RESUMO
Linker histone H1 has been correlated with transcriptional inhibition, but the mechanistic basis of the inhibition and its reversal during gene activation has remained enigmatic. We report that H1-compacted chromatin, reconstituted in vitro, blocks transcription by abrogating core histone modifications by p300 but not activator and p300 binding. Transcription from H1-bound chromatin is elicited by the H1 chaperone NAP1, which is recruited in a gene-specific manner through direct interactions with activator-bound p300 that facilitate core histone acetylation (by p300) and concomitant eviction of H1 and H2A-H2B. An analysis in B cells confirms the strong dependency on NAP1-mediated H1 eviction for induction of the silent CD40 gene and further demonstrates that H1 eviction, seeded by activator-p300-NAP1-H1 interactions, is propagated over a CCCTC-binding factor (CTCF)-demarcated region through a distinct mechanism that also involves NAP1. Our results confirm direct transcriptional inhibition by H1 and establish a gene-specific H1 eviction mechanism through an activatorâp300âNAP1âH1 pathway.
Assuntos
Fator de Ligação a CCCTC/genética , Proteína p300 Associada a E1A/genética , Proteínas/genética , Transcrição Gênica , Acetilação , Linfócitos B/química , Sítios de Ligação , Fator de Ligação a CCCTC/química , Antígenos CD40/genética , Cromatina/química , Cromatina/genética , Proteína p300 Associada a E1A/química , Código das Histonas , Histonas/química , Histonas/genética , Humanos , Chaperonas Moleculares/química , Chaperonas Moleculares/genética , Nucleossomos/química , Nucleossomos/genética , Regiões Promotoras Genéticas , Ligação Proteica/genética , Proteínas/química , tRNA MetiltransferasesRESUMO
B-cell receptor (BCR) signaling is crucial for chronic lymphocytic leukemia (CLL) biology. IGLV3-21-expressing B cells may acquire a single point mutation (R110) that triggers autonomous BCR signaling, conferring aggressive behavior. Epigenetic studies have defined 3 CLL subtypes based on methylation signatures reminiscent of naïve-like (n-CLL), intermediate (i-CLL), and memory-like (m-CLL) B cells with different biological features. i-CLL carries a borderline IGHV mutational load and significantly higher use of IGHV3-21/IGLV3-21. To determine the clinical and biological features of IGLV3-21R110 CLL and its relationship to these epigenetic subtypes, we characterized the immunoglobulin gene of 584 CLL cases using whole-genome/exome and RNA sequencing. IGLV3-21R110 was detected in 6.5% of cases: 30 (38%) of 79 i-CLLs, 5 (1.7%) of 291 m-CLLs, and 1 (0.5%) of 189 n-CLLs. All stereotype subset 2 cases carried IGLV3-21R110, whereas 62% of IGLV3-21R110 i-CLL cases had nonstereotyped BCR immunoglobulins. IGLV3-21R110 i-CLL had a significantly higher number of SF3B1 and ATM mutations and total number of driver alterations. However, the R110 mutation was the sole alteration in 1 i-CLL and was accompanied only by del(13q) in 3. Although IGHV mutational status varied, IGLV3-21R110 i-CLL transcriptomically resembled n-CLL/unmutated IGHV CLL with a specific signature including WNT5A/B overexpression. In contrast, i-CLL lacking IGLV3-21R110 mirrored m-CLL/mutated IGHV. Patients with IGLV3-21R110 i-CLL had a short time to first treatment and overall survival similar to those of n-CLL/unmutated IGHV patients, whereas patients with non-IGLV3-21R110 i-CLL had a good prognosis similar to that of patients with m-CLL/mutated IGHV. IGLV3-21R110 defines a CLL subgroup with specific biological features and an unfavorable prognosis independent of IGHV mutational status and epigenetic subtype.
