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1.
Annu Rev Immunol ; 40: 323-348, 2022 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-35113729

RESUMO

The diverse biological activity of interleukin-6 (IL-6) contributes to the maintenance of homeostasis. Emergent infection or tissue injury induces rapid production of IL-6 and activates host defense through augmentation of acute-phase proteins and immune responses. However, excessive IL-6 production and uncontrolled IL-6 receptor signaling are critical to pathogenesis. Over the years, therapeutic agents targeting IL-6 signaling, such as tocilizumab, a humanized anti-IL-6 receptor antibody, have shown remarkable efficacy for rheumatoid arthritis, Castleman disease, and juvenile idiopathic arthritis, and their efficacy in other diseases is continually being reported. Emerging evidence has demonstrated the benefit of tocilizumab for several types of acute inflammatory diseases, including cytokine storms induced by chimeric antigen receptor T cell therapy and coronavirus disease 2019 (COVID-19). Here, we refocus attention on the biology of IL-6 and summarize the distinct pathological roles of IL-6 signaling in several acute and chronic inflammatory diseases.


Assuntos
Artrite Reumatoide , COVID-19 , Animais , Artrite Reumatoide/terapia , COVID-19/terapia , Humanos , Imunoterapia Adotiva , Interleucina-6/metabolismo , Transdução de Sinais
2.
Annu Rev Immunol ; 37: 349-375, 2019 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-30673536

RESUMO

Detection of double-stranded RNAs (dsRNAs) is a central mechanism of innate immune defense in many organisms. We here discuss several families of dsRNA-binding proteins involved in mammalian antiviral innate immunity. These include RIG-I-like receptors, protein kinase R, oligoadenylate synthases, adenosine deaminases acting on RNA, RNA interference systems, and other proteins containing dsRNA-binding domains and helicase domains. Studies suggest that their functions are highly interdependent and that their interdependence could offer keys to understanding the complex regulatory mechanisms for cellular dsRNA homeostasis and antiviral immunity. This review aims to highlight their interconnectivity, as well as their commonalities and differences in their dsRNA recognition mechanisms.


Assuntos
Imunidade Inata/genética , RNA de Cadeia Dupla/genética , Viroses/imunologia , 2',5'-Oligoadenilato Sintetase/metabolismo , Animais , Proteína DEAD-box 58/metabolismo , Humanos , Imunomodulação , Mamíferos , Nucleotídeo Desaminases/metabolismo , Interferência de RNA , eIF-2 Quinase/metabolismo
3.
Annu Rev Biochem ; 93(1): 21-46, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38594943

RESUMO

DNA replication and transcription occur in all living cells across all domains of life. Both essential processes occur simultaneously on the same template, leading to conflicts between the macromolecular machines that perform these functions. Numerous studies over the past few decades demonstrate that this is an inevitable problem in both prokaryotic and eukaryotic cells. We have learned that conflicts lead to replication fork reversal, breaks in the DNA, R-loop formation, topological stress, and mutagenesis and can ultimately impact evolution. Recent studies have also provided insight into the various mechanisms that mitigate, resolve, and allow tolerance of conflicts and how conflicts result in pathological consequences across divergent species. In this review, we summarize our current knowledge regarding the outcomes of the encounters between replication and transcription machineries and explore how these clashes are dealt with across species.


Assuntos
Replicação do DNA , Transcrição Gênica , Humanos , Animais , Cromossomos/metabolismo , Cromossomos/genética , Cromossomos/química , Estruturas R-Loop , DNA/metabolismo , DNA/genética , DNA/química
4.
Cell ; 187(19): 5238-5252.e20, 2024 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-39208796

RESUMO

Fanzor (Fz) is an ωRNA-guided endonuclease extensively found throughout the eukaryotic domain with unique gene editing potential. Here, we describe the structures of Fzs from three different organisms. We find that Fzs share a common ωRNA interaction interface, regardless of the length of the ωRNA, which varies considerably across species. The analysis also reveals Fz's mode of DNA recognition and unwinding capabilities as well as the presence of a non-canonical catalytic site. The structures demonstrate how protein conformations of Fz shift to allow the binding of double-stranded DNA to the active site within the R-loop. Mechanistically, examination of structures in different states shows that the conformation of the lid loop on the RuvC domain is controlled by the formation of the guide/DNA heteroduplex, regulating the activation of nuclease and DNA double-stranded displacement at the single cleavage site. Our findings clarify the mechanism of Fz, establishing a foundation for engineering efforts.


