RESUMEN
Missense mutations of the tumor suppressor Neurofibromin 2 (NF2/Merlin/schwannomin) result in sporadic to frequent occurrences of tumorigenesis in multiple organs. However, the underlying pathogenicity of NF2-related tumorigenesis remains mostly unknown. Here we found that NF2 facilitated innate immunity by regulating YAP/TAZ-mediated TBK1 inhibition. Unexpectedly, patient-derived individual mutations in the FERM domain of NF2 (NF2m) converted NF2 into a potent suppressor of cGAS-STING signaling. Mechanistically, NF2m gained extreme associations with IRF3 and TBK1 and, upon innate nucleic acid sensing, was directly induced by the activated IRF3 to form cellular condensates, which contained the PP2A complex, to eliminate TBK1 activation. Accordingly, NF2m robustly suppressed STING-initiated antitumor immunity in cancer cell-autonomous and -nonautonomous murine models, and NF2m-IRF3 condensates were evident in human vestibular schwannomas. Our study reports phase separation-mediated quiescence of cGAS-STING signaling by a mutant tumor suppressor and reveals gain-of-function pathogenesis for NF2-related tumors by regulating antitumor immunity.
Asunto(s)
Inmunidad Innata , Proteínas de la Membrana/metabolismo , Mutación Missense , Neoplasias/metabolismo , Neurofibromina 2/metabolismo , Nucleotidiltransferasas/metabolismo , Escape del Tumor , Animales , Femenino , Regulación Neoplásica de la Expresión Génica , Células HCT116 , Células HEK293 , Humanos , Factor 3 Regulador del Interferón/genética , Factor 3 Regulador del Interferón/metabolismo , Macrófagos Peritoneales/inmunología , Macrófagos Peritoneales/metabolismo , Masculino , Melanoma Experimental/genética , Melanoma Experimental/inmunología , Melanoma Experimental/metabolismo , Melanoma Experimental/patología , Proteínas de la Membrana/genética , Ratones Endogámicos C57BL , Ratones Transgénicos , Neoplasias/genética , Neoplasias/inmunología , Neoplasias/patología , Neurofibromina 2/genética , Nucleotidiltransferasas/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de SeñalRESUMEN
Mitochondrial morphology shifts rapidly to manage cellular metabolism, organelle integrity, and cell fate. It remains unknown whether innate nucleic acid sensing, the central and general mechanisms of monitoring both microbial invasion and cellular damage, can reprogram and govern mitochondrial dynamics and function. Here, we unexpectedly observed that upon activation of RIG-I-like receptor (RLR)-MAVS signaling, TBK1 directly phosphorylated DRP1/DNM1L, which disabled DRP1, preventing its high-order oligomerization and mitochondrial fragmentation function. The TBK1-DRP1 axis was essential for assembly of large MAVS aggregates and healthy antiviral immunity and underlay nutrient-triggered mitochondrial dynamics and cell fate determination. Knockin (KI) strategies mimicking TBK1-DRP1 signaling produced dominant-negative phenotypes reminiscent of human DRP1 inborn mutations, while interrupting the TBK1-DRP1 connection compromised antiviral responses. Thus, our findings establish an unrecognized function of innate immunity governing both morphology and physiology of a major organelle, identify a lacking loop during innate RNA sensing, and report an elegant mechanism of shaping mitochondrial dynamics.
Asunto(s)
Dinaminas/metabolismo , Mitocondrias/fisiología , Proteínas Serina-Treonina Quinasas/metabolismo , ARN/metabolismo , Pez Cebra/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Proteína 58 DEAD Box/genética , Proteína 58 DEAD Box/metabolismo , Dinaminas/genética , Células HCT116 , Células HEK293 , Humanos , Masculino , Ratones , Ratones Transgénicos , Mutación , Proteínas Serina-Treonina Quinasas/genética , ARN/genética , Transducción de Señal/genética , Pez Cebra/genética , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismoRESUMEN
Teleost fish type I IFNs and the associated receptors from the cytokine receptor family B (CRFB) are characterized by remarkable diversity and complexity. How the fish type I IFNs bind to their receptors is still not fully understood. In this study, we demonstrate that CRFB1 and CRFB5 constitute the receptor pair through which type I subgroup d IFN (IFNd) from large yellow croaker, Larimichthys crocea, activates the conserved JAK-STAT signaling pathway as a part of the antiviral response. Our data suggest that L. crocea IFNd (LcIFNd) has a higher binding affinity with L. crocea CRFB5 (LcCRFB5) than with LcCRFB1. Furthermore, we report the crystal structure of LcIFNd at a 1.49-Å resolution and construct structural models of LcIFNd in binary complexes with predicted structures of extracellular regions of LcCRFB1 and LcCRFB5, respectively. Despite striking similarities in overall architectures of LcIFNd and its ortholog human IFN-ω, the receptor binding patterns between LcIFNd and its receptors show that teleost and mammalian type I IFNs may have differentially selected helices that bind to their homologous receptors. Correspondingly, key residues mediating binding of LcIFNd to LcCRFB1 and LcCRFB5 are largely distinct from the receptor-interacting residues in other fish and mammalian type I IFNs. Our findings reveal a ligand/receptor complex binding mechanism of IFNd in teleost fish, thus providing new insights into the function and evolution of type I IFNs.
