RESUMEN
Activation-induced cytidine deaminase (AID) is responsible for the initiation of somatic hypermutation (SHM) and class-switch recombination (CSR), which result in antibody affinity maturation and isotype switching, thus producing pathogen-specific antibodies. Chromatin dynamics and accessibility play a significant role in determining AID expression and its targeting. Chromatin remodelers contribute to the accessibility of the chromatin structure, thereby influencing the targeting of AID to Ig genes. Epigenetic modifications, including DNA methylation, histone modifications, and miRNA expression, profoundly impact the regulation of AID and chromatin remodelers targeting Ig genes. Additionally, epigenetic modifications lead to chromatin rearrangement and thereby can change AID expression levels and its preferential targeting to Ig genes. This interplay is symbolized as the ACE phenomenon encapsulates three interconnected aspects: AID, Chromatin remodelers, and Epigenetic modifications. This review emphasizes the importance of understanding the intricate relationship between these aspects to unlock the therapeutic potential of these molecular processes and molecules.
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Diversidad de Anticuerpos , Ensamble y Desensamble de Cromatina , Citidina Desaminasa , Epigénesis Genética , Citidina Desaminasa/metabolismo , Citidina Desaminasa/genética , Humanos , Animales , Diversidad de Anticuerpos/genética , Cambio de Clase de Inmunoglobulina/genética , Metilación de ADN , Hipermutación Somática de Inmunoglobulina , Cromatina/metabolismo , Cromatina/genética , Regulación de la Expresión GénicaRESUMEN
BACKGROUND: RNA-DNA hybrids or R-loops are associated with deleterious genomic instability and protective immunoglobulin class switch recombination (CSR). However, the underlying phenomenon regulating the two contrasting functions of R-loops is unknown. Notably, the underlying mechanism that protects R-loops from classic RNase H-mediated digestion thereby promoting persistence of CSR-associated R-loops during CSR remains elusive. RESULTS: Here, we report that during CSR, R-loops formed at the immunoglobulin heavy (IgH) chain are modified by ribose 2'-O-methylation (2'-OMe). Moreover, we find that 2'-O-methyltransferase fibrillarin (FBL) interacts with activation-induced cytidine deaminase (AID) associated snoRNA aSNORD1C to facilitate the 2'-OMe. Moreover, deleting AID C-terminal tail impairs its association with aSNORD1C and FBL. Disrupting FBL, AID or aSNORD1C expression severely impairs 2'-OMe, R-loop stability and CSR. Surprisingly, FBL, AID's interaction partner and aSNORD1C promoted AID targeting to the IgH locus. CONCLUSION: Taken together, our results suggest that 2'-OMe stabilizes IgH-associated R-loops to enable productive CSR. These results would shed light on AID-mediated CSR and explain the mechanism of R-loop-associated genomic instability.
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Citidina Desaminasa , Cambio de Clase de Inmunoglobulina , Estructuras R-Loop , Cambio de Clase de Inmunoglobulina/genética , Citidina Desaminasa/metabolismo , Citidina Desaminasa/genética , Citidina Desaminasa/química , Animales , Ratones , Metilación , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/metabolismo , Recombinación Genética , ARN/metabolismo , ARN/genéticaRESUMEN
Activation induced cytidine deaminase (AID) is a key element of the adaptive immune system, required for immunoglobulin isotype switching and affinity maturation of B-cells as they undergo the germinal center (GC) reaction in peripheral lymphoid tissue. The inherent DNA damaging activity of this enzyme can also have off-target effects in B-cells, producing lymphomagenic chromosomal translocations that are characteristic features of various classes of non-Hodgkin B-cell lymphoma (B-NHL), and generating oncogenic mutations, so-called aberrant somatic hypermutation (aSHM). Additionally, AID has been found to affect gene expression through demethylation as well as altered interactions between gene regulatory elements. These changes have been most thoroughly studied in B-NHL arising from GC B-cells. Here, we describe the most common classes of GC-derived B-NHL and explore the consequences of on- and off-target AID activity in B and plasma cell neoplasms. The relationships between AID expression, including effects of infection and other exposures/agents, mutagenic activity and lymphoma biology are also discussed.