Assuntos
Clivagem do DNA , DNA , DNA/metabolismo , DNA/química , Domínio Catalítico , Modelos Moleculares , RNA Guia de Sistemas CRISPR-Cas/metabolismo , RNA Guia de Sistemas CRISPR-Cas/química , Humanos , Endodesoxirribonucleases/metabolismo , Endodesoxirribonucleases/química , Edição de Genes , Sistemas CRISPR-Cas
5.
Cell ; 187(13): 3249-3261.e14, 2024 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-38781968

RESUMO

Thermostable clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas9) enzymes could improve genome-editing efficiency and delivery due to extended protein lifetimes. However, initial experimentation demonstrated Geobacillus stearothermophilus Cas9 (GeoCas9) to be virtually inactive when used in cultured human cells. Laboratory-evolved variants of GeoCas9 overcome this natural limitation by acquiring mutations in the wedge (WED) domain that produce >100-fold-higher genome-editing levels. Cryoelectron microscopy (cryo-EM) structures of the wild-type and improved GeoCas9 (iGeoCas9) enzymes reveal extended contacts between the WED domain of iGeoCas9 and DNA substrates. Biochemical analysis shows that iGeoCas9 accelerates DNA unwinding to capture substrates under the magnesium-restricted conditions typical of mammalian but not bacterial cells. These findings enabled rational engineering of other Cas9 orthologs to enhance genome-editing levels, pointing to a general strategy for editing enzyme improvement. Together, these results uncover a new role for the Cas9 WED domain in DNA unwinding and demonstrate how accelerated target unwinding dramatically improves Cas9-induced genome-editing activity.


Assuntos
Proteína 9 Associada à CRISPR , Sistemas CRISPR-Cas , Microscopia Crioeletrônica , DNA , Edição de Genes , Humanos , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/química , Proteína 9 Associada à CRISPR/metabolismo , Proteína 9 Associada à CRISPR/genética , Sistemas CRISPR-Cas/genética , DNA/metabolismo , DNA/genética , Edição de Genes/métodos , Geobacillus stearothermophilus/genética , Geobacillus stearothermophilus/metabolismo , Células HEK293 , Domínios Proteicos , Genoma Humano , Modelos Moleculares , Estrutura Terciária de Proteína , Conformação de Ácido Nucleico , Biocatálise , Magnésio/química , Magnésio/metabolismo
6.
Cell ; 187(17): 4674-4689.e18, 2024 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-38981481

RESUMO

All-RNA-mediated targeted gene integration methods, rendering reduced immunogenicity, effective deliverability with non-viral vehicles, and a low risk of random mutagenesis, are urgently needed for next-generation gene addition technologies. Naturally occurring R2 retrotransposons hold promise in this context due to their site-specific integration profile. Here, we systematically analyzed the biodiversity of R2 elements and screened several R2 orthologs capable of full-length gene insertion in mammalian cells. Robust R2 system gene integration efficiency was attained using combined donor RNA and protein engineering. Importantly, the all-RNA-delivered engineered R2 system showed effective integration activity, with efficiency over 60% in mouse embryos. Unbiased high-throughput sequencing demonstrated that the engineered R2 system exhibited high on-target integration specificity (99%). In conclusion, our study provides engineered R2 tools for applications based on hit-and-run targeted DNA integration and insights for further optimization of retrotransposon systems.