Assuntos
Metilação de DNA , Genes de Cadeia Leve de Imunoglobulina/genética , Leucemia Linfocítica Crônica de Células B/genética , Mutação Puntual , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos B/química , Feminino , Humanos , Leucemia Linfocítica Crônica de Células B/classificação , Leucemia Linfocítica Crônica de Células B/patologia , Masculino , Pessoa de Meia-Idade , Mutação , Adulto JovemRESUMO
Burkitt lymphoma (BL) is a highly aggressive, B-cell, non-Hodgkin lymphoma categorized into endemic, sporadic, and immunodeficiency-associated subtypes. BL has distinct pathologic and clinical features, characterized by rapidly progressive tumors with high rates of extranodal involvement. Next-generation-sequencing analyses have further characterized the genomic landscape of BL and our understanding of disease pathogenesis, although these findings have yet to influence treatment. Although most patients are cured with intensive combination chemotherapy, given the paucity of randomized trials, optimal therapy has not been defined. Furthermore, treatment of elderly patients, patients with central nervous system involvement, or those with relapsed disease remains an unmet need. In this review, we highlight the clinical, pathologic, and genomic features, as well as standard and emerging treatment options for adult patients with BL.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linfoma de Burkitt/tratamento farmacológico , Adulto , África Subsaariana/epidemiologia , Aloenxertos , Antígenos CD/análise , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Linfócitos B/química , Linfócitos B/patologia , Biomarcadores Tumorais , Linfoma de Burkitt/epidemiologia , Linfoma de Burkitt/patologia , Linfoma de Burkitt/virologia , Sistema Nervoso Central/patologia , Gerenciamento Clínico , Doenças Endêmicas , Infecções por Vírus Epstein-Barr/epidemiologia , Europa (Continente)/epidemiologia , Perfilação da Expressão Gênica , Genes myc , Transplante de Células-Tronco Hematopoéticas , Herpesvirus Humano 4/isolamento & purificação , Humanos , Mutação , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Prognóstico , Rituximab/administração & dosagem , Terapias em Estudo , Síndrome de Lise Tumoral/etiologia , Estados Unidos/epidemiologiaRESUMO
The immunoglobulin heavy-chain (Igh) locus is organized into distinct regions that contain multiple variable (V(H)), diversity (D(H)), joining (J(H)) and constant (C(H)) coding elements. How the Igh locus is structured in 3D space is unknown. To probe the topography of the Igh locus, spatial distance distributions were determined between 12 genomic markers that span the entire Igh locus. Comparison of the distance distributions to computer simulations of alternative chromatin arrangements predicted that the Igh locus is organized into compartments containing clusters of loops separated by linkers. Trilateration and triple-point angle measurements indicated the mean relative 3D positions of the V(H), D(H), J(H), and C(H) elements, showed compartmentalization and striking conformational changes involving V(H) and D(H)-J(H) elements during early B cell development. In pro-B cells, the entire repertoire of V(H) regions (2 Mbp) appeared to have merged and juxtaposed to the D(H) elements, mechanistically permitting long-range genomic interactions to occur with relatively high frequency.
Assuntos
Genes de Cadeia Pesada de Imunoglobulina , Animais , Linfócitos B/química , Linfócitos B/metabolismo , Linhagem da Célula , Células Cultivadas , Camundongos , Camundongos Endogâmicos C57BL , Modelos Moleculares , Conformação de Ácido Nucleico , Éxons VDJRESUMO
Despite its popularity, chromatin immunoprecipitation followed by sequencing (ChIP-seq) remains a tedious (>2 d), manually intensive, low-sensitivity and low-throughput approach. Here, we combine principles of microengineering, surface chemistry, and molecular biology to address the major limitations of standard ChIP-seq. The resulting technology, FloChIP, automates and miniaturizes ChIP in a beadless fashion while facilitating the downstream library preparation process through on-chip chromatin tagmentation. FloChIP is fast (<2 h), has a wide dynamic range (from 106 to 500 cells), is scalable and parallelized, and supports antibody- or sample-multiplexed ChIP on both histone marks and transcription factors. In addition, FloChIP's interconnected design allows for straightforward chromatin reimmunoprecipitation, which allows this technology to also act as a microfluidic sequential ChIP-seq system. Finally, we ran FloChIP for the transcription factor MEF2A in 32 distinct human lymphoblastoid cell lines, providing insights into the main factors driving collaborative DNA binding of MEF2A and into its role in B cell-specific gene regulation. Together, our results validate FloChIP as a flexible and reproducible automated solution for individual or sequential ChIP-seq.