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Linfocitos B , Citidina Desaminasa , Centro Germinal , Linfoma de Células B , Humanos , Citidina Desaminasa/metabolismo , Citidina Desaminasa/genética , Centro Germinal/inmunología , Linfoma de Células B/genética , Linfoma de Células B/inmunología , Animales , Linfocitos B/inmunología , Hipermutación Somática de Inmunoglobulina , Regulación Neoplásica de la Expresión Génica , Cambio de Clase de InmunoglobulinaRESUMEN
PURPOSE: Molecular residual disease (MRD) is the main cause of postoperative recurrence of breast cancer. However, the baseline tumor genomic characteristics and therapeutic implications of breast cancer patients with detectable MRD after surgery are still unknown. MATERIALS AND METHODS: In this study, we enrolled 80 patients with breast cancer who underwent next-generation sequencing-based genetic testing of 1,021 cancer-related genes performed on baseline tumor and postoperative plasma, among which 18 patients had detectable MRD after surgery. RESULTS: Baseline clinical characteristics found that patients with higher clinical stages were more likely to have detectable MRD. Analysis of single nucleotide variations and small insertions/deletions in baseline tumors showed that somatic mutations in MAP3K1, ATM, FLT1, GNAS, POLD1, SPEN, and WWP2 were significantly enriched in patients with detectable MRD. Oncogenic signaling pathway analysis revealed that alteration of the Cell cycle pathway was more likely to occur in patients with detectable MRD (p=0.012). Mutational signature analysis showed that defective DNA mismatch repair and activation-induced cytidine deaminase (AID) mediated somatic hypermutation (SHM) were associated with detectable MRD. According to the OncoKB database, 77.8% (14/18) of patients with detectable MRD had U.S. Food and Drug Administration-approved mutational biomarkers and targeted therapy. CONCLUSION: Our study reports genomic characteristics of breast cancer patients with detectable MRD. The cell cycle pathway, defective DNA mismatch repair, and AID-mediated SHM were found to be the possible causes of detectable MRD. We also found the vast majority of patients with detectable MRD have the opportunity to access targeted therapy.
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Neoplasias de la Mama , Humanos , Femenino , Neoplasias de la Mama/genética , Neoplasias de la Mama/terapia , Hipermutación Somática de Inmunoglobulina , Mutación , Reparación de la Incompatibilidad de ADN , Genómica , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
Activation-induced cytidine deaminase (AID) plays a crucial role in promoting B cell diversification through somatic hypermutation (SHM) and class switch recombination (CSR). While AID is primarily associated with the physiological function of humoral immune response, it has also been linked to the initiation and progression of lymphomas. Abnormalities in AID have been shown to disrupt gene networks and signaling pathways in both B-cell and T-cell lineage lymphoblastic leukemia, although the full extent of its role in carcinogenesis remains unclear. This review proposes an alternative role for AID and explores its off-target effects in regulating tumorigenesis. In this review, we first provide an overview of the physiological function of AID and its regulation. AID plays a crucial role in promoting B cell diversification through SHM and CSR. We then discuss the off-target effects of AID, which includes inducing mutations of non-Igs, epigenetic modification, and the alternative role as a cofactor. We also explore the networks that keep AID in line. Furthermore, we summarize the off-target effects of AID in autoimmune diseases and hematological neoplasms. Finally, we assess the off-target effects of AID in solid tumors. The primary focus of this review is to understand how and when AID targets specific gene loci and how this affects carcinogenesis. Overall, this review aims to provide a comprehensive understanding of the physiological and off-target effects of AID, which will contribute to the development of novel therapeutic strategies for autoimmune diseases, hematological neoplasms, and solid tumors.
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Enfermedades Autoinmunes , Citidina Desaminasa , Neoplasias Hematológicas , Humanos , Carcinogénesis , Transformación Celular Neoplásica/genéticaRESUMEN
Previously, we established a platform for antibody/protein affinity maturation based on CHO cell display. The gene of interest was mutated by activation-induced cytidine deaminase (AID), and then, a mutation library mainly containing G/C to A/T conversion was obtained by simply proliferating cells. However, the AID-induced G/C to A/T conversion limits the diversity space of the mutation library. In contrast to AID, adenine deaminase (ADA) can convert A/T to G/C. In this study, we demonstrated that ADA could efficiently induce random A/T to G/C mutations on the target gene in the CHO cell display and could be applied in affinity maturation. Our data also showed that more mutant types were obtained through the combined use of AID and ADA, thus offering an opportunity to acquire new mutants offering higher affinities than those obtained by only using AID. Examples presented in this study showed that ADA contributed to the improvement of antibody affinity either with or without AID in CHO display. KEY POINTS: ⢠ADA is able to induce random mutations on antibody gene in mammalian cells. ⢠ADA induces mutations on A/T bases to compensate AID which can induce mutation on G/C. ⢠Combination of AID and ADA can increase mutation types and maturation efficiencies.