Assuntos
RNA , Retroelementos , Animais , Retroelementos/genética , Camundongos , Humanos , RNA/genética , RNA/metabolismo , Células HEK293 , Engenharia Genética/métodos , Marcação de Genes/métodos
7.
Cell ; 187(16): 4305-4317.e18, 2024 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-38936360

RESUMO

Interleukin (IL)-23 and IL-17 are well-validated therapeutic targets in autoinflammatory diseases. Antibodies targeting IL-23 and IL-17 have shown clinical efficacy but are limited by high costs, safety risks, lack of sustained efficacy, and poor patient convenience as they require parenteral administration. Here, we present designed miniproteins inhibiting IL-23R and IL-17 with antibody-like, low picomolar affinities at a fraction of the molecular size. The minibinders potently block cell signaling in vitro and are extremely stable, enabling oral administration and low-cost manufacturing. The orally administered IL-23R minibinder shows efficacy better than a clinical anti-IL-23 antibody in mouse colitis and has a favorable pharmacokinetics (PK) and biodistribution profile in rats. This work demonstrates that orally administered de novo-designed minibinders can reach a therapeutic target past the gut epithelial barrier. With high potency, gut stability, and straightforward manufacturability, de novo-designed minibinders are a promising modality for oral biologics.


Assuntos
Colite , Interleucina-17 , Células Th17 , Animais , Administração Oral , Camundongos , Humanos , Ratos , Colite/tratamento farmacológico , Interleucina-17/metabolismo , Interleucina-17/antagonistas & inibidores , Células Th17/imunologia , Receptores de Interleucina/metabolismo , Receptores de Interleucina/antagonistas & inibidores , Camundongos Endogâmicos C57BL , Masculino , Interleucina-23/metabolismo , Interleucina-23/antagonistas & inibidores , Distribuição Tecidual , Feminino , Ratos Sprague-Dawley
8.
Cell ; 187(13): 3390-3408.e19, 2024 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-38754421

RESUMO

Clinical trials have identified ARID1A mutations as enriched among patients who respond favorably to immune checkpoint blockade (ICB) in several solid tumor types independent of microsatellite instability. We show that ARID1A loss in murine models is sufficient to induce anti-tumor immune phenotypes observed in ARID1A mutant human cancers, including increased CD8+ T cell infiltration and cytolytic activity. ARID1A-deficient cancers upregulated an interferon (IFN) gene expression signature, the ARID1A-IFN signature, associated with increased R-loops and cytosolic single-stranded DNA (ssDNA). Overexpression of the R-loop resolving enzyme, RNASEH2B, or cytosolic DNase, TREX1, in ARID1A-deficient cells prevented cytosolic ssDNA accumulation and ARID1A-IFN gene upregulation. Further, the ARID1A-IFN signature and anti-tumor immunity were driven by STING-dependent type I IFN signaling, which was required for improved responsiveness of ARID1A mutant tumors to ICB treatment. These findings define a molecular mechanism underlying anti-tumor immunity in ARID1A mutant cancers.


Assuntos
Linfócitos T CD8-Positivos , Proteínas de Ligação a DNA , Interferon Tipo I , Proteínas de Membrana , Neoplasias , Transdução de Sinais , Fatores de Transcrição , Animais , Humanos , Camundongos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/metabolismo , Exodesoxirribonucleases/metabolismo , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Interferon Tipo I/metabolismo , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Camundongos Endogâmicos C57BL , Mutação , Neoplasias/imunologia , Neoplasias/genética , Proteínas Nucleares/metabolismo , Fosfoproteínas/metabolismo , Fatores de Transcrição/metabolismo , Masculino , Quimiocinas/genética , Quimiocinas/metabolismo
9.
Cell ; 186(13): 2865-2879.e20, 2023 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-37301196