Assuntos
Automação/métodos , Sequenciamento de Cromatina por Imunoprecipitação/métodos , Histonas/metabolismo , Fatores de Transcrição MEF2/metabolismo , Automação/instrumentação , Linfócitos B/química , Linfócitos B/metabolismo , Linhagem Celular Tumoral , Sequenciamento de Cromatina por Imunoprecipitação/instrumentação , Histonas/química , Histonas/genética , Humanos , Fatores de Transcrição MEF2/química , Fatores de Transcrição MEF2/genética , Ligação ProteicaRESUMO
The naïve antibody/B-cell receptor (BCR) repertoires of different individuals ought to exhibit significant functional commonality, given that most pathogens trigger an effective antibody response to immunodominant epitopes. Sequence-based repertoire analysis has so far offered little evidence for this phenomenon. For example, a recent study estimated the number of shared ('public') antibody clonotypes in circulating baseline repertoires to be around 0.02% across ten unrelated individuals. However, to engage the same epitope, antibodies only require a similar binding site structure and the presence of key paratope interactions, which can occur even when their sequences are dissimilar. Here, we search for evidence of geometric similarity/convergence across human antibody repertoires. We first structurally profile naïve ('baseline') antibody diversity using snapshots from 41 unrelated individuals, predicting all modellable distinct structures within each repertoire. This analysis uncovers a high (much greater than random) degree of structural commonality. For instance, around 3% of distinct structures are common to the ten most diverse individual samples ('Public Baseline' structures). Our approach is the first computational method to find levels of BCR commonality commensurate with epitope immunodominance and could therefore be harnessed to find more genetically distant antibodies with same-epitope complementarity. We then apply the same structural profiling approach to repertoire snapshots from three individuals before and after flu vaccination, detecting a convergent structural drift indicative of recognising similar epitopes ('Public Response' structures). We show that Antibody Model Libraries derived from Public Baseline and Public Response structures represent a powerful geometric basis set of low-immunogenicity candidates exploitable for general or target-focused therapeutic antibody screening.
Assuntos
Anticorpos , Diversidade de Anticorpos , Linfócitos B , Bases de Dados Genéticas , Epitopos Imunodominantes , Linfócitos B/química , Linfócitos B/imunologia , Linfócitos B/metabolismo , Biologia Computacional , HumanosRESUMO
The E-protein transcription factors guide immune cell differentiation, with E12 and E47 (hereafter called E2A) being essential for B-cell specification and maturation. E2A and the oncogenic chimera E2A-PBX1 contain three transactivation domains (ADs), with AD1 and AD2 having redundant, independent, and cooperative functions in a cell-dependent manner. AD1 and AD2 both mediate their functions by binding to the KIX domain of the histone acetyltransferase paralogues CREB-binding protein (CBP) and E1A-binding protein P300 (p300). This interaction is necessary for B-cell maturation and oncogenesis by E2A-PBX1 and occurs through conserved ΦXXΦΦ motifs (with Φ denoting a hydrophobic amino acid) in AD1 and AD2. However, disruption of this interaction via mutation of the KIX domain in CBP/p300 does not completely abrogate binding of E2A and E2A-PBX1. Here, we determined that E2A-AD1 and E2A-AD2 also interact with the TAZ2 domain of CBP/p300. Characterization of the TAZ2:E2A-AD1(1-37) complex indicated that E2A-AD1 adopts an α-helical structure and uses its ΦXXΦΦ motif to bind TAZ2. Whereas this region overlapped with the KIX recognition region, key KIX-interacting E2A-AD1 residues were exposed, suggesting that E2A-AD1 could simultaneously bind both the KIX and TAZ2 domains. However, we did not detect a ternary complex involving E2A-AD1, KIX, and TAZ2 and found that E2A containing both intact AD1 and AD2 is required to bind to CBP/p300. Our findings highlight the structural plasticity and promiscuity of E2A-AD1 and suggest that E2A binds both the TAZ2 and KIX domains of CBP/p300 through AD1 and AD2.
Assuntos
Proteína de Ligação a CREB/química , Proteína p300 Associada a E1A/genética , Domínios Proteicos/genética , Fator 3 de Transcrição/química , Linfócitos B/química , Linfócitos B/metabolismo , Proteína de Ligação a CREB/genética , Proteína de Ligação a CREB/ultraestrutura , Proteína p300 Associada a E1A/química , Proteína p300 Associada a E1A/ultraestrutura , Proteínas de Homeodomínio/química , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/ultraestrutura , Humanos , Mutação/genética , Proteínas de Fusão Oncogênica/química , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/ultraestrutura , Ligação Proteica/genética , Conformação Proteica , Fator 3 de Transcrição/genética , Fator 3 de Transcrição/ultraestruturaRESUMO
The identification of each cell type is essential for understanding multicellular communities. Antibodies set as biomarkers have been the main toolbox for cell-type recognition, and chemical probes are emerging surrogates. Herein we report the first small-molecule probe, CDgB, to discriminate B lymphocytes from T lymphocytes, which was previously impossible without the help of antibodies. Through the study of the origin of cell specificity, we discovered an unexpected novel mechanism of membrane-oriented live-cell distinction. B cells maintain higher flexibility in their cell membrane than T cells and accumulate the lipid-like probe CDgB more preferably. Because B and T cells share common ancestors, we tracked the cell membrane changes of the progenitor cells and disclosed the dynamic reorganization of the membrane properties over the lymphocyte differentiation progress. This study casts an orthogonal strategy for the small-molecule cell identifier and enriches the toolbox for live-cell distinction from complex cell communities.