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Aminohidrolasas , Hidrolasas , Cricetinae , Animales , Afinidad de Anticuerpos , Mutación , Células CHO , CricetulusRESUMEN
Transcriptional factors (TFs) and their regulons make up the gene regulatory networks. Here, we developed a method based on TF-directed activation-induced cytidine deaminase (AID) mutagenesis in combination with genome sequencing, called AIDmut-Seq, to detect TF targets on the genome. AIDmut-Seq involves only three simple steps, including the expression of the AID-TF fusion protein, whole-genome sequencing, and single nucleotide polymorphism (SNP) profiling, making it easy for junior and interdisciplinary researchers to use. Using AIDmut-Seq for the major quorum sensing regulator LasR in Pseudomonas aeruginosa, we confirmed that a few TF-guided C-T (or G-A) conversions occurred near their binding boxes on the genome, and a number of previously characterized and uncharacterized LasR-binding sites were detected. Further verification of AIDmut-Seq using various transcriptional regulators demonstrated its high efficiency for most transcriptional activators (FleQ, ErdR, GacA, ExsA). We confirmed the binding of LasR, FleQ, and ErdR to 100%, 50%, and 86% of their newly identified promoters by using in vitro protein-DNA binding assay. And real-time RT-PCR data validated the intracellular activity of these TFs to regulate the transcription of those newly found target promoters. However, AIDmut-Seq exhibited low efficiency for some small transcriptional repressors such as RsaL and AmrZ, with possible reasons involving fusion-induced TF dysfunction as well as low transcription rates of target promoters. Although there are false-positive and false-negative results in the AIDmut-Seq data, preliminary results have demonstrated the value of AIDmut-Seq to act as a complementary tool for existing methods. IMPORTANCE Protein-DNA interactions (PDI) play a central role in gene regulatory networks (GRNs). However, current techniques for studying genome-wide PDI usually involve complex experimental procedures, which prevent their broad use by scientific researchers. In this study, we provide a in vivo method called AIDmut-Seq. AIDmut-Seq involves only three simple steps that are easy to operate for researchers with basic skills in molecular biology. The efficiency of AIDmut-Seq was tested and confirmed using multiple transcription factors in Pseudomonas aeruginosa. Although there are still some defects regarding false-positive and false-negative results, AIDmut-Seq will be a good choice in the early stage of PDI study.
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Proteínas Bacterianas , Factores de Transcripción , Proteínas Bacterianas/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Unión Proteica , ADN/metabolismo , Sitios de UniónRESUMEN
Sonodynamic therapy (SDT) triggered by ultrasound (US) has attracted increasing attention owing to its ability to overcome critical limitations, including low tissue-penetration depth and phototoxicity in photodynamic therapy (PDT). Biogenic metal oxide nanoparticles (NPs) have been used as anti-cancer drugs due to their biocompatibility properties with most biological systems. Here, sonosensitizer MWO4-PEG NPs (M = Fe Mn Co Ni) were synthesized as inhibitors to activation-induced cytidine deaminase (AID), thus neutralizing the extensive carcinogenesis of AID in diffuse large B-cell lymphoma (DLBCL). The physiological properties of these nanomaterials were examined using transmission electron microscopy (TEM). The inhibition of NPs to AID was primarily identified by the affinity interaction prediction between reactive oxygen species (ROS) and AID through molecular dynamics and molecular docking technology. The cell apoptosis and ROS generation in US-triggered NPs treated DLBCL cells (with high levels of AID) were also detected to indicate the sonosensitivity and toxicity of MWO4-PEG NPs to DLBCL cells. The anti-lymphoma studies using DLBCL and AID-deficient DLBCL cell lines indicated a concentration-dependent profile. The synthesized MWO4-PEG NPs in this study manifested good sonodynamic inhibitory effects to AID and well treatment for AID-positive hematopoietic cancers.
