RESUMO
NLRs constitute a large, highly conserved family of cytosolic pattern recognition receptors that are central to health and disease, making them key therapeutic targets. NLRC5 is an enigmatic NLR with mutations associated with inflammatory and infectious diseases, but little is known about its function as an innate immune sensor and cell death regulator. Therefore, we screened for NLRC5's role in response to infections, PAMPs, DAMPs, and cytokines. We identified that NLRC5 acts as an innate immune sensor to drive inflammatory cell death, PANoptosis, in response to specific ligands, including PAMP/heme and heme/cytokine combinations. NLRC5 interacted with NLRP12 and PANoptosome components to form a cell death complex, suggesting an NLR network forms similar to those in plants. Mechanistically, TLR signaling and NAD+ levels regulated NLRC5 expression and ROS production to control cell death. Furthermore, NLRC5-deficient mice were protected in hemolytic and inflammatory models, suggesting that NLRC5 could be a potential therapeutic target.
Assuntos
Inflamação , Peptídeos e Proteínas de Sinalização Intracelular , NAD , Animais , Camundongos , Inflamação/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , NAD/metabolismo , Humanos , Imunidade Inata , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio/metabolismo , Camundongos Knockout , Transdução de Sinais , Células HEK293 , Inflamassomos/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas Reguladoras de Apoptose/genética , Receptores Toll-Like/metabolismo , Masculino , Citocinas/metabolismo , Proteínas de Ligação ao CálcioRESUMO
Searching for factors to improve knockin efficiency for therapeutic applications, biotechnology, and generation of non-human primate models of disease, we found that the strand exchange protein RAD51 can significantly increase Cas9-mediated homozygous knockin in mouse embryos through an interhomolog repair (IHR) mechanism. IHR is a hallmark of meiosis but only occurs at low frequencies in somatic cells, and its occurrence in zygotes is controversial. Using multiple approaches, we provide evidence for an endogenous IHR mechanism in the early embryo that can be enhanced by RAD51. This process can be harnessed to generate homozygotes from wild-type zygotes using exogenous donors and to convert heterozygous alleles into homozygous alleles without exogenous templates. Furthermore, we identify additional IHR-promoting factors and describe features of IHR events. Together, our findings show conclusive evidence for IHR in mouse embryos and describe an efficient method for enhanced gene conversion.
Assuntos
Reparo do DNA/genética , Conversão Gênica , Rad51 Recombinase/metabolismo , Alelos , Animais , Sequência de Bases , Proteína 9 Associada à CRISPR/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ciclo Celular/metabolismo , Cromossomos de Mamíferos/genética , Quebras de DNA de Cadeia Dupla , Embrião de Mamíferos , Feminino , Loci Gênicos , Recombinação Homóloga/genética , Homozigoto , Humanos , Mutação INDEL/genética , Camundongos Endogâmicos C57BL , Mosaicismo , Proteínas Nucleares/metabolismo , Polimorfismo de Nucleotídeo Único/genética , Ribonucleoproteínas/metabolismo , Zigoto/metabolismoRESUMO
Inflammasomes are cytosolic multiprotein complexes that assemble in response to a variety of infectious and noxious insults. Inflammasomes play a critical role in the initiation of innate immune responses, primarily by serving as platforms for the activation of inflammatory caspase proteases. One such caspase, CASPASE-1 (CASP1), initiates innate immune responses by cleaving pro-IL-1ß and pro-IL-18, leading to their activation and release. CASP1 and another inflammatory caspase termed CASP11 can also initiate a rapid and inflammatory form of cell death termed pyroptosis. Several distinct inflammasomes have been described, each of which contains a unique sensor protein of the NLR (nucleotide-binding domain, leucine-rich repeat-containing) superfamily or the PYHIN (PYRIN and HIN-200 domain-containing) superfamily. Here we describe the surprisingly diverse mechanisms by which NLR/PYHIN proteins sense bacteria and initiate innate immune responses. We conclude that inflammasomes represent a highly adaptable scaffold ideally suited for detecting and initiating rapid innate responses to diverse and rapidly evolving bacteria.
