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1.
Eur J Immunol ; 45(8): 2365-76, 2015 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-25912253

RESUMEN

Activation induced deaminase (AID) initiates somatic hypermutation and class switch recombination of the Ig genes in antigen-activated B cells, underpinning antibody affinity maturation and isotype switching. AID can also be pathogenic by contributing to autoimmune diseases and oncogenic mutations. Moreover, AID can exert noncanonical functions when aberrantly expressed in epithelial cells. The lack of specific inhibitors prevents therapeutic applications to modulate AID functions. Here, we have exploited our previous finding that the HSP90 molecular chaperoning pathway stabilizes AID in B cells, to test whether HSP90 inhibitors could target AID in vivo. We demonstrate that chronic administration of HSP90 inhibitors decreases AID protein levels and isotype switching in immunized mice. HSP90 inhibitors also reduce disease severity in a mouse model of acute B-cell lymphoblastic leukemia in which AID accelerates disease progression. We further show that human AID protein levels are sensitive to HSP90 inhibition in normal and leukemic B cells, and that HSP90 inhibition prevents AID-dependent epithelial to mesenchymal transition in a human breast cancer cell line in vitro. Thus, we provide proof-of-concept that HSP90 inhibitors indirectly target AID in vivo and that endogenous human AID is widely sensitive to them, which could have therapeutic applications.


Asunto(s)
Linfocitos B/inmunología , Neoplasias de la Mama/inmunología , Citidina Desaminasa/inmunología , Proteínas HSP90 de Choque Térmico/inmunología , Proteínas de Neoplasias/inmunología , Neoplasias Experimentales/inmunología , Leucemia-Linfoma Linfoblástico de Células Precursoras/inmunología , Animales , Linfocitos B/patología , Neoplasias de la Mama/patología , Línea Celular Tumoral , Transición Epitelial-Mesenquimal/inmunología , Femenino , Humanos , Ratones , Ratones Noqueados , Neoplasias Experimentales/patología , Leucemia-Linfoma Linfoblástico de Células Precursoras/patología
2.
J Exp Med ; 220(9)2023 09 04.
Artículo en Inglés | MEDLINE | ID: mdl-37310381

RESUMEN

Positively selected germinal center B cells (GCBC) can either resume proliferation and somatic hypermutation or differentiate. The mechanisms dictating these alternative cell fates are incompletely understood. We show that the protein arginine methyltransferase 1 (Prmt1) is upregulated in murine GCBC by Myc and mTORC-dependent signaling after positive selection. Deleting Prmt1 in activated B cells compromises antibody affinity maturation by hampering proliferation and GCBC light zone to dark zone cycling. Prmt1 deficiency also results in enhanced memory B cell generation and plasma cell differentiation, albeit the quality of these cells is compromised by the GCBC defects. We further demonstrate that Prmt1 intrinsically limits plasma cell differentiation, a function co-opted by B cell lymphoma (BCL) cells. Consistently, PRMT1 expression in BCL correlates with poor disease outcome, depends on MYC and mTORC1 activity, is required for cell proliferation, and prevents differentiation. Collectively, these data identify PRMT1 as a determinant of normal and cancerous mature B cell proliferation and differentiation balance.


Asunto(s)
Linfocitos B , Proteína-Arginina N-Metiltransferasas , Animales , Ratones , Afinidad de Anticuerpos , Diferenciación Celular , Centro Germinal , Proteína-Arginina N-Metiltransferasas/genética , Proliferación Celular
3.
Sci Immunol ; 6(55)2021 01 08.
Artículo en Inglés | MEDLINE | ID: mdl-33419790

RESUMEN

Influenza is a deadly and costly infectious disease, even during flu seasons when an effective vaccine has been developed. To improve vaccines against respiratory viruses, a better understanding of the immune response at the site of infection is crucial. After influenza infection, clonally expanded T cells take up permanent residence in the lung, poised to rapidly respond to subsequent infection. Here, we characterized the dynamics and transcriptional regulation of lung-resident CD4+ T cells during influenza infection and identified a long-lived, Bcl6-dependent population that we have termed T resident helper (TRH) cells. TRH cells arise in the lung independently of lymph node T follicular helper cells but are dependent on B cells, with which they tightly colocalize in inducible bronchus-associated lymphoid tissue (iBALT). Deletion of Bcl6 in CD4+ T cells before heterotypic challenge infection resulted in redistribution of CD4+ T cells outside of iBALT areas and impaired local antibody production. These results highlight iBALT as a homeostatic niche for TRH cells and advocate for vaccination strategies that induce TRH cells in the lung.