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Linfoma de Células B Grandes Difuso , Nanopartículas del Metal , Nanopartículas , Terapia por Ultrasonido , Humanos , Especies Reactivas de Oxígeno/metabolismo , Línea Celular Tumoral , Simulación del Acoplamiento Molecular , Linfoma de Células B Grandes Difuso/tratamiento farmacológico , Linfoma de Células B Grandes Difuso/metabolismo , Linfoma de Células B Grandes Difuso/patología , Óxidos , Polietilenglicoles/uso terapéuticoRESUMEN
Affinity maturation, an essential component of antibody engineering, is crucial for developing therapeutic antibodies. Cell display system coupled with somatic hypermutation (SHM) initiated by activation-induced cytidine deaminase (AID) is a commonly used technique for affinity maturation. AID introduces targeted DNA lesions into hotspots of immunoglobulin (Ig) gene loci followed by erroneous DNA repair, leading to biased mutations in the complementary determining regions. However, systems that use an in vivo mimicking mechanism often require several rounds of selection to enrich clones possessing accumulated mutations. We previously described the human ADLib® system, which features autonomous, AID-mediated diversification in Ig gene loci of a chicken B cell line DT40 and streamlines human antibody generation and optimization in one integrated platform. In this study, we further engineered DT40 capable of receiving exogenous antibody genes and examined whether the antibody could be affinity matured. The Ig genes of three representative anti-hVEGF-A antibodies originating from the human ADLib® were introduced; the resulting human IgG1 antibodies had up to 76.4-fold improvement in binding affinities (sub-picomolar KD) within just one round of optimization, owing to efficient accumulation of functional mutations. Moreover, we successfully improved the affinity of a mouse hybridoma-derived anti-hCDCP1 antibody using the engineered DT40, and the observed mutations remained effective in the post-humanized antibody as exhibited by an 8.2-fold increase of in vitro cytotoxicity without compromised physical stability. These results demonstrated the versatility of the novel B cell-based affinity maturation system as an easy-to-use antibody optimization tool regardless of the species of origin.Abbreviations: ADLib®: Autonomously diversifying library, ADLib® KI-AMP: ADLib® knock-in affinity maturation platform, AID: activation-induced cytidine deaminase, CDRs: complementary-determining regions, DIVAC: diversification activator, ECD: extracellular domain, FACS: fluorescence-activated cell sorting, FCM: flow cytometry, HC: heavy chainIg: immunoglobulin, LC: light chain, NGS: next-generation sequencing, PBD: pyrrolobenzodiazepine, SHM: somatic hypermutation, SPR: surface plasmon resonance.
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Citidina Desaminasa , Hipermutación Somática de Inmunoglobulina , Animales , Humanos , Ratones , Linfocitos B , Citidina Desaminasa/genética , Citidina Desaminasa/metabolismo , ADN , Inmunoglobulina G/genéticaRESUMEN
Activation-induced cytidine deaminase (AID) is a key player that initiates antibody diversification in activated B-cell. AID mediates somatic hypermutation (SHM) and class switch recombination (CSR) via the deamination of cytosine to uracil at the Ig locus, resulting in the production of high-affinity antibodies. AID is predominantly restricted to Ig genes, whereas off-targeting of AID leads to lymphocyte-related malignancies. Interestingly, apart from FL-AID other splice isoforms of AID are highly expressed in the lymphocyte malignancies. In our study, we found that the heterologous expression of hAID-FL in E. coli cells produced two induced bands of hAID as demonstrated by SDS-PAGE and western blotting. Remarkably, peptide mapping data predicted that one band is hAID-FL and the other is its splice isoform, hAIDδE4a. To get an insight into why E. coli cells expressed hAID-FL and hAID variant, we mutated the 5' and 3' splice site of a putative intron of hAID, but it failed to produce only hAID-FL. Incidentally, hAID expressed with fusion partners also displayed two bands, and peptide mapping data strongly suggest that besides hAID-FL, the lower band showed a significant number of amino acids missing towards the C-terminal domain (named as hAIDδC). Our results are the first report to show that expression of recombinant hAID alone or irrespective of solubilization tags in E. coli cells produced hAID-FL and hAIDδC. It will be fascinating to explore the potential mechanism underlying the expression of hAIDδC from recombinant hAID plasmid in E. coli cells.