RESUMO

Retroelements are the widespread jumping elements considered as major drivers for genome evolution, which can also be repurposed as gene-editing tools. Here, we determine the cryo-EM structures of eukaryotic R2 retrotransposon with ribosomal DNA target and regulatory RNAs. Combined with biochemical and sequencing analysis, we reveal two essential DNA regions, Drr and Dcr, required for recognition and cleavage. The association of 3' regulatory RNA with R2 protein accelerates the first-strand cleavage, blocks the second-strand cleavage, and initiates the reverse transcription starting from the 3'-tail. Removing 3' regulatory RNA by reverse transcription allows the association of 5' regulatory RNA and initiates the second-strand cleavage. Taken together, our work explains the DNA recognition and RNA supervised sequential retrotransposition mechanisms by R2 machinery, providing insights into the retrotransposon and application reprogramming.


Assuntos
RNA , Retroelementos , RNA/metabolismo , Clivagem do DNA , DNA Polimerase Dirigida por RNA/metabolismo , Transcrição Reversa
10.
Cell ; 186(6): 1162-1178.e20, 2023 03 16.
Artigo em Inglês | MEDLINE | ID: mdl-36931244

RESUMO

Germline histone H3.3 amino acid substitutions, including H3.3G34R/V, cause severe neurodevelopmental syndromes. To understand how these mutations impact brain development, we generated H3.3G34R/V/W knock-in mice and identified strikingly distinct developmental defects for each mutation. H3.3G34R-mutants exhibited progressive microcephaly and neurodegeneration, with abnormal accumulation of disease-associated microglia and concurrent neuronal depletion. G34R severely decreased H3K36me2 on the mutant H3.3 tail, impairing recruitment of DNA methyltransferase DNMT3A and its redistribution on chromatin. These changes were concurrent with sustained expression of complement and other innate immune genes possibly through loss of non-CG (CH) methylation and silencing of neuronal gene promoters through aberrant CG methylation. Complement expression in G34R brains may lead to neuroinflammation possibly accounting for progressive neurodegeneration. Our study reveals that H3.3G34-substitutions have differential impact on the epigenome, which underlie the diverse phenotypes observed, and uncovers potential roles for H3K36me2 and DNMT3A-dependent CH-methylation in modulating synaptic pruning and neuroinflammation in post-natal brains.


Assuntos
DNA Metiltransferase 3A , Histonas , Animais , Camundongos , DNA (Citosina-5-)-Metiltransferases/genética , Metilação de DNA/genética , Metilases de Modificação do DNA/genética , Histonas/metabolismo , Doenças Neuroinflamatórias
11.
Cell ; 186(12): 2593-2609.e18, 2023 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-37209683

RESUMO

Here, we describe an approach to correct the genetic defect in fragile X syndrome (FXS) via recruitment of endogenous repair mechanisms. A leading cause of autism spectrum disorders, FXS results from epigenetic silencing of FMR1 due to a congenital trinucleotide (CGG) repeat expansion. By investigating conditions favorable to FMR1 reactivation, we find MEK and BRAF inhibitors that induce a strong repeat contraction and full FMR1 reactivation in cellular models. We trace the mechanism to DNA demethylation and site-specific R-loops, which are necessary and sufficient for repeat contraction. A positive feedback cycle comprising demethylation, de novo FMR1 transcription, and R-loop formation results in the recruitment of endogenous DNA repair mechanisms that then drive excision of the long CGG repeat. Repeat contraction is specific to FMR1 and restores the production of FMRP protein. Our study therefore identifies a potential method of treating FXS in the future.


Assuntos
Síndrome do Cromossomo X Frágil , Expansão das Repetições de Trinucleotídeos , Humanos , Estruturas R-Loop , Metilação de DNA , Síndrome do Cromossomo X Frágil/genética , Epigênese Genética , Proteína do X Frágil da Deficiência Intelectual/genética , Proteína do X Frágil da Deficiência Intelectual/metabolismo
12.
Cell ; 185(17): 3186-3200.e17, 2022 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-35907403