Assuntos
Bactérias/patogenicidade , Inflamassomos/metabolismo , Animais , Bacillus anthracis/patogenicidade , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Proteínas Adaptadoras de Sinalização CARD/fisiologia , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/fisiologia , Flagelos/metabolismo , Flagelos/fisiologia , Humanos , Inflamassomos/genética , Inflamassomos/fisiologia , Legionella pneumophila/patogenicidade , Listeria monocytogenes/patogenicidade , Salmonella typhimurium/patogenicidadeRESUMO
Social impairment is frequently associated with mitochondrial dysfunction and altered neurotransmission. Although mitochondrial function is crucial for brain homeostasis, it remains unknown whether mitochondrial disruption contributes to social behavioral deficits. Here, we show that Drosophila mutants in the homolog of the human CYFIP1, a gene linked to autism and schizophrenia, exhibit mitochondrial hyperactivity and altered group behavior. We identify the regulation of GABA availability by mitochondrial activity as a biologically relevant mechanism and demonstrate its contribution to social behavior. Specifically, increased mitochondrial activity causes gamma aminobutyric acid (GABA) sequestration in the mitochondria, reducing GABAergic signaling and resulting in social deficits. Pharmacological and genetic manipulation of mitochondrial activity or GABA signaling corrects the observed abnormalities. We identify Aralar as the mitochondrial transporter that sequesters GABA upon increased mitochondrial activity. This study increases our understanding of how mitochondria modulate neuronal homeostasis and social behavior under physiopathological conditions.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Drosophila/metabolismo , Mitocôndrias/metabolismo , Ácido gama-Aminobutírico/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Animais Geneticamente Modificados , Ácido Aspártico/metabolismo , Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/fisiologia , Proteínas de Drosophila/fisiologia , Drosophila melanogaster/metabolismo , Glucose/metabolismo , Homeostase , Humanos , Masculino , Mitocôndrias/genética , Proteínas de Transporte da Membrana Mitocondrial/genética , Proteínas Mitocondriais/metabolismo , Neurônios/metabolismo , Comportamento Social , Transmissão Sináptica , Ácido gama-Aminobutírico/genéticaRESUMO
Varying pH of luminal fluid along the female reproductive tract is a physiological cue that modulates sperm motility. CatSper is a sperm-specific, pH-sensitive calcium channel essential for hyperactivated motility and male fertility. Multi-subunit CatSper channel complexes organize linear Ca2+ signaling nanodomains along the sperm tail. Here, we identify EF-hand calcium-binding domain-containing protein 9 (EFCAB9) as a bifunctional, cytoplasmic machine modulating the channel activity and the domain organization of CatSper. Knockout mice studies demonstrate that EFCAB9, in complex with the CatSper subunit, CATSPERζ, is essential for pH-dependent and Ca2+-sensitive activation of the CatSper channel. In the absence of EFCAB9, sperm motility and fertility is compromised, and the linear arrangement of the Ca2+ signaling domains is disrupted. EFCAB9 interacts directly with CATSPERζ in a Ca2+-dependent manner and dissociates at elevated pH. These observations suggest that EFCAB9 is a long-sought, intracellular, pH-dependent Ca2+ sensor that triggers changes in sperm motility.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Motilidade dos Espermatozoides/fisiologia , Animais , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Sinalização do Cálcio/fisiologia , Proteínas de Ligação ao Cálcio/fisiologia , Linhagem Celular , Membrana Celular/metabolismo , Fertilidade , Células HEK293 , Humanos , Concentração de Íons de Hidrogênio , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Espermatozoides/metabolismoRESUMO
