus de la Leucemia Murina de Friend/patogenicidad , Virión/patogenicidad , Animales , Antivirales/farmacología , Citidina Desaminasa/genética , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Fenotipo , Plasma/virología , Regiones Promotoras Genéticas , ARN Viral/genética , ARN Viral/metabolismo , Transcripción Reversa , Análisis de Secuencia de ARN , Carga ViralRESUMEN
APOBEC1 is a cytidine deaminase involved in cholesterol metabolism that has been linked to retrovirus restriction, analogous to the evolutionarily-related APOBEC3 proteins. In particular, murine APOBEC1 was shown to inhibit Friend retrovirus (FV) in vitro, generating high levels of C-to-T and G-to-A mutations. These observations raised the possibility that FV infection might be altered in APOBEC1-null mice. To examine this question directly, we infected wild-type and APOBEC1-null mice with FV complex and evaluated acute infection levels. Surprisingly, APOBEC1-null mice exhibited similar cellular infection levels and plasma viremia relative to wild-type mice. Moreover, next-generation sequencing analyses revealed that in contrast to APOBEC3, APOBEC1 did not enhance retroviral C-to-T and G-to-A mutational frequencies in genomic DNA. Thus, APOBEC1 neither inhibited nor significantly drove the molecular evolution of FV in vivo. Our findings reinforce that not all retrovirus restriction factors characterized as potent in vitro may be functionally relevant in vivo.
Asunto(s)
Citidina Desaminasa/metabolismo , Virus de la Leucemia Murina de Friend/fisiología , Leucemia Experimental/virología , Infecciones por Retroviridae/virología , Infecciones Tumorales por Virus/virología , Desaminasas APOBEC-1 , Animales , Citidina Desaminasa/genética , Virus de la Leucemia Murina de Friend/genética , Regulación de la Expresión Génica/fisiología , Leucemia Experimental/genética , Leucemia Experimental/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Mutación , Infecciones por Retroviridae/genética , Infecciones por Retroviridae/metabolismo , Infecciones Tumorales por Virus/genética , Infecciones Tumorales por Virus/metabolismo , Carga Viral , Viremia , Replicación ViralRESUMEN
Understanding the host genetics of the immune response in retrovirus infection models could provide insights for basic HIV vaccine discovery. In Friend retrovirus (FV) infection of mice, Fv1 differentially inhibits N-tropic versus B-tropic FV infection by mediating a capsid-dependent post-entry block, Fv2 susceptibility governs splenomegaly induction, and Rfv3 resistance primes a stronger neutralizing antibody response due to more potent Apobec3 activity. Apobec3 polymorphisms in inbred mouse strains correlate with Rfv3 resistance and susceptibility, with one unresolved exception. The 129/OlaHsd (129P2) mouse strain is Fv2 and Rfv3 susceptible based on genotyping, but infection of 129P2 mice with B-tropic FV resulted in strong neutralizing antibody responses and no splenomegaly. Here we confirm that 129P2 mice are Fv1(nr/nr), explaining its resistance to B-tropic FV. Infection of 129P2 mice with NB-tropic FV, which can efficiently infect mice independent of Fv1 genotype, resulted in severe splenomegaly, high levels of viremia and weak neutralizing antibody responses regardless of Apobec3 status. Notably, high-dose B-tropic FV infection of 129P2 Apobec3-deficient mice induced significant adaptive immune responses and conferred high levels of protection following challenge with pathogenic NB-tropic FV. This immunological protection complemented previous studies that N-tropic FV can act as a live-attenuated vaccine in Fv1 (b/b) mice. Altogether, the results obtained in 129P2 mice strengthen the conclusion that Rfv3 is encoded by Apobec3, and highlight Fv1 incompatibility as a retroviral vaccine paradigm in mice. Due to its susceptibility to disease that allows for pathogenic challenge studies, B-tropic FV infection of 129P2 mice may be a useful model to study the immunological pathways induced by retroviral capsid restriction.
Asunto(s)
Citidina Desaminasa/deficiencia , Proteínas/metabolismo , Animales , Línea Celular , Citometría de Flujo , Genotipo , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Proteínas/genética , Infecciones por Retroviridae/inmunologíaRESUMEN
Major conceptual roadblocks impede the development of an HIV-1 vaccine that can stimulate a potent neutralizing antibody response. Animal models that support HIV-1 replication and allow for host genetic manipulation would be an ideal platform for testing various immunological hypotheses, but progress on this research front has been slow and disappointing. In contrast, many valuable concepts emerged from more than 50 years of studying the Friend retrovirus model. This was recently exemplified by the identification of an innate restriction gene, Apobec3, that could promote the retrovirus-specific neutralizing antibody response. Here we review both classical and recent data on humoral immunity against Friend retrovirus infection, and highlight the potential of this model for unraveling novel aspects of the retrovirus-specific antibody response that may guide HIV-1 vaccine development efforts.
Asunto(s)
Virus de la Leucemia Murina de Friend/inmunología , Infecciones por Retroviridae/inmunología , Animales , Modelos Animales de Enfermedad , Humanos , Inmunidad HumoralRESUMEN
Ribonuclease L (RNase L) is a type I interferon regulated factor that can significantly inhibit retroviruses in vitro and may activate cytoplasmic sensing pathways to augment adaptive immunity. However, the antiretroviral activity of RNase L remains to be validated in vivo. We investigated the role of RNaseL in counteracting Friend retrovirus (FV) infection relative to a well-described restriction factor, Apobec3. C57BL/6 wild-type (WT) and RNaseL knock-out (KO) mice exhibited similar acute FV infection levels despite significant transcriptional induction of oligoadenylate synthetase 1, which produces activators of RNase L. Apobec3 KO mice showed higher FV infection levels relative to WT mice, but deletion of RNaseL in Apobec3 KO mice did not augment FV infection. Moreover, RNaseL did not influence FV-specific IgG responses and recovery from viremia by 28 days post-infection. The results suggest that RNase L is not an evolutionarily-conserved host defense mechanism to counteract retroviruses in vivo.