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Citidina Desaminasa , Hipermutación Somática de Inmunoglobulina , Citidina Desaminasa/genética , Citidina Desaminasa/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Cambio de Clase de Inmunoglobulina , Isoformas de Proteínas/genéticaRESUMEN
In activated B cells, activation-induced cytidine deaminase (AID) generates programmed DNA lesions required for antibody class switch recombination (CSR), which may also threaten genome integrity. AID dynamically shuttles between cytoplasm and nucleus, and the majority stays in the cytoplasm due to active nuclear export mediated by its C-terminal peptide. In immunodeficient-patient cells expressing mutant AID lacking its C-terminus, a catalytically active AID-delC protein accumulates in the nucleus but nevertheless fails to support CSR. To resolve this apparent paradox, we dissected the function of AID-delC proteins in the CSR process and found that they cannot efficiently target antibody genes. We demonstrate that AID-delC proteins form condensates both in vivo and in vitro, dependent on its N-terminus and on a surface arginine-rich patch. Co-expression of AID-delC and wild-type AID leads to an unbalanced nuclear AID-delC/AID ratio, with AID-delC proteins able to trap wild-type AID in condensates, resulting in a dominant-negative phenotype that could contribute to immunodeficiency. The co-condensation model of mutant and wild-type proteins could be an alternative explanation for the dominant-negative effect in genetic disorders.
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Citidina Desaminasa , Cambio de Clase de Inmunoglobulina , Linfocitos B , Citidina Desaminasa/genética , Citidina Desaminasa/metabolismo , ADN/metabolismo , Humanos , Cambio de Clase de Inmunoglobulina/genéticaRESUMEN
H. pylori is one of the major causes of chronic gastritis, peptic ulcer disease (PUD), gastric mucosa-associated lymphoid tissue lymphoma (MALT) and gastric carcinoma. H. pylori toxin VacA is responsible for host cell apoptosis, whereas CagA is known to aberrantly induce expression of activation-induced cytidine deaminase (AID) in gastric epithelial cells that causes mutations in oncogenes and tumour suppressor genes, leading to the transformation of normal cells into cancerous cells. Although, a significant amount of research has been conducted to understand the role of bacterial factors modulating deregulated host cell pathways, the interaction between H. pylori and immune cells of the marginal zone and its consequences are still not well understood. HomB and HomA, outer membrane proteins (OMPs) from H. pylori, which assist in the adhesion of bacteria to host cells, are found to be associated with H. pylori virulent strains and promote inflammation. Interestingly, we observed that the interaction of HomB/HomA OMPs with B-cells transiently downregulates AID expression and Ig switch germline transcription. Downregulation of AID leads to impairment of class switch recombination (CSR), resulting in significantly reduced switching to IgG and IgA antibodies. Besides, we examined the immune-suppressive response of B-cells and observed that the cells stimulated with HomA/B show upregulation in the levels of IL10, IL35, as well as PDL1, a T-cell inhibition marker. Our study suggests the potential role of OMPs in immune response modulation strategies used by the pathogen to evade the immune response. These results provide a better understanding of H. pylori pathogenesis and assist in identifying novel targets for therapy.
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Linfocitos B/inmunología , Proteínas de la Membrana Bacteriana Externa/metabolismo , Citidina Desaminasa/metabolismo , Helicobacter pylori/inmunología , Evasión Inmune/inmunología , Cambio de Clase de Inmunoglobulina/genética , Apoptosis/fisiología , Antígeno B7-H1/metabolismo , Proteínas de la Membrana Bacteriana Externa/genética , Proteínas de la Membrana Bacteriana Externa/inmunología , Línea Celular Tumoral , Genes de Inmunoglobulinas/genética , Infecciones por Helicobacter/patología , Humanos , Inmunoglobulina A/inmunología , Inmunoglobulina G/inmunología , Interleucina-10/metabolismo , Interleucinas/metabolismoRESUMEN
Activation-induced deaminase (AID) is required for somatic hypermutation in immunoglobulin genes, but also induces off-target mutations. Follicular lymphoma (FL) and chronic lymphocytic leukemia (CLL), the most frequent types of indolent B-cell tumors, are exposed to AID activity during lymphomagenesis. We designed a workflow integrating de novo mutational signatures extraction and fitting of COSMIC (Catalogue Of Somatic Mutations In Cancer) signatures, with tridimensional chromatin conformation data (Hi-C). We applied the workflow to exome sequencing data from lymphoma samples. In 33 FL and 30 CLL samples, 42% and 34% of the contextual mutations could be traced to a known AID motif. We demonstrate that both CLL and FL share mutational processes dominated by spontaneous deamination, failures in DNA repair, and AID activity. The processes had equiproportional distribution across active and nonactive chromatin compartments in CLL. In contrast, canonical AID activity and failures in DNA repair pathways in FL were significantly higher within the active chromatin compartment. Analysis of DNA repair genes revealed a higher prevalence of base excision repair gene mutations (p = 0.02) in FL than CLL. These data indicate that AID activity drives the genetic landscapes of FL and CLL. However, the final result of AID-induced mutagenesis differs between these lymphomas depending on chromatin compartmentalization and mutations in DNA repair pathways.