Asunto(s)
Vacunas contra la Influenza/inmunología , Gripe Humana/inmunología , Subgrupos de Linfocitos T/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Animales , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunidad Mucosa , Virus de la Influenza A/inmunología , Vacunas contra la Influenza/administración & dosificación , Gripe Humana/prevención & control , Gripe Humana/virología , Pulmón/inmunología , Pulmón/patología , Pulmón/virología , Masculino , Ratones , Ratones Transgénicos , Proteínas Proto-Oncogénicas c-bcl-6/genética , Proteínas Proto-Oncogénicas c-bcl-6/metabolismo , Subgrupos de Linfocitos T/metabolismo , Linfocitos T Colaboradores-Inductores/metabolismo
4.
Sci Immunol ; 5(45)2020 03 06.
Artículo en Inglés | MEDLINE | ID: mdl-32144185

RESUMEN

CD4+ memory T cells play an important role in protective immunity and are a key target in vaccine development. Many studies have focused on T central memory (Tcm) cells, whereas the existence and functional significance of long-lived T follicular helper (Tfh) cells are controversial. Here, we show that Tfh cells are highly susceptible to NAD-induced cell death (NICD) during isolation from tissues, leading to their underrepresentation in prior studies. NICD blockade reveals the persistence of abundant Tfh cells with high expression of hallmark Tfh markers to at least 400 days after infection, by which time Tcm cells are no longer found. Using single-cell RNA-seq, we demonstrate that long-lived Tfh cells are transcriptionally distinct from Tcm cells, maintain stemness and self-renewal gene expression, and, in contrast to Tcm cells, are multipotent after recall. At the protein level, we show that folate receptor 4 (FR4) robustly discriminates long-lived Tfh cells from Tcm cells. Unexpectedly, long-lived Tfh cells concurrently express a distinct glycolytic signature similar to trained immune cells, including elevated expression of mTOR-, HIF-1-, and cAMP-regulated genes. Late disruption of glycolysis/ICOS signaling leads to Tfh cell depletion concomitant with decreased splenic plasma cells and circulating antibody titers, demonstrating both unique homeostatic regulation of Tfh and their sustained function during the memory phase of the immune response. These results highlight the metabolic heterogeneity underlying distinct long-lived T cell subsets and establish Tfh cells as an attractive target for the induction of durable adaptive immunity.


Asunto(s)
Inmunidad Humoral/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Animales , Muerte Celular/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , NAD/farmacología , Receptores Purinérgicos P2X7/deficiencia , Receptores Purinérgicos P2X7/inmunología , Linfocitos T Colaboradores-Inductores/efectos de los fármacos
5.
Nat Commun ; 10(1): 22, 2019 01 03.
Artículo en Inglés | MEDLINE | ID: mdl-30604754

RESUMEN

Mechanisms regulating B cell development, activation, education in the germinal center (GC) and differentiation, underpin the humoral immune response. Protein arginine methyltransferase 5 (Prmt5), which catalyzes most symmetric dimethyl arginine protein modifications, is overexpressed in B cell lymphomas but its function in normal B cells is poorly defined. Here we show that Prmt5 is necessary for antibody responses and has essential but distinct functions in all proliferative B cell stages in mice. Prmt5 is necessary for B cell development by preventing p53-dependent and p53-independent blocks in Pro-B and Pre-B cells, respectively. By contrast, Prmt5 protects, via p53-independent pathways, mature B cells from apoptosis during activation, promotes GC expansion, and counters plasma cell differentiation. Phenotypic and RNA-seq data indicate that Prmt5 regulates GC light zone B cell fate by regulating transcriptional programs, achieved in part by ensuring RNA splicing fidelity. Our results establish Prmt5 as an essential regulator of B cell biology.