Signaling through Notch receptors intrinsically regulates tumor cell development and growth. Here, we studied the role of the Notch ligand Jagged2 on immune evasion in non-small cell lung cancer (NSCLC). Higher expression of JAG2 in NSCLC negatively correlated with survival. In NSCLC pre-clinical models, deletion of Jag2, but not Jag1, in cancer cells attenuated tumor growth and activated protective anti-tumor T cell responses. Jag2-/- lung tumors exhibited higher frequencies of macrophages that expressed immunostimulatory mediators and triggered T cell-dependent anti-tumor immunity. Mechanistically, Jag2 ablation promoted Nr4a-mediated induction of Notch ligands DLL1/4 on cancer cells. DLL1/4-initiated Notch1/2 signaling in macrophages induced the expression of transcription factor IRF4 and macrophage immunostimulatory functionality. IRF4 expression was required for the anti-tumor effects of Jag2 deletion in lung tumors. Antibody targeting of Jagged2 inhibited tumor growth and activated IRF4-driven macrophage-mediated anti-tumor immunity. Thus, Jagged2 orchestrates immunosuppressive systems in NSCLC that can be overcome to incite macrophage-mediated anti-tumor immunity.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Fatores Reguladores de Interferon , Proteína Jagged-2 , Neoplasias Pulmonares , Camundongos Knockout , Macrófagos Associados a Tumor , Animais , Humanos , Camundongos , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/genética , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Linhagem Celular Tumoral , Fatores Reguladores de Interferon/metabolismo , Fatores Reguladores de Interferon/genética , Proteína Jagged-1/metabolismo , Proteína Jagged-1/genética , Proteína Jagged-2/metabolismo , Proteína Jagged-2/genética , Proteína Jagged-2/imunologia , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/genética , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Receptor Notch1/metabolismo , Receptor Notch1/genética , Receptores Notch/metabolismo , Transdução de Sinais , Evasão Tumoral/imunologia , Macrófagos Associados a Tumor/imunologia , Macrófagos Associados a Tumor/metabolismoRESUMO
As a critical step during innate response, the cytoplasmic ß subunit (IFN-γR2) of interferon-γ receptor (IFN-γR) is induced and translocates to plasma membrane to join α subunit to form functional IFN-γR to mediate IFN-γ signaling. However, the mechanism driving membrane translocation and its significance remain largely unknown. We found, unexpectedly, that mice deficient in E-selectin, an endothelial cell-specific adhesion molecule, displayed impaired innate activation of macrophages upon Listeria monocytogenes infection yet had increased circulating IFN-γ. Inflammatory macrophages from E-selectin-deficient mice had less surface IFN-γR2 and impaired IFN-γ signaling. BTK elicited by extrinsic E-selectin engagement phosphorylates cytoplasmic IFN-γR2, facilitating EFhd2 binding and promoting IFN-γR2 trafficking from Golgi to cell membrane. Our findings demonstrate that membrane translocation of cytoplasmic IFN-γR2 is required to activate macrophage innate response against intracellular bacterial infection, identifying the assembly of functional cytokine receptors on cell membrane as an important layer in innate activation and cytokine signaling.
Assuntos
Selectina E/metabolismo , Imunidade Inata , Receptores de Interferon/metabolismo , Animais , Proteínas de Ligação ao Cálcio/antagonistas & inibidores , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Membrana Celular/metabolismo , Selectina E/deficiência , Selectina E/genética , Complexo de Golgi/metabolismo , Interferon gama/sangue , Interferon gama/metabolismo , Listeria/patogenicidade , Ativação de Macrófagos , Macrófagos/citologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosforilação , Transporte Proteico , Células RAW 264.7 , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Receptores de Interferon/deficiência , Receptores de Interferon/genética , Transdução de Sinais , Receptor de Interferon gamaRESUMO
The cerebral cortex underwent rapid expansion and increased complexity during recent hominid evolution. Gene duplications constitute a major evolutionary force, but their impact on human brain development remains unclear. Using tailored RNA sequencing (RNA-seq), we profiled the spatial and temporal expression of hominid-specific duplicated (HS) genes in the human fetal cortex and identified a repertoire of 35 HS genes displaying robust and dynamic patterns during cortical neurogenesis. Among them NOTCH2NL, human-specific paralogs of the NOTCH2 receptor, stood out for their ability to promote cortical progenitor maintenance. NOTCH2NL promote the clonal expansion of human cortical progenitors, ultimately leading to higher neuronal output. At the molecular level, NOTCH2NL function by activating the Notch pathway through inhibition of cis Delta/Notch interactions. Our study uncovers a large repertoire of recently evolved genes active during human corticogenesis and reveals how human-specific NOTCH paralogs may have contributed to the expansion of the human cortex.