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Citidina Desaminasa/genética , Leucemia Linfocítica Crónica de Células B/patología , Linfoma Folicular/patología , Alelos , Cromatina/metabolismo , Análisis Mutacional de ADN , Reparación del ADN/genética , Bases de Datos Genéticas , Frecuencia de los Genes , Humanos , Leucemia Linfocítica Crónica de Células B/genética , Linfoma Folicular/genética , Polimorfismo de Nucleótido SimpleRESUMEN
OBJECTIVE: IgG4-related disease (IgG4-RD) is a systemic condition characterized by an elevated serum IgG4 level, localized infiltration of IgG4-positive plasma cells, and lesions in various organs. IgG4-RD also shows high rates of complication with allergic diseases and is often accompanied by elevated serum IgE levels. Reports in recent years have also shown high rates of complication with chronic rhinosinusitis (CRS) and its characteristic nasal manifestations. Accordingly, we speculate that there may be a distinct form of CRS that, as an IgG4-RD, differs from other CRS. Here, we investigated whether the elevated levels of factors that are thought to be important in the pathogenesis of IgG4-RD are also seen in the sinus mucosa of IgG4-RD-associated CRS patients. METHODS: Ethmoid sinus mucosa specimens from 9 IgG4-RD (6 Mikulicz disease and 3 Küttner's tumor) patients with elevated serum IgG4 and IgE and from 22 control CRS patients were examined immunohistochemically for Treg cytokines (IL-10 and TGF-ß), activation-induced cytidine deaminase (AID), and immunocompetent cells. The 22 control CRS patients were divided into 3 subgroups based on the serological findings for IgG4 and IgE. Quantitative real-time PCR was performed to examine the expression of AID. RESULTS: The ethmoid sinus mucosa from patients with IgG4-RD-associated CRS showed, in comparison with the 3 CRS control subgroups, significantly elevated AID production. Their mucosa also showed significantly increased infiltration of CD-20-positive immunocompetent cells compared with the controls. On the other hand, immunohistochemical examination found no significant differences in the number of IL-10- or TGF-ß-positive cells. CONCLUSION: Ethmoid sinus mucosa from IgG4-RD-associated CRS patients showed clearly increased AID production, suggesting AID involvement in class-switching to IgG4 in those local sites. This implies the existence of a distinct form of CRS that is an IgG4-RD.
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Enfermedad Relacionada con Inmunoglobulina G4 , Rinitis , Sialadenitis , Sinusitis , Enfermedad Crónica , Citidina Desaminasa , Humanos , Enfermedad Relacionada con Inmunoglobulina G4/complicaciones , Rinitis/complicaciones , Sinusitis/complicacionesRESUMEN
Background/aim: Activation-induced cytidine deaminase (AID) enables antibody diversity in B lymphocytes. It may also have an effect on MDS pathogenesis by causing somatic mutations and by inducing epigenetic changes in myeloid cells. This study aimed to compare AID expression of MDS patients with healthy controls, of MDS patients in different risk groups, and of MDS patients according to their treatment. Materials and methods: Total RNA was isolated and complementary DNA (cDNA) was transcribed from the peripheral blood samples of MDS patients and healthy controls. AID and the reference gene HPRT1 were analyzed using quantitative real-time PCR (QRT-PCR). AID expression relative to HPRT1 was calculated. Patients were classified into "lower risk" and "higher risk" subgroups according to their initial IPSS and IPSS-R scores and their MDS subtypes at the time of study. Patients were also divided into two groups based on receiving treatment with hypomethylating agents. AID expressions of different groups were compared using the MannWhitney U test. Results: Thirty MDS patients and thirty healthy controls were included. AID expression in MDS patients was significantly higher compared to healthy controls (p < 0.001). There was no significant difference in AID expression of "lower risk" and "higher risk" subgroups of patients. Patients that received hypomethylating agents did not have a significant difference in AID expression compared with patients that did not receive hypomethylating agents. Conclusion: AID expression is increased in the peripheral blood of MDS patients compared to healthy controls. However, AID expression is not significantly different in "lower risk" and "higher risk" subgroups and in patients treated with hypomethylating agents. Increased AID expression may be an early step in MDS pathogenesis.