Assuntos
Linfócitos B/citologia , Membrana Celular/metabolismo , Corantes Fluorescentes/química , Linfócitos T/citologia , Animais , Linfócitos B/química , Linfócitos B/imunologia , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Diferenciação Celular , Membrana Celular/química , Citometria de Fluxo , Lipidômica , Camundongos , Linfócitos T/química , Linfócitos T/imunologiaRESUMO
To acquire a better understanding of clonal evolution of acute myeloid leukemia (AML) and to identify the clone(s) responsible for disease recurrence, we have comparatively studied leukemia-specific mutations by whole-exome-sequencing (WES) of both the leukemia and the nonleukemia compartments derived from the bone marrow of AML patients. The T-lymphocytes, B-lymphocytes and the functionally normal hematopoietic stem cells (HSC), that is, CD34+ /CD38- /ALDH+ cells for AML with rare-ALDH+ blasts (<1.9% ALDH+ cells) were defined as the nonleukemia compartments. WES identified 62 point-mutations in the leukemia compartment derived from 12 AML-patients at the time of diagnosis and 73 mutations in 3 matched relapse cases. Most patients (8/12) showed 4 to 6 point-mutations per sample at diagnosis. Other than the mutations in the recurrently mutated genes such as DNMT3A, NRAS and KIT, we were able to identify novel point-mutations that have not yet been described in AML. Some leukemia-specific mutations and cytogenetic abnormalities including DNMT3A(R882H), EZH2(I146T) and inversion(16) were also detectable in the respective T-lymphocytes, B-lymphocytes and HSC in 5/12 patients, suggesting that preleukemia HSC might represent the source of leukemogenesis for these cases. The leukemic evolution was reconstructed for five cases with detectable preleukemia clones, which were tracked in follow-up and relapse samples. Four of the five patients with detectable preleukemic mutations developed relapse. The presence of leukemia-specific mutations in these nonleukemia compartments, especially after chemotherapy or after allogeneic stem cell transplantation, is highly relevant, as these could be responsible for relapse. This discovery may facilitate the identification of novel targets for long-term cure.
Assuntos
Biomarcadores Tumorais/genética , Resistencia a Medicamentos Antineoplásicos , Sequenciamento do Exoma/métodos , Leucemia Mieloide Aguda/genética , Mutação Puntual , Lesões Pré-Cancerosas/genética , Idoso , Linfócitos B/química , Evolução Clonal , DNA (Citosina-5-)-Metiltransferases/genética , DNA Metiltransferase 3A , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Feminino , GTP Fosfo-Hidrolases/genética , Células-Tronco Hematopoéticas/química , Humanos , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-kit/genética , Linfócitos T/químicaRESUMO
We assessed the concordance between immunohistochemistry (IHC) and gene expression profiling (GEP) for determining diffuse large B-cell lymphoma (DLBCL) cell of origin (COO) in the phase III PHOENIX trial of rituximab plus cyclophosphamide, doxorubicin, vincristine and prednisone (R-CHOP) with or without ibrutinib. Among 910 of 1114 screened patients with non-germinal centre B cell-like (non-GCB) DLBCL by IHC, the concordance with GEP for non-GCB calls was 82·7%, with 691 (75·9%) identified as activated B cell-like (ABC), and 62 (6·8%) as unclassified. Among 746 of 837 enrolled patients with verified non-GCB DLBCL by IHC, the concordance with GEP was 82·8%, with 567 (76·0%) identified as ABC and 51 (6·8%) unclassified; survival outcomes were similar regardless of COO or treatment, whereas among patients with ABC DLBCL aged <60 years, the overall and event-free survival were substantially better with ibrutinib versus placebo plus R-CHOP [hazard ratio (HR) 0·365, 95% confidence interval (CI) 0·147-0·909, P = 0·0305; HR 0·561, 95% CI 0·326-0·967, P = 0·0348, respectively]. IHC and GEP showed high concordance and consistent survival outcomes among tested patients, indicating centralised IHC may be used to enrich populations for response to ibrutinib plus R-CHOP.