Assuntos
Córtex Cerebral/metabolismo , Regulação da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas de Membrana/metabolismo , Neurogênese , Neurônios/metabolismo , Receptor Notch2/genética , Sequência de Aminoácidos , Proteínas de Ligação ao Cálcio , Diferenciação Celular/genética , Análise por Conglomerados , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Hibridização In Situ , Células-Tronco Neurais/metabolismo , Transdução de SinaisRESUMO
Inflammasome activation is critical for host defenses against various microbial infections. Activation of the NLRC4 inflammasome requires detection of flagellin or type III secretion system (T3SS) components by NLR family apoptosis inhibitory proteins (NAIPs); yet how this pathway is regulated is unknown. Here, we found that interferon regulatory factor 8 (IRF8) is required for optimal activation of the NLRC4 inflammasome in bone-marrow-derived macrophages infected with Salmonella Typhimurium, Burkholderia thailandensis, or Pseudomonas aeruginosa but is dispensable for activation of the canonical and non-canonical NLRP3, AIM2, and Pyrin inflammasomes. IRF8 governs the transcription of Naips to allow detection of flagellin or T3SS proteins to mediate NLRC4 inflammasome activation. Furthermore, we found that IRF8 confers protection against bacterial infection in vivo, owing to its role in inflammasome-dependent cytokine production and pyroptosis. Altogether, our findings suggest that IRF8 is a critical regulator of NAIPs and NLRC4 inflammasome activation for defense against bacterial infection.
Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Inflamassomos/metabolismo , Fatores Reguladores de Interferon/metabolismo , Proteína Inibidora de Apoptose Neuronal/metabolismo , Animais , Proteínas Reguladoras de Apoptose/genética , Proteínas de Ligação ao Cálcio/genética , Células Cultivadas , Citocinas/metabolismo , Ensaio de Desvio de Mobilidade Eletroforética , Flagelina/metabolismo , Fatores Reguladores de Interferon/antagonistas & inibidores , Fatores Reguladores de Interferon/genética , Macrófagos/citologia , Macrófagos/metabolismo , Macrófagos/microbiologia , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteína Inibidora de Apoptose Neuronal/genética , Regiões Promotoras Genéticas , Ligação Proteica , Pseudomonas aeruginosa/patogenicidade , Piroptose , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Salmonella typhimurium/patogenicidade , Transcrição GênicaRESUMO
Cerebral cortex size differs dramatically between reptiles, birds, and mammals, owing to developmental differences in neuron production. In mammals, signaling pathways regulating neurogenesis have been identified, but genetic differences behind their evolution across amniotes remain unknown. We show that direct neurogenesis from radial glia cells, with limited neuron production, dominates the avian, reptilian, and mammalian paleocortex, whereas in the evolutionarily recent mammalian neocortex, most neurogenesis is indirect via basal progenitors. Gain- and loss-of-function experiments in mouse, chick, and snake embryos and in human cerebral organoids demonstrate that high Slit/Robo and low Dll1 signaling, via Jag1 and Jag2, are necessary and sufficient to drive direct neurogenesis. Attenuating Robo signaling and enhancing Dll1 in snakes and birds recapitulates the formation of basal progenitors and promotes indirect neurogenesis. Our study identifies modulation in activity levels of conserved signaling pathways as a primary mechanism driving the expansion and increased complexity of the mammalian neocortex during amniote evolution.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neurogênese/genética , Receptores Imunológicos/genética , Receptores Imunológicos/metabolismo , Animais , Proteínas de Ligação ao Cálcio , Córtex Cerebral/metabolismo , Embrião de Galinha , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas de Homeodomínio , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Proteína Jagged-1 , Proteína Jagged-2 , Mamíferos/embriologia , Camundongos , Camundongos Endogâmicos C57BL , Neocórtex/fisiologia , Células-Tronco Neurais , Neurogênese/fisiologia , Neuroglia/fisiologia , Neurônios , Fator de Transcrição PAX6/metabolismo , Proteínas Repressoras , Transdução de Sinais , Serpentes/embriologia , Proteínas RoundaboutRESUMO