Asunto(s)
Interferón Tipo I , Perciformes , Animales , Humanos , Filogenia , Peces/metabolismo , Interferón Tipo I/metabolismo , Proteínas de Peces/genética , Mamíferos/metabolismoRESUMEN
The Legionella pneumophila effector MavC induces ubiquitination of the E2 ubiquitin-conjugating enzyme UBE2N by transglutamination, thereby abolishing its function in the synthesis of K63 -type polyubiquitin chains. The inhibition of UBE2N activity creates a conundrum because this E2 enzyme is important in multiple signaling pathways, including some that are important for intracellular L. pneumophila replication. Here, we show that prolonged inhibition of UBE2N activity by MavC restricts intracellular bacterial replication and that the activity of UBE2N is restored by MvcA, an ortholog of MavC (50% identity) with ubiquitin deamidase activity. MvcA functions to deubiquitinate UBE2N-Ub using the same catalytic triad required for its deamidase activity. Structural analysis of the MvcA-UBE2N-Ub complex reveals a crucial role of the insertion domain in MvcA in substrate recognition. Our study establishes a deubiquitination mechanism catalyzed by a deamidase, which, together with MavC, imposes temporal regulation of the activity of UBE2N during L. pneumophila infection.
Asunto(s)
Proteínas Bacterianas/metabolismo , Legionella pneumophila/fisiología , Transducción de Señal , Enzimas Ubiquitina-Conjugadoras/metabolismo , Ubiquitina/metabolismo , Proteínas Bacterianas/genética , Células HEK293 , Humanos , Legionella pneumophila/enzimología , Legionella pneumophila/genética , Legionella pneumophila/patogenicidad , Poliubiquitina/metabolismo , Sistemas de Secreción Tipo IV , Enzimas Ubiquitina-Conjugadoras/genética , UbiquitinaciónRESUMEN
In mammals, type I IFNs, which commonly contain one or two disulfide bonds, activate the JAK-STAT signaling pathway through binding to the common cell surface receptor formed by IFN-α/ß receptor (IFNAR)1 and IFNAR2 subunits. Although type I IFNs are also known to be essential for antiviral defense in teleost fish, very little is known about mechanisms underlying the recognition of fish type I IFNs by associated receptors. In this study, we demonstrate that a type I IFN of large yellow croaker Larimichthys crocea (LcIFNi), belonging to a new subgroup of fish type I IFNs, triggers antiviral response via the conserved JAK-STAT pathway through stable binding with a heterodimeric receptor comprising subunits LcCRFB5 and LcCRFB2. LcIFNi binds to LcCRFB5 with a much higher affinity than to LcCRFB2. Furthermore, we determined the crystal structure of LcIFNi at a 1.39 Å resolution. The high-resolution structure is, to our knowledge, the first reported structure of a type I IFN with three disulfide bonds, all of which were found to be indispensable for folding and stability of LcIFNi. Using structural analysis, mutagenesis, and biochemical assays, we identified key LcIFNi residues involved in receptor interaction and proposed a structural model of LcIFNi bound to the LcCRFB2-LcCRFB5 receptor. The results show that LcIFNi-LcCRFB2 exhibits a similar binding pattern to human IFN-ω-IFNAR2, whereas the binding pattern of LcIFNi-LcCRFB5 is quite different from that of IFN-ω-IFNAR1. Altogether, our findings reveal the structural basis for receptor interaction and signaling of a type I IFN with three disulfide bonds and provide new insights into the mechanisms underlying type I IFN recognition in teleosts.
Asunto(s)
Perciformes , Transducción de Señal , Animales , Antivirales , Disulfuros/metabolismo , Peces/metabolismo , Humanos , Quinasas Janus/metabolismo , Mamíferos/metabolismo , Receptor de Interferón alfa y beta/metabolismo , Factores de Transcripción STAT/metabolismoRESUMEN
Proteasomes are highly abundant and conserved protease complexes that eliminate unwanted proteins in the cells. As a single-chain ATP-independent nuclear proteasome activator, proteasome activator 200 (PA200) associates with 20S core particle to form proteasome complex that catalyzes polyubiquitin-independent degradation of acetylated histones, thus playing a pivotal role in DNA repair and spermatogenesis. Here, we present cryo-electron microscopy (cryo-EM) structures of the human PA200-20S complex and PA200 at 2.72 Å and 3.75 Å, respectively. PA200 exhibits a dome-like architecture that caps 20S and uses its C-terminal YYA (Tyr-Tyr-Ala) to induce the α-ring rearrangements and partial opening of the 20S gate. Our structural data also indicate that PA200 has two openings formed by numerous positively charged residues that respectively bind (5,6)-bisdiphosphoinositol tetrakisphosphate (5,6[PP]2-InsP4) and inositol hexakisphosphate (InsP6) and are likely to be the gates that lead unfolded proteins through PA200 and into the 20S. Besides, our structural analysis of PA200 found that the bromodomain (BRD)-like (BRDL) domain of PA200 shows considerable sequence variation in comparison to other human BRDs, as it contains only 82 residues because of a short ZA loop, and cannot be classified into any of the eight typical human BRD families. Taken together, the results obtained from this study provide important insights into human PA200-induced 20S gate opening for substrate degradation and the opportunities to explore the mechanism for its recognition of H4 histone in acetylation-mediated proteasomal degradation.