Asunto(s)
Linfocitos B/fisiología , Proliferación Celular/fisiología , Centro Germinal/fisiología , Inmunidad Humoral/fisiología , Proteína-Arginina N-Metiltransferasas/fisiología , Animales , Apoptosis/inmunología , Linfocitos B/inmunología , Linfocitos B/metabolismo , Diferenciación Celular/inmunología , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Técnicas de Silenciamiento del Gen , Centro Germinal/citología , Humanos , Activación de Linfocitos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Cultivo Primario de Células , Proteína-Arginina N-Metiltransferasas/genética , Proteína-Arginina N-Metiltransferasas/metabolismo , Transducción de Señal/fisiología , Trichostrongyloidea/inmunología , Tricostrongiloidiasis/inmunología , Tricostrongiloidiasis/parasitología , Proteína p53 Supresora de Tumor/metabolismo
6.
Nat Commun ; 9(1): 1248, 2018 03 28.
Artículo en Inglés | MEDLINE | ID: mdl-29593215

RESUMEN

Activation-induced deaminase (AID) mutates the immunoglobulin (Ig) genes to initiate somatic hypermutation (SHM) and class switch recombination (CSR) in B cells, thus underpinning antibody responses. AID mutates a few hundred other loci, but most AID-occupied genes are spared. The mechanisms underlying productive deamination versus non-productive AID targeting are unclear. Here we show that three clustered arginine residues define a functional AID domain required for SHM, CSR, and off-target activity in B cells without affecting AID deaminase activity or Escherichia coli mutagenesis. Both wt AID and mutants with single amino acid replacements in this domain broadly associate with Spt5 and chromatin and occupy the promoter of AID target genes. However, mutant AID fails to occupy the corresponding gene bodies and loses association with transcription elongation factors. Thus AID mutagenic activity is determined not by locus occupancy but by a licensing mechanism, which couples AID to transcription elongation.


Asunto(s)
Linfocitos B/metabolismo , Citidina Desaminasa/metabolismo , Cambio de Clase de Inmunoglobulina , Mutagénesis , Elongación de la Transcripción Genética , Animales , Arginina/química , Línea Celular Tumoral , Cromatina/química , ADN/química , Desaminación , Escherichia coli/metabolismo , Genes de Inmunoglobulinas , Humanos , Inmunoglobulinas/química , Lipopolisacáridos/química , Ratones , Microscopía Confocal , Mutación , Dominios Proteicos , Hipermutación Somática de Inmunoglobulina , Transcripción Genética
7.
J Exp Med ; 212(4): 581-96, 2015 Apr 06.
Artículo en Inglés | MEDLINE | ID: mdl-25824822

RESUMEN

Activation-induced deaminase (AID) initiates mutagenic pathways to diversify the antibody genes during immune responses. The access of AID to the nucleus is limited by CRM1-mediated nuclear export and by an uncharacterized mechanism of cytoplasmic retention. Here, we define a conformational motif in AID that dictates its cytoplasmic retention and demonstrate that the translation elongation factor eukaryotic elongation factor 1 α (eEF1A) is necessary for AID cytoplasmic sequestering. The mechanism is independent of protein synthesis but dependent on a tRNA-free form of eEF1A. Inhibiting eEF1A prevents the interaction with AID, which accumulates in the nucleus and increases class switch recombination as well as chromosomal translocation byproducts. Most AID is associated to unspecified cytoplasmic complexes. We find that the interactions of AID with eEF1A and heat-shock protein 90 kD (HSP90) are inversely correlated. Despite both interactions stabilizing AID, the nature of the AID fractions associated with HSP90 or eEF1A are different, defining two complexes that sequentially produce and store functional AID in the cytoplasm. In addition, nuclear export and cytoplasmic retention cooperate to exclude AID from the nucleus but might not be functionally equivalent. Our results elucidate the molecular basis of AID cytoplasmic retention, define its functional relevance and distinguish it from other mechanisms regulating AID.


Asunto(s)
Citidina Desaminasa/metabolismo , Citoplasma/metabolismo , Factor 1 de Elongación Peptídica/metabolismo , Transporte Activo de Núcleo Celular/genética , Secuencias de Aminoácidos , Animales , Núcleo Celular/genética , Núcleo Celular/metabolismo , Citidina Desaminasa/genética , Citoplasma/genética , Células HEK293 , Humanos , Ratones , Ratones Noqueados , Complejos Multiproteicos/genética , Complejos Multiproteicos/metabolismo , Factor 1 de Elongación Peptídica/genética , Translocación Genética
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