RESUMO

Upon stress, eukaryotes typically reprogram their translatome through GCN2-mediated phosphorylation of the eukaryotic translation initiation factor, eIF2α, to inhibit general translation initiation while selectively translating essential stress regulators. Unexpectedly, in plants, pattern-triggered immunity (PTI) and response to other environmental stresses occur independently of the GCN2/eIF2α pathway. Here, we show that while PTI induces mRNA decapping to inhibit general translation, defense mRNAs with a purine-rich element ("R-motif") are selectively translated using R-motif as an internal ribosome entry site (IRES). R-motif-dependent translation is executed by poly(A)-binding proteins (PABPs) through preferential association with the PTI-activating eIFiso4G over the repressive eIF4G. Phosphorylation by PTI regulators mitogen-activated protein kinase 3 and 6 (MPK3/6) inhibits eIF4G's activity while enhancing PABP binding to the R-motif and promoting eIFiso4G-mediated defense mRNA translation, establishing a link between PTI signaling and protein synthesis. Given its prevalence in both plants and animals, the PABP/R-motif translation initiation module may have a broader role in reprogramming the stress translatome.


Assuntos
Fator de Iniciação Eucariótico 4G , Proteínas de Ligação a Poli(A) , Animais , Fator de Iniciação Eucariótico 4G/genética , Fator de Iniciação Eucariótico 4G/metabolismo , Fatores de Iniciação em Eucariotos/metabolismo , Proteínas de Ligação a Poli(A)/metabolismo , Biossíntese de Proteínas , Purinas , RNA Mensageiro/metabolismo
13.
Cell ; 185(20): 3671-3688.e23, 2022 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-36113466

RESUMO

Bacteria encode reverse transcriptases (RTs) of unknown function that are closely related to group II intron-encoded RTs. We found that a Pseudomonas aeruginosa group II intron-like RT (G2L4 RT) with YIDD instead of YADD at its active site functions in DNA repair in its native host and when expressed in Escherichia coli. G2L4 RT has biochemical activities strikingly similar to those of human DNA repair polymerase Î¸ and uses them for translesion DNA synthesis and double-strand break repair (DSBR) via microhomology-mediated end-joining (MMEJ). We also found that a group II intron RT can function similarly in DNA repair, with reciprocal active-site substitutions showing isoleucine favors MMEJ and alanine favors primer extension in both enzymes. These DNA repair functions utilize conserved structural features of non-LTR-retroelement RTs, including human LINE-1 and other eukaryotic non-LTR-retrotransposon RTs, suggesting such enzymes may have inherent ability to function in DSBR in a wide range of organisms.


Assuntos
DNA Polimerase Dirigida por RNA , Retroelementos , Alanina/genética , Reparo do DNA por Junção de Extremidades , Reparo do DNA , RNA Polimerases Dirigidas por DNA/genética , Humanos , Íntrons , Isoleucina/genética , DNA Polimerase Dirigida por RNA/química
14.
Cell ; 184(7): 1757-1774.e14, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33761328

RESUMO

The central pathogen-immune interface in tuberculosis is the granuloma, a complex host immune structure that dictates infection trajectory and physiology. Granuloma macrophages undergo a dramatic transition in which entire epithelial modules are induced and define granuloma architecture. In tuberculosis, relatively little is known about the host signals that trigger this transition. Using the zebrafish-Mycobacterium marinum model, we identify the basis of granuloma macrophage transformation. Single-cell RNA-sequencing analysis of zebrafish granulomas and analysis of Mycobacterium tuberculosis-infected macaques reveal that, even in the presence of robust type 1 immune responses, countervailing type 2 signals associate with macrophage epithelialization. We find that type 2 immune signaling, mediated via stat6, is absolutely required for epithelialization and granuloma formation. In mixed chimeras, stat6 acts cell autonomously within macrophages, where it is required for epithelioid transformation and incorporation into necrotic granulomas. These findings establish the signaling pathway that produces the hallmark structure of mycobacterial infection.