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Citidina Desaminasa/genética , Síndromes Mielodisplásicos/genética , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Femenino , Humanos , Leucemia Mieloide Aguda , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/sangre , Síndromes Mielodisplásicos/tratamiento farmacológico , PronósticoRESUMEN
Immunoglobulin G4-related disease (IgG4-RD) is a systemic disorder characterized by tissue fibrosis and intense lymphoplasmacytic infiltration, causing progressive organ dysfunction. Activation-induced cytidine deaminase (AID), a deaminase normally expressed in activated B-cells in germinal centers, edits ribonucleotides to induce somatic hypermutation and class switching of immunoglobulin. While AID expression is strictly controlled under physiological conditions, chronic inflammation has been noted to induce its upregulation to propel oncogenesis. We examined AID expression in IgG4-related ophthalmic disease (IgG4-ROD; n = 16), marginal zone lymphoma with IgG4-positive cells (IgG4+ MZL; n = 11), and marginal zone lymphoma without IgG4-positive cells (IgG4- MZL; n = 12) of ocular adnexa using immunohistochemical staining. Immunohistochemistry revealed significantly higher AID-intensity index in IgG4-ROD and IgG4+ MZL than IgG4- MZL (p < 0.001 and = 0.001, respectively). The present results suggest that IgG4-RD has several specific causes of AID up-regulation in addition to inflammation, and AID may be a driver of oncogenesis in IgG4-ROD to IgG4+ MZL.
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Citidina Desaminasa/genética , Neoplasias del Ojo/enzimología , Neoplasias del Ojo/genética , Inmunoglobulina G/metabolismo , Linfoma de Células B de la Zona Marginal/enzimología , Linfoma de Células B de la Zona Marginal/genética , Regulación hacia Arriba , Neoplasias del Ojo/patología , Femenino , Humanos , Linfoma de Células B de la Zona Marginal/patología , Masculino , Persona de Mediana Edad , Regulación hacia Arriba/genéticaRESUMEN
Activation-induced cytidine deaminase (AID) encoded by the Aicda gene initiates class-switch recombination and somatic hypermutation of immunoglobulin genes. In addition to this function, AID is also implicated in the epigenetic regulation in pluripotent stem cells and in the oncogenesis of lymphoid and non-lymphoid origins. To examine AID's role in specific cell types, we developed mouse strains of conditional knockout (Aicda-FL) and knock-in with a red fluorescent protein gene (RFP) inserted into the Aicda locus (Aicda-RFP). These two strains were obtained from a single targeting event in embryonic stem cells by a three-loxP or tri-lox strategy. Partial and complete recombination among the three loxP sites in the Aicda-RFP locus gave rise to Aicda-FL and AID-deficient loci (Aicda-KO), respectively, after mating Aicda-RFP mice with Cre-expressing mice driven by tissue-non-specific alkaline phosphate promoter. We confirmed RFP expression in B cells of germinal centers of intestine-associated lymphoid tissue. Mice homozygous for each allele were obtained and were checked for AID activity by class-switch and hypermutation assays. AID activity was normal for Aicda-FL but partially and completely absent for Aicda-RFP and Aicda-KO, respectively. Aicda-FL and Aicda-RFP mice would be useful for studying AID function in subpopulations of B cells and in non-lymphoid cells.