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Perfilação da Expressão Gênica , Imuno-Histoquímica , Linfoma Difuso de Grandes Células B/classificação , Adenina/administração & dosagem , Adenina/análogos & derivados , Adulto , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Linfócitos B/química , Linfócitos B/patologia , Ciclofosfamida/administração & dosagem , Doxorrubicina/administração & dosagem , Centro Germinativo/patologia , Humanos , Estimativa de Kaplan-Meier , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/patologia , Pessoa de Meia-Idade , Células-Tronco Neoplásicas/química , Células-Tronco Neoplásicas/patologia , Piperidinas/administração & dosagem , Prednisona/administração & dosagem , Prognóstico , Intervalo Livre de Progressão , Rituximab/administração & dosagem , Resultado do Tratamento , Vincristina/administração & dosagemRESUMO
Carbohydrate-protein interactions define a multitude of cellular recognition events. We present herein synthetic glycovesicles as cell-surface mimics in order to switch the nature of lectin recognition. The covalent glycovesicles, constituted with diacetylene monomers of various ligand densities at their surfaces, are prepared through photo-polymerization. Vesicles with sparsely imbedded ligands engage in a lectin interaction leading to the formation of a dense, crosslinked multimeric complex. On the other hand, vesicles with many ligands, or completely covered with them, switch the lectin interaction to form a fully soluble monomeric complex, without crosslinking. Nanomolar dissociation constants govern these interactions, as assessed by a ligand-displacement assay. The study demonstrates the switching nature - between monomeric and multimeric - of the interaction as a function of ligand density in the vesicles; the results are directly relevant to understanding such a phenomenon occurring at cell surfaces.
Assuntos
Glicosídeos/química , Lectinas/análise , Linfócitos B/química , Glicosídeos/síntese química , Humanos , Ligantes , Estrutura Molecular , Propriedades de SuperfícieRESUMO
BACKGROUND: Flow cytometry plays a key role in detecting bone marrow (BM) involvement in patients with diffuse large B-cell lymphoma (DLBCL). To improve its detection sensitivity, we need to explore novel markers. In this study, we detected the expression CD54 on lymphoma cells in BM specimens from DLBCL patients and clarified its diagnostic significance in BM involvement by DLBCL. METHODS: We collected BM specimens from 76 patients with DLBCL (germinal center B-cell (GCB) = 25, non-GCB = 51) and 10 control patients without lymphoma. We detected and compared the expression of CD54 on lymphoma cells and normal mature B cells by using 10-color panels. RESULTS: Normal plasma cells expressed a higher level of CD54 as compared with hematogones (p < 0.05) and normal mature B cells (p < 0.05). Among 76 patients, 23 of them (GCB = 12, non-GCB = 11) had BM involvement. Lymphoma B cells from 12 cases (GBC = 4, non-GCB = 8) expressed a higher level of CD54 compared to normal mature B cells (p < 0.05). Additionally, lymphoma cells of the non-GCB subtype frequently expressed a higher level of CD54 in comparison to the GCB subtype (p < 0.05). And the high expression of CD54 was not related to plasmacytoid differentiation. CONCLUSION: Aberrant expression of CD54 on lymphoma cells is frequently seen in patients' BM specimens involved by DLBCL, especially in the non-GCB subtype. CD54 could be used as a new marker to gate on lymphoma cells and improve the detection sensitivity of BM involvement in patients with DLBCL.