The Notch signaling pathway comprises multiple ligands that are used in distinct biological contexts. In principle, different ligands could activate distinct target programs in signal-receiving cells, but it is unclear how such ligand discrimination could occur. Here, we show that cells use dynamics to discriminate signaling by the ligands Dll1 and Dll4 through the Notch1 receptor. Quantitative single-cell imaging revealed that Dll1 activates Notch1 in discrete, frequency-modulated pulses that specifically upregulate the Notch target gene Hes1. By contrast, Dll4 activates Notch1 in a sustained, amplitude-modulated manner that predominantly upregulates Hey1 and HeyL. Ectopic expression of Dll1 or Dll4 in chick neural crest produced opposite effects on myogenic differentiation, showing that ligand discrimination can occur in vivo. Finally, analysis of chimeric ligands suggests that ligand-receptor clustering underlies dynamic encoding of ligand identity. The ability of the pathway to utilize ligands as distinct communication channels has implications for diverse Notch-dependent processes.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/metabolismo , Receptor Notch1/metabolismo , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/biossíntese , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Células CHO , Proteínas de Ligação ao Cálcio , Proteínas de Ciclo Celular/biossíntese , Proteínas de Ciclo Celular/genética , Embrião de Galinha , Cricetulus , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Ligantes , Proteínas de Membrana/genética , Camundongos , Receptor Notch1/genética , Proteínas Repressoras/biossíntese , Proteínas Repressoras/genética , Regulação para CimaRESUMO
Synapses are fundamental units of communication in the brain. The prototypical synapse-organizing complex neurexin-neuroligin mediates synapse development and function and is central to a shared genetic risk pathway in autism and schizophrenia. Neurexin's role in synapse development is thought to be mediated purely by its protein domains, but we reveal a requirement for a rare glycan modification. Mice lacking heparan sulfate (HS) on neurexin-1 show reduced survival, as well as structural and functional deficits at central synapses. HS directly binds postsynaptic partners neuroligins and LRRTMs, revealing a dual binding mode involving intrinsic glycan and protein domains for canonical synapse-organizing complexes. Neurexin HS chains also bind novel ligands, potentially expanding the neurexin interactome to hundreds of HS-binding proteins. Because HS structure is heterogeneous, our findings indicate an additional dimension to neurexin diversity, provide a molecular basis for fine-tuning synaptic function, and open therapeutic directions targeting glycan-binding motifs critical for brain development.
Assuntos
Heparitina Sulfato/metabolismo , Moléculas de Adesão de Célula Nervosa/metabolismo , Sinapses/metabolismo , Sequência de Aminoácidos , Animais , Proteínas de Ligação ao Cálcio , Moléculas de Adesão Celular Neuronais/antagonistas & inibidores , Moléculas de Adesão Celular Neuronais/genética , Moléculas de Adesão Celular Neuronais/metabolismo , Drosophila , Proteínas de Drosophila/antagonistas & inibidores , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Feminino , Glicopeptídeos/análise , Heparitina Sulfato/química , Humanos , Proteínas de Membrana , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso , Moléculas de Adesão de Célula Nervosa/antagonistas & inibidores , Moléculas de Adesão de Célula Nervosa/genética , Neurônios/citologia , Neurônios/metabolismo , Ligação Proteica , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Ratos , Alinhamento de SequênciaRESUMO
NOD-like receptors (NLRs) are pattern recognition receptors for diverse innate immune responses. Self-oligomerization after engagement with a ligand is a generally accepted model for the activation of each NLR. We report here that a catalyzer was required for NLR self-oligomerization. PELO, a well-known surveillance factor in translational quality control and/or ribosome rescue, interacted with all cytosolic NLRs and activated their ATPase activity. In the case of flagellin-initiated NLRC4 inflammasome activation, flagellin-bound NAIP5 recruited the first NLRC4 and then PELO was required for correctly assembling the rest of NLRC4s into the NLRC4 complex, one by one, by activating the NLRC4 ATPase activity. Stoichiometric and functional data revealed that PELO was not a structural constituent of the NLRC4 inflammasome but a powerful catalyzer for its assembly. The catalytic role of PELO in the activation of cytosolic NLRs provides insight into NLR activation and provides a direction for future studies of NLR family members.