Assuntos
Granuloma/patologia , Imunidade/fisiologia , Infecções por Mycobacterium não Tuberculosas/patologia , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/metabolismo , Caderinas/genética , Caderinas/metabolismo , Diferenciação Celular , Modelos Animais de Doenças , Células Epitelioides/citologia , Células Epitelioides/imunologia , Células Epitelioides/metabolismo , Granuloma/imunologia , Granuloma/metabolismo , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Interferon gama/metabolismo , Interleucina-12/metabolismo , Macrófagos/citologia , Macrófagos/imunologia , Macrófagos/metabolismo , Infecções por Mycobacterium não Tuberculosas/imunologia , Mycobacterium marinum/isolamento & purificação , Mycobacterium marinum/fisiologia , Necrose , RNA Guia de Cinetoplastídeos/metabolismo , Receptores de Interleucina-4/antagonistas & inibidores , Receptores de Interleucina-4/genética , Receptores de Interleucina-4/metabolismo , Fator de Transcrição STAT6/antagonistas & inibidores , Fator de Transcrição STAT6/genética , Fator de Transcrição STAT6/metabolismo , Transdução de Sinais , Peixe-Zebra/crescimento & desenvolvimento , Peixe-Zebra/metabolismo
15.
Cell ; 184(4): 931-942.e18, 2021 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-33571431

RESUMO

The D1- and D2-dopamine receptors (D1R and D2R), which signal through Gs and Gi, respectively, represent the principal stimulatory and inhibitory dopamine receptors in the central nervous system. D1R and D2R also represent the main therapeutic targets for Parkinson's disease, schizophrenia, and many other neuropsychiatric disorders, and insight into their signaling is essential for understanding both therapeutic and side effects of dopaminergic drugs. Here, we report four cryoelectron microscopy (cryo-EM) structures of D1R-Gs and D2R-Gi signaling complexes with selective and non-selective dopamine agonists, including two currently used anti-Parkinson's disease drugs, apomorphine and bromocriptine. These structures, together with mutagenesis studies, reveal the conserved binding mode of dopamine agonists, the unique pocket topology underlying ligand selectivity, the conformational changes in receptor activation, and potential structural determinants for G protein-coupling selectivity. These results provide both a molecular understanding of dopamine signaling and multiple structural templates for drug design targeting the dopaminergic system.


Assuntos
Receptores de Dopamina D1/química , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/química , Receptores de Dopamina D2/metabolismo , Transdução de Sinais , 2,3,4,5-Tetra-Hidro-7,8-Di-Hidroxi-1-Fenil-1H-3-Benzazepina/análogos & derivados , 2,3,4,5-Tetra-Hidro-7,8-Di-Hidroxi-1-Fenil-1H-3-Benzazepina/farmacologia , Sequência de Aminoácidos , Sequência Conservada , Microscopia Crioeletrônica , AMP Cíclico/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Células HEK293 , Humanos , Ligantes , Modelos Moleculares , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Receptores Adrenérgicos beta 2/metabolismo , Receptores de Dopamina D1/ultraestrutura , Receptores de Dopamina D2/ultraestrutura , Homologia Estrutural de Proteína
16.
Cell ; 184(13): 3426-3437.e8, 2021 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-33991487

RESUMO

We identified an emerging severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant by viral whole-genome sequencing of 2,172 nasal/nasopharyngeal swab samples from 44 counties in California, a state in the western United States. Named B.1.427/B.1.429 to denote its two lineages, the variant emerged in May 2020 and increased from 0% to >50% of sequenced cases from September 2020 to January 2021, showing 18.6%-24% increased transmissibility relative to wild-type circulating strains. The variant carries three mutations in the spike protein, including an L452R substitution. We found 2-fold increased B.1.427/B.1.429 viral shedding in vivo and increased L452R pseudovirus infection of cell cultures and lung organoids, albeit decreased relative to pseudoviruses carrying the N501Y mutation common to variants B.1.1.7, B.1.351, and P.1. Antibody neutralization assays revealed 4.0- to 6.7-fold and 2.0-fold decreases in neutralizing titers from convalescent patients and vaccine recipients, respectively. The increased prevalence of a more transmissible variant in California exhibiting decreased antibody neutralization warrants further investigation.