Assuntos
Biomarcadores Tumorais/análise , Medula Óssea/química , Citometria de Fluxo , Molécula 1 de Adesão Intercelular/análise , Linfoma Difuso de Grandes Células B/química , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos B/química , Linfócitos B/metabolismo , Biomarcadores Tumorais/metabolismo , Biópsia , Medula Óssea/metabolismo , Medula Óssea/patologia , Moléculas de Adesão Celular/análise , Moléculas de Adesão Celular/metabolismo , Diferenciação Celular , Feminino , Centro Germinativo/química , Centro Germinativo/metabolismo , Centro Germinativo/patologia , Humanos , Imunofenotipagem , Molécula 1 de Adesão Intercelular/metabolismo , Linfoma Difuso de Grandes Células B/metabolismo , Linfoma Difuso de Grandes Células B/patologia , Masculino , Pessoa de Meia-Idade , Plasmócitos/citologiaRESUMO
Treatment options for relapsed or refractory B-lymphoblastic leukaemia (r/r B-ALL) are limited and the prognosis of these patients remains dismal, but novel immunotherapeutic options such as the anti-CD22 antibody-drug-conjugate Inotuzumab-Ozogamicin (InO) have improved outcomes in these patients. Flow cytometry is essential to assess antigen-expression prior to treatment initiation of antigen-directed immunotherapies. Here, we present flow cytometric and clinical data of three adult patients with r/r B-ALL who failed treatment with InO associated with reduced or lost antigen-expression. In addition, we present comparative data on two different diagnostic CD22-specific antibody clones that exhibit significant differences in staining intensities.
Assuntos
Antineoplásicos Imunológicos/uso terapêutico , Linfócitos B/química , Inotuzumab Ozogamicina/uso terapêutico , Subpopulações de Linfócitos/química , Leucemia-Linfoma Linfoblástico de Células Precursoras B/tratamento farmacológico , Lectina 2 Semelhante a Ig de Ligação ao Ácido Siálico/análise , Adulto , Idoso de 80 Anos ou mais , Aloenxertos , Anticorpos Biespecíficos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linfócitos B/patologia , Células Clonais , Feminino , Citometria de Fluxo , Transplante de Células-Tronco Hematopoéticas , Humanos , Mesilato de Imatinib/administração & dosagem , Imunofenotipagem , Subpopulações de Linfócitos/patologia , Masculino , Pessoa de Meia-Idade , Leucemia-Linfoma Linfoblástico de Células Precursoras B/sangue , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologia , Recidiva , Terapia de Salvação , Sorafenibe/uso terapêutico , Falha de Tratamento , Adulto JovemRESUMO
B-cell epitope information is critical to immune therapy and vaccine design. Protein epitopes can be significantly affected by glycosylation, while no methods have considered this till now. Based on previous versions of Spatial Epitope Prediction of Protein Antigens (SEPPA), we here present an enhanced tool SEPPA 3.0, enabling glycoprotein antigens. Parameters were updated based on the latest and largest dataset. Then, additional micro-environmental features of glycosylation triangles and glycosylation-related amino acid indexes were added as important classifiers, coupled with final calibration based on neighboring antigenicity. Logistic regression model was retained as SEPPA 2.0. The AUC value of 0.794 was obtained through 10-fold cross-validation on internal validation. Independent testing on general protein antigens resulted in AUC of 0.740 with BA (balanced accuracy) of 0.657 as baseline of SEPPA 3.0. Most importantly, when tested on independent glycoprotein antigens only, SEPPA 3.0 gave an AUC of 0.749 and BA of 0.665, leading the top performance among peers. As the first server enabling accurate epitope prediction for glycoproteins, SEPPA 3.0 shows significant advantages over popular peers on both general protein and glycoprotein antigens. It can be accessed at http://bidd2.nus.edu.sg/SEPPA3/ or at http://www.badd-cao.net/seppa3/index.html. Batch query is supported.
Assuntos
Antígenos/química , Mapeamento de Epitopos/métodos , Epitopos de Linfócito B/química , Glicoproteínas/química , Proteína gp120 do Envelope de HIV/química , Processamento de Proteína Pós-Traducional , Software , Algoritmos , Antígenos/imunologia , Antígenos/metabolismo , Área Sob a Curva , Linfócitos B/química , Linfócitos B/imunologia , Bases de Dados de Proteínas , Conjuntos de Dados como Assunto , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito B/metabolismo , Glicoproteínas/imunologia , Glicoproteínas/metabolismo , Glicosilação , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp120 do Envelope de HIV/metabolismo , Humanos , Internet , Modelos Logísticos , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre ProteínasRESUMO
BACKGROUND: Lung aging is characterized by a number of structural alterations including fibrosis, chronic inflammation and the alteration of inflammatory cell composition. Chronic exposure to cigarette smoke (CS) is known to induce similar alterations and may contribute to premature lung aging. Additionally, aging and CS exposure are associated with transcriptional alterations in the lung. The current work aims to explore the interaction between age- and CS- associated transcriptomic perturbations and develop a transcriptomic clock able to predict the biological age and the impact of external factors on lung aging. RESULTS: Our investigations revealed a substantial overlap between transcriptomic response to CS exposure and age-related transcriptomic alterations in the murine lung. Of particular interest is the strong upregulation of immunoglobulin genes with increased age and in response to CS exposure, indicating an important implication of B-cells in lung inflammation associated with aging and smoking. Furthermore, we used a machine learning approach based on Lasso regression to build a transcriptomic age model that can accurately predict chronological age in untreated mice and the deviations associated with certain exposures. Interestingly, CS-exposed-mice were predicted to be prematurely aged in contrast to mice exposed to fresh air or to heated tobacco products (HTPs). The accelerated aging rate associated with CS was reversed upon smoking cessation or switching to HTPs. Additionally, our model was able to predict premature aging associated with thoracic irradiation from an independent public dataset. CONCLUSIONS: Aging and CS exposure share common transcriptional alteration patterns in the murine lung. The massive upregulation of B-cell restricted genes during these processes shed light on the contribution of cell composition and particularly immune cells to the measured transcriptomic signal. Through machine learning approach, we show that gene expression changes can be used to accurately monitor the biological age and the modulations associated with certain exposures. Our findings also suggest that the premature lung aging is reversible upon the reduction of harmful exposures.