Assuntos
Proteínas Reguladoras de Apoptose , Inflamassomos , Adenosina Trifosfatases/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Ligação ao Cálcio/química , Proteínas de Ligação ao Cálcio/metabolismo , Flagelina/metabolismo , Inflamassomos/metabolismo , Proteína Inibidora de Apoptose Neuronal/química , Proteína Inibidora de Apoptose Neuronal/metabolismo , Proteínas NLR/metabolismoAssuntos
Calcineurina , Calcineurina/metabolismo , Calcineurina/genética , Animais , Humanos , Camundongos , Regulação da Expressão Gênica , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/genética , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição NFATC/metabolismo , Fatores de Transcrição NFATC/genéticaRESUMO
Resolution of inflammation is essential for tissue homeostasis and represents a promising approach to inflammatory disorders. Here we found that developmental endothelial locus-1 (DEL-1), a secreted protein that inhibits leukocyte-endothelial adhesion and inflammation initiation, also functions as a non-redundant downstream effector in inflammation clearance. In human and mouse periodontitis, waning of inflammation was correlated with DEL-1 upregulation, whereas resolution of experimental periodontitis failed in DEL-1 deficiency. This concept was mechanistically substantiated in acute monosodium-urate-crystal-induced inflammation, where the pro-resolution function of DEL-1 was attributed to effective apoptotic neutrophil clearance (efferocytosis). DEL-1-mediated efferocytosis induced liver X receptor-dependent macrophage reprogramming to a pro-resolving phenotype and was required for optimal production of at least certain specific pro-resolving mediators. Experiments in transgenic mice with cell-specific overexpression of DEL-1 linked its anti-leukocyte-recruitment action to endothelial cell-derived DEL-1 and its efferocytic/pro-resolving action to macrophage-derived DEL-1. Thus, the compartmentalized expression of DEL-1 facilitates distinct homeostatic functions in an appropriate context that can be harnessed therapeutically.
Assuntos
Proteínas de Transporte/metabolismo , Inflamação/imunologia , Macrófagos/fisiologia , Neutrófilos/imunologia , Periodontite/imunologia , Adulto , Animais , Proteínas de Ligação ao Cálcio , Proteínas de Transporte/genética , Moléculas de Adesão Celular , Reprogramação Celular , Citocinas/metabolismo , Regulação da Expressão Gênica , Humanos , Inflamação/induzido quimicamente , Peptídeos e Proteínas de Sinalização Intercelular , Células K562 , Receptores X do Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , FagocitoseRESUMO
During neurodegenerative disease, the toxic accumulation of aggregates and misfolded proteins is often accompanied with widespread changes in peripheral metabolism, even in cells in which the aggregating protein is not present. The mechanism by which the central nervous system elicits a distal reaction to proteotoxic stress remains unknown. We hypothesized that the endocrine communication of neuronal stress plays a causative role in the changes in mitochondrial homeostasis associated with proteotoxic disease states. We find that an aggregation-prone protein expressed in the neurons of C. elegans binds to mitochondria, eliciting a global induction of a mitochondrial-specific unfolded protein response (UPR(mt)), affecting whole-animal physiology. Importantly, dense core vesicle release and secretion of the neurotransmitter serotonin is required for the signal's propagation. Collectively, these data suggest the commandeering of a nutrient sensing network to allow for cell-to-cell communication between mitochondria in response to protein folding stress in the nervous system.
Assuntos
Homeostase , Transdução de Sinais , Resposta a Proteínas não Dobradas , Animais , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Comunicação Celular , Mitocôndrias/metabolismo , Células Neuroendócrinas/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Peptídeos/metabolismo , Dobramento de Proteína , Serotonina/metabolismoRESUMO
Proper establishment of synapses is critical for constructing functional circuits. Interactions between presynaptic neurexins and postsynaptic neuroligins coordinate the formation of synaptic adhesions. An isoform code determines the direct interactions of neurexins and neuroligins across the synapse. However, whether extracellular linker proteins can expand such a code is unknown. Using a combination of in vitro and in vivo approaches, we found that hevin, an astrocyte-secreted synaptogenic protein, assembles glutamatergic synapses by bridging neurexin-1alpha and neuroligin-1B, two isoforms that do not interact with each other. Bridging of neurexin-1alpha and neuroligin-1B via hevin is critical for the formation and plasticity of thalamocortical connections in the developing visual cortex. These results show that astrocytes promote the formation of synapses by modulating neurexin/neuroligin adhesions through hevin secretion. Our findings also provide an important mechanistic insight into how mutations in these genes may lead to circuit dysfunction in diseases such as autism.