Assuntos
Anticorpos Neutralizantes/imunologia , COVID-19/imunologia , COVID-19/transmissão , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Humanos , Mutação/genética , Sequenciamento Completo do Genoma/métodos
17.
Cell ; 184(24): 5869-5885.e25, 2021 11 24.
Artigo em Inglês | MEDLINE | ID: mdl-34758294

RESUMO

RTN4-binding proteins were widely studied as "NoGo" receptors, but their physiological interactors and roles remain elusive. Similarly, BAI adhesion-GPCRs were associated with numerous activities, but their ligands and functions remain unclear. Using unbiased approaches, we observed an unexpected convergence: RTN4 receptors are high-affinity ligands for BAI adhesion-GPCRs. A single thrombospondin type 1-repeat (TSR) domain of BAIs binds to the leucine-rich repeat domain of all three RTN4-receptor isoforms with nanomolar affinity. In the 1.65 Å crystal structure of the BAI1/RTN4-receptor complex, C-mannosylation of tryptophan and O-fucosylation of threonine in the BAI TSR-domains creates a RTN4-receptor/BAI interface shaped by unusual glycoconjugates that enables high-affinity interactions. In human neurons, RTN4 receptors regulate dendritic arborization, axonal elongation, and synapse formation by differential binding to glial versus neuronal BAIs, thereby controlling neural network activity. Thus, BAI binding to RTN4/NoGo receptors represents a receptor-ligand axis that, enabled by rare post-translational modifications, controls development of synaptic circuits.


Assuntos
Inibidores da Angiogênese/metabolismo , Encéfalo/metabolismo , Neurogênese , Neurônios/metabolismo , Proteínas Nogo/metabolismo , Receptores Nogo/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Adipocinas/metabolismo , Sequência de Aminoácidos , Animais , Axônios/metabolismo , Adesão Celular , Moléculas de Adesão Celular Neuronais/metabolismo , Complemento C1q/metabolismo , Dendritos/metabolismo , Glicosilação , Células HEK293 , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Ligantes , Camundongos Endogâmicos C57BL , Rede Nervosa/metabolismo , Polissacarídeos/metabolismo , Ligação Proteica , Domínios Proteicos , Deleção de Sequência , Sinapses/metabolismo , Transmissão Sináptica/fisiologia
18.
Cell ; 184(15): 3981-3997.e22, 2021 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-34157301

RESUMO

A fraction of mature T cells can be activated by peripheral self-antigens, potentially eliciting host autoimmunity. We investigated homeostatic control of self-activated T cells within unperturbed tissue environments by combining high-resolution multiplexed and volumetric imaging with computational modeling. In lymph nodes, self-activated T cells produced interleukin (IL)-2, which enhanced local regulatory T cell (Treg) proliferation and inhibitory functionality. The resulting micro-domains reciprocally constrained inputs required for damaging effector responses, including CD28 co-stimulation and IL-2 signaling, constituting a negative feedback circuit. Due to these local constraints, self-activated T cells underwent transient clonal expansion, followed by rapid death ("pruning"). Computational simulations and experimental manipulations revealed the feedback machinery's quantitative limits: modest reductions in Treg micro-domain density or functionality produced non-linear breakdowns in control, enabling self-activated T cells to subvert pruning. This fine-tuned, paracrine feedback process not only enforces immune homeostasis but also establishes a sharp boundary between autoimmune and host-protective T cell responses.