Assuntos
Senilidade Prematura/genética , Perfilação da Expressão Gênica/métodos , Pulmão/química , Fumaça/efeitos adversos , Senilidade Prematura/induzido quimicamente , Animais , Linfócitos B/química , Ritmo Circadiano , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Aprendizado de Máquina , Camundongos , Camundongos Endogâmicos C57BL , Análise de Sequência com Séries de Oligonucleotídeos , Produtos do TabacoRESUMO
CD22 (Siglec-2) is a B-cell surface inhibitory protein capable of selectively recognising sialylated glycans, thus dampening autoimmune responses against self-antigens. Here we have characterised the dynamic recognition of complex-type N-glycans by human CD22 by means of orthogonal approaches including NMR spectroscopy, computational methods and biophysical assays. We provide new molecular insights into the binding mode of sialoglycans in complex with h-CD22, highlighting the role of the sialic acid galactose moieties in the recognition process, elucidating the conformational behaviour of complex-type N-glycans bound to Siglec-2 and dissecting the formation of CD22 homo-oligomers on the B-cell surface. Our results could enable the development of additional therapeutics capable of modulating the activity of h-CD22 in autoimmune diseases and malignancies derived from B-cells.
Assuntos
Simulação de Dinâmica Molecular , Polissacarídeos/química , Lectina 2 Semelhante a Ig de Ligação ao Ácido Siálico/química , Linfócitos B/química , Configuração de Carboidratos , Galactose/química , HumanosRESUMO
The Myc family of oncogenic transcription factors regulates myriad cellular functions. Myc proteins contain a basic region/helix-loop-helix/leucine zipper domain that mediates DNA binding and heterodimerization with its partner Max. Among the Myc proteins, c-Myc is the most widely expressed and relevant in primary B lymphocytes. There is evidence suggesting that c-Myc can perform some of its functions in the absence of Max in different cellular contexts. However, the functional in vivo interplay between c-Myc and Max during B lymphocyte differentiation is not well understood. Using in vivo and ex vivo models, we show that while c-Myc requires Max in primary B lymphocytes, several key biological processes, such as cell differentiation and DNA replication, can initially progress without the formation of c-Myc/Max heterodimers. We also describe that B lymphocytes lacking Myc, Max, or both show upregulation of signaling pathways associated with the B-cell receptor. These data suggest that c-Myc/Max heterodimers are not essential for the initiation of a subset of important biological processes in B lymphocytes, but are required for fine-tuning the initial response after activation.
Assuntos
Linfócitos B/química , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Diferenciação Celular/genética , Proteínas Proto-Oncogênicas c-myc/genética , Sequência de Aminoácidos/genética , Animais , Linfócitos B/metabolismo , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/química , Replicação do DNA/genética , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Dimerização , Sequências Hélice-Alça-Hélice/genética , Humanos , Zíper de Leucina/genética , Camundongos , Ligação Proteica/genética , Proteínas Proto-Oncogênicas c-myc/química , Ativação Transcricional/genéticaRESUMO
BACKGROUND: To investigate the roles of T, B, and natural killer (NK) cells in pregnancy outcome of women with recurrent implantation failure (RIF). METHODS: This retrospective cohort study enrolled 196 patients with RIF. Peripheral lymphocyte subsets were measured before and during pregnancy. The relationship between pregnancy outcome and level of lymphocytes was analyzed. RESULTS: Peripheral CD19+ B cells in women who experienced miscarriage were significantly lower than those who subsequently had live birth. After adjusting for potential confounders in the multiple logistic regression models, each 1% increment in the peripheral CD19+ B cells before pregnancy [odds ratio (OR): 0.93] and during early pregnancy (OR: 0.83) was associated with a significantly decreased risk of miscarriage (p < 0.05). The risk of mis-carriage in patients with ≥ 15% CD19+ B cells before and during pregnancy was 39% and 21% lower, respectively, than in their counterparts with < 15% CD19+ B cells. The association between CD19+ B cells and the risk of miscarriage was nonlinear. CONCLUSIONS: Measurement of peripheral CD19+ subsets may help predict the pregnancy outcome in women with RIF.