Assuntos
Astrócitos/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Moléculas de Adesão Celular Neuronais/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Moléculas de Adesão de Célula Nervosa/metabolismo , Tálamo/metabolismo , Animais , Células COS , Chlorocebus aethiops , Dominância Ocular , Humanos , Camundongos , Camundongos Knockout , Doenças do Sistema Nervoso/metabolismo , Neurônios/metabolismo , Isoformas de Proteínas/metabolismo , Transdução de Sinais , Sinapses/metabolismoRESUMO
The selectivity with which proprioceptive sensory neurons innervate their central and peripheral targets implies that they exhibit distinctions in muscle-type identity. The molecular correlates of proprioceptor identity and its origins remain largely unknown, however. In screens to define muscle-type proprioceptor character, we find all-or-none differences in gene expression for proprioceptors that control antagonistic muscles at a single hindlimb joint. Analysis of three of these genes, cadherin13 (cdh13), semaphorin5a (sema5a), and cartilage-acidic protein-1 (crtac1), reveals expression in proprioceptor subsets that supply muscle groups located at restricted dorsoventral and proximodistal domains of the limb. Genetically altering the dorsoventral character of the limb mesenchyme elicits a change in the profile of proprioceptor cdh13, sema5a, and crtac1 expression. These findings indicate that proprioceptors acquire aspects of their muscle-type identity in response to mesenchymal signals expressed in restricted proximodistal and dorsoventral domains of the developing limb.
Assuntos
Extremidades/embriologia , Mesoderma/metabolismo , Propriocepção , Animais , Caderinas/genética , Proteínas de Ligação ao Cálcio/genética , Embrião de Mamíferos/metabolismo , Extremidades/fisiologia , Camundongos , Músculo Esquelético/inervação , Neurônios/metabolismo , Semaforinas/genética , Transdução de Sinais , TranscriptomaRESUMO
The ubiquitin ligase CUL3 is an essential regulator of neural crest specification whose aberrant activation has been linked to autism, schizophrenia, and hypertension. CUL3 exerts its roles by pairing with â¼90 distinct substrate adaptors, yet how the different CUL3-complexes are activated is poorly understood. Here, we show that CUL3 and its adaptor KLHL12 require two calcium-binding proteins, PEF1 and ALG2, for recognition of their substrate SEC31. PEF1 and ALG2 form a target-specific co-adaptor that translates a transient rise in cytosolic calcium levels into more persistent SEC31 ubiquitylation, which in turn triggers formation of large COPII coats and promotes collagen secretion. As calcium also instructs chondrocyte differentiation and collagen synthesis, calcium-dependent control of CUL3KLHL12 integrates collagen secretion into broader programs of craniofacial bone formation. Our work, therefore, identifies both calcium and CUL3 co-adaptors as important regulators of ubiquitylation events that control human development.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Cálcio/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas Culina/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Proteínas dos Microfilamentos/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Vesículas Revestidas pelo Complexo de Proteína do Envoltório/metabolismo , Colágeno/metabolismo , Células HEK293 , Humanos , Especificidade por Substrato , Ubiquitinação , Proteínas de Transporte Vesicular/metabolismoRESUMO
Viable yet damaged cells can accumulate during development and aging. Although eliminating those cells may benefit organ function, identification of this less fit cell population remains challenging. Previously, we identified a molecular mechanism, based on "fitness fingerprints" displayed on cell membranes, which allows direct fitness comparison among cells in Drosophila. Here, we study the physiological consequences of efficient cell selection for the whole organism. We find that fitness-based cell culling is naturally used to maintain tissue health, delay aging, and extend lifespan in Drosophila. We identify a gene, azot, which ensures the elimination of less fit cells. Lack of azot increases morphological malformations and susceptibility to random mutations and accelerates tissue degeneration. On the contrary, improving the efficiency of cell selection is beneficial for tissue health and extends lifespan.