Assuntos
Retroalimentação Fisiológica , Homeostase/imunologia , Ativação Linfocitária/imunologia , Linfócitos T Reguladores/imunologia , Animais , Autoantígenos/imunologia , Linfócitos T CD4-Positivos/imunologia , Proliferação de Células , Interleucina-2/metabolismo , Microdomínios da Membrana/metabolismo , Camundongos Endogâmicos C57BL , Modelos Imunológicos , Comunicação Parácrina , Transdução de Sinais
19.
Cell ; 183(6): 1617-1633.e22, 2020 12 10.
Artigo em Inglês | MEDLINE | ID: mdl-33259802

RESUMO

Histone H3.3 glycine 34 to arginine/valine (G34R/V) mutations drive deadly gliomas and show exquisite regional and temporal specificity, suggesting a developmental context permissive to their effects. Here we show that 50% of G34R/V tumors (n = 95) bear activating PDGFRA mutations that display strong selection pressure at recurrence. Although considered gliomas, G34R/V tumors actually arise in GSX2/DLX-expressing interneuron progenitors, where G34R/V mutations impair neuronal differentiation. The lineage of origin may facilitate PDGFRA co-option through a chromatin loop connecting PDGFRA to GSX2 regulatory elements, promoting PDGFRA overexpression and mutation. At the single-cell level, G34R/V tumors harbor dual neuronal/astroglial identity and lack oligodendroglial programs, actively repressed by GSX2/DLX-mediated cell fate specification. G34R/V may become dispensable for tumor maintenance, whereas mutant-PDGFRA is potently oncogenic. Collectively, our results open novel research avenues in deadly tumors. G34R/V gliomas are neuronal malignancies where interneuron progenitors are stalled in differentiation by G34R/V mutations and malignant gliogenesis is promoted by co-option of a potentially targetable pathway, PDGFRA signaling.


Assuntos
Neoplasias Encefálicas/genética , Carcinogênese/genética , Glioma/genética , Histonas/genética , Interneurônios/metabolismo , Mutação/genética , Células-Tronco Neurais/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Neoplasias Encefálicas/patologia , Carcinogênese/patologia , Linhagem da Célula , Reprogramação Celular/genética , Cromatina/metabolismo , Embrião de Mamíferos/metabolismo , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Glioma/patologia , Histonas/metabolismo , Lisina/metabolismo , Camundongos Endogâmicos C57BL , Modelos Biológicos , Gradação de Tumores , Oligodendroglia/metabolismo , Regiões Promotoras Genéticas/genética , Prosencéfalo/embriologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Transcrição Gênica , Transcriptoma/genética
20.
Cell ; 181(2): 442-459.e29, 2020 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-32302573

RESUMO

Single-cell RNA sequencing (scRNA-seq) is a powerful tool for defining cellular diversity in tumors, but its application toward dissecting mechanisms underlying immune-modulating therapies is scarce. We performed scRNA-seq analyses on immune and stromal populations from colorectal cancer patients, identifying specific macrophage and conventional dendritic cell (cDC) subsets as key mediators of cellular cross-talk in the tumor microenvironment. Defining comparable myeloid populations in mouse tumors enabled characterization of their response to myeloid-targeted immunotherapy. Treatment with anti-CSF1R preferentially depleted macrophages with an inflammatory signature but spared macrophage populations that in mouse and human expresses pro-angiogenic/tumorigenic genes. Treatment with a CD40 agonist antibody preferentially activated a cDC population and increased Bhlhe40+ Th1-like cells and CD8+ memory T cells. Our comprehensive analysis of key myeloid subsets in human and mouse identifies critical cellular interactions regulating tumor immunity and defines mechanisms underlying myeloid-targeted immunotherapies currently undergoing clinical testing.


Assuntos
Neoplasias do Colo/patologia , Células Mieloides/metabolismo , Análise de Célula Única/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Sequência de Bases/genética , Linfócitos T CD8-Positivos/imunologia , China , Neoplasias do Colo/terapia , Neoplasias Colorretais/patologia , Células Dendríticas/imunologia , Feminino , Humanos , Imunoterapia , Macrófagos/imunologia , Masculino , Camundongos , Pessoa de Meia-Idade , Análise de Sequência de RNA/métodos , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia
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