Assuntos
Aborto Espontâneo/epidemiologia , Antígenos CD19/sangue , Linfócitos B/química , Perda do Embrião/epidemiologia , Adulto , Linfócitos B/citologia , Feminino , Humanos , Subpopulações de Linfócitos/química , Gravidez , Estudos Retrospectivos , Fatores de RiscoRESUMO
PURPOSE: Immunohistochemistry of orbital tissues offers a correlation between the microscopic changes and macroscopic clinical manifestation of Graves' orbitopathy (GO). Summarizing the participation of different molecules will help us to understand the pathogenesis of GO. METHODS: The pertinent and current literature on immunohistochemistry of human orbital tissue in GO was reviewed using the NCBI PubMed database. RESULTS: 33 articles comprising over 700 orbital tissue samples were included in this review. The earliest findings included the demonstration of HLA-DR and T cell (to a lesser extent B cell) markers in GO orbital tissues. Subsequent investigators further contributed by characterizing cellular infiltration, confirming the presence of HLA-DR and TSHR, as well as revealing the participation of cytokines, growth factors, adhesion molecules and miscellaneous substances. HLA-DR and TSHR are over-expressed in orbital tissues of GO patients. The inflammatory infiltration mainly comprises CD4 + T cells and macrophages. Cytokine profile suggests the importance of Th1 (especially in early active phase) and Th17 immunity in the pathogenesis of GO. Upregulation of proinflammatory/profibrotic cytokines, adhesion molecules and growth factors finally culminate in activation of orbital fibroblasts and perpetuation of orbital inflammation. The molecular status of selected parameters correlates with the clinical presentation of GO. CONCLUSION: Further investigation is warranted to define precisely the role of different molecules and ongoing search for new players yet to be discovered is also important. Unfolding the molecular mechanisms behind GO will hopefully provide insights into the development of novel therapeutic strategies and optimize our clinical management of the disease.
Assuntos
Oftalmopatia de Graves/metabolismo , Mediadores da Inflamação/metabolismo , Órbita/química , Órbita/metabolismo , Animais , Linfócitos B/química , Linfócitos B/metabolismo , Citocinas/análise , Citocinas/metabolismo , Oftalmopatia de Graves/patologia , Humanos , Mediadores da Inflamação/análise , Órbita/patologia , Linfócitos T/química , Linfócitos T/metabolismoRESUMO
Common differentially expressed genes (DEGs) in acute lymphoblastic leukemia (ALL) and chronic lymphocytic leukemia (CLL) might play critical roles in the pathogenesis and process of leukemia. We collected RNA sequencing (RNA-seq) data of human CLL, ALL samples, and normal peripheral blood CD19+ B cells as well as thymus samples, and analyzed similarities and differences between their transcriptomes using Cuffdiff2, DESeq, and edgeR. Compared with the RNA-seq data of normal peripheral blood CD19+ B cells and thymus samples, there were a large number of DEGs in ALL and CLL. DEGs in ALL and CLL not only have their distinguished features but also have a similar pattern. To figure out the common DEGs between CLL and ALL, we further identified 26 overlapped genes between CLL and ALL, among which 10 genes showed similar expression variation profiles whereas 16 genes showed opposite variation. The expression levels of 10 genes (SCML4, TNF-α, CD1C, FGFR1, MYO7B, DUSP1, PAP1GAP, MAN1C1, SLFN5, and CD8A) among the 26 genes were further confirmed by experiments, which was consistent with the results obtained by analyzing the RNA-seq data. The current study contributes to better understanding the pathophysiology of leukemia and unearthing novel potential prognostic markers and therapeutic targets of leukemia.