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Citidina Desaminasa/genética , Animales , Linfocitos B/metabolismo , Epigénesis Genética/genética , Femenino , Centro Germinal/metabolismo , Cambio de Clase de Inmunoglobulina/genética , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Regiones Promotoras Genéticas/genéticaRESUMEN
Immunoglobulins and T cell receptors (TCR) have obvious structural similarities as well as similar immunogenetic diversification and selection mechanisms. Nevertheless, the two receptor systems and the loci that encode them are distinct in humans and classical murine models, and the gene segments comprising each repertoire are mutually exclusive. Additionally, while both B and T cells employ recombination-activating genes (RAG) for primary diversification, immunoglobulins are afforded a supplementary set of activation-induced cytidine deaminase (AID)-mediated diversification tools. As the oldest-emerging vertebrates sharing the same adaptive B and T cell receptor systems as humans, extant cartilaginous fishes allow a potential view of the ancestral immune system. In this review, we discuss breakthroughs we have made in studies of nurse shark (Ginglymostoma cirratum) T cell receptors demonstrating substantial integration of loci and diversification mechanisms in primordial B and T cell repertoires. We survey these findings in this shark model where they were first described, while noting corroborating examples in other vertebrate groups. We also consider other examples where the gnathostome common ancestry of the B and T cell receptor systems have allowed dovetailing of genomic elements and AID-based diversification approaches for the TCR. The cartilaginous fish seem to have retained this T/B cell plasticity to a greater extent than more derived vertebrate groups, but representatives in all vertebrate taxa except bony fish and placental mammals show such plasticity.
Asunto(s)
Inmunoglobulinas/genética , Mamíferos/inmunología , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos/genética , Tiburones/inmunología , Inmunidad Adaptativa , Animales , Citidina Desaminasa/inmunología , Evolución Molecular , Humanos , Mamíferos/genética , Tiburones/genéticaRESUMEN
B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is the most common childhood malignancy. The two-step BCP-ALL pathogenesis requires in utero-induced chromosomal aberrations and additional mutagenic events for overt leukemia. In mouse models, activation-induced cytidine deaminase (AID/AICDA) was suggested to contribute to BCP-ALL pathogenesis by off-target mutagenic activity. The role of AID in patients, however, remains unclear. Moreover, AID is usually not expressed in precursor B-cells but in germinal center B-cells, where it is induced upon T-helper (Th) cell stimulation. We have previously demonstrated that autologous Th-cells supportively interacted with BCP-ALL-cells. Here, we hypothesize that this interaction additionally induces AID expression in BCP-ALL-cells, leading to off-target mutagenic activity. We show that co-culture with autologous bone marrow Th-cells induced high AICDA expression in primary BCP-ALL-cells. This induction was mediated by a mechanism similar to the induction in mature B-cells involving IL-13/Stat6, CD40L/NF-κB and TGFß/Smad2/3 signaling. Even though Th-cell-induced AID seemed to be active in vitro in a BCP-ALL reporter cell line, extensive mutational signature analysis revealed no major contribution of AID activity to the mutational landscape in BCP-ALL patients. AID activity was neither detected in mutation clusters nor in known AID targets. Moreover, no recurrently mutated gene showed a relevant enrichment of mutations in the AID motif. Together, the lack of AID-induced mutational consequences argues towards a Th-cell-promoted yet AID-independent BCP-ALL pathogenesis and favors therapeutic research focusing on Th-cell-derived support of BCP-ALL-cells rather than AID-induced effects.
Asunto(s)
Médula Ósea/inmunología , Citidina Desaminasa/inmunología , Linfoma de Células B/inmunología , Mutagénesis/inmunología , Leucemia-Linfoma Linfoblástico de Células Precursoras B/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Adolescente , Adulto , Linfocitos B/inmunología , Línea Celular Tumoral , Células Cultivadas , Niño , Preescolar , Femenino , Humanos , Lactante , Masculino , Mutación/inmunología , Transducción de Señal/inmunología , Adulto JovenRESUMEN
There has been a longstanding question of whether affinity maturation occurs in ectotherms, and if it does, where in tissues this happens. Although cold-blooded vertebrates (ectotherms) lack histologically discernible germinal centers, they have a fully functional Ig gene mutator enzyme (activation-induced cytidine deaminase: AID or Aicda). Protein and Ig cDNA transcript analyses provide evidence that ectotherms can, under certain conditions, demonstrate antibody affinity maturation, and somatic hypermutation of their Ig genes during secondary immune responses. Here, we review the evidence for antibody affinity maturation and somatic hypermutation of Ig V(D)J exons. We argue that past evidence of long-term intact antigen retention, and recent studies of in situ expression of AID transcripts, point to fish melanomacrophage clusters as sites functionally analogous to a germinal center. Recent work in zebrafish provides a way forward to test these predictions through V(D)J repertoire analyses on isolated, intact melanomacrophage clusters. This work has implications not only for vaccine use in aquaculture, but also for antibody affinity maturation processes in all ectothermic vertebrates.