Asunto(s)
Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Complejo de la Endopetidasa Proteasomal/química , Complejo de la Endopetidasa Proteasomal/metabolismo , Secuencia de Aminoácidos , Microscopía por Crioelectrón , Humanos , Fosfatos de Inositol/metabolismo , Modelos Moleculares , Unión Proteica , Conformación Proteica , Dominios Proteicos , Proteolisis , Relación Estructura-ActividadRESUMEN
Biological motors, ubiquitous in living systems, convert chemical energy into different kinds of mechanical motions critical to cellular functions. Gene product 16 (gp16) in bacteriophage Ï29 is among the most powerful biomotors known, which adopts a multisubunit ring-shaped structure and hydrolyzes ATP to package double-stranded DNA (dsDNA) into a preformed procapsid. Here we report the crystal structure of the C-terminal domain of gp16 (gp16-CTD). Structure-based alignment and molecular dynamics simulations revealed an essential binding surface of gp16-CTD for prohead RNA, a unique component of the motor complex. Furthermore, our simulations highlighted a dynamic interplay between the N-terminal domain and the CTD of gp16, which may play a role in driving movement of DNA into the procapsid. Lastly, we assembled an atomic structural model of the complete Ï29 dsDNA packaging motor complex by integrating structural and experimental data from multiple sources. Collectively, our findings provided a refined inchworm-revolution model for dsDNA translocation in bacteriophage Ï29 and suggested how the individual domains of gp16 work together to power such translocation.
Asunto(s)
Adenosina Trifosfatasas/química , Adenosina Trifosfatasas/metabolismo , Bacteriófagos/genética , Bacteriófagos/metabolismo , Empaquetamiento del ADN , Bacteriófagos/fisiología , ADN Viral/metabolismo , ARN Viral/metabolismo , Ensamble de VirusRESUMEN
Spring viremia of carp virus (SVCV) is a highly pathogenic Vesiculovirus in the common carp. The phosphoprotein (P protein) of SVCV is a multifunctional protein that acts as a polymerase cofactor and an antagonist of cellular interferon (IFN) response. Here, we report the 1.5-Å-resolution crystal structure of the P protein central domain (PCD) of SVCV (SVCVPCD). The PCD monomer consists of two ß sheets, an α helix, and another two ß sheets. Two PCD monomers pack together through their hydrophobic surfaces to form a dimer. The mutations of residues on the hydrophobic surfaces of PCD disrupt the dimer formation to different degrees and affect the expression of host IFN consistently. Therefore, the oligomeric state formation of the P protein of SVCV is an important mechanism to negatively regulate host IFN response.IMPORTANCE SVCV can cause spring viremia of carp with up to 90% lethality, and it is the homologous virus of the notorious vesicular stomatitis virus (VSV). There are currently no drugs that effectively cure this disease. P proteins of negative-strand RNA viruses (NSVs) play an essential role in many steps during the replication cycle and an additional role in immunosuppression as a cofactor. All P proteins of NSVs are oligomeric, but the studies on the role of this oligomerization mainly focus on the process of virus transcription or replication, and there are few studies on the role of PCD in immunosuppression. Here, we present the crystal structure of SVCVPCD A new mechanism of immune evasion is clarified by exploring the relationship between SVCVPCD and host IFN response from a structural biology point of view. These findings may provide more accurate target sites for drug design against SVCV and provide new insights into the function of NSVPCD.
Asunto(s)
Fosfoproteínas/química , Rhabdoviridae/química , Proteínas Virales/química , Animales , Cristalografía por Rayos X , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina betaRESUMEN
Coronaviruses (CoV) have caused a number of major epidemics in humans and animals, including the current pandemic of coronavirus disease 2019 (COVID-19), which has brought a renewed focus on the evolution and interspecies transmission of coronaviruses. Swine acute diarrhea syndrome coronavirus (SADS-CoV), which was recently identified in piglets in southern China, is an alphacoronavirus that originates from the same genus of horseshoe bats as severe acute respiratory syndrome CoV (SARS-CoV) and that was reported to be capable of infecting cells from a broad range of species, suggesting a considerable potential for interspecies transmission. Given the importance of the coronavirus spike (S) glycoprotein in host range determination and viral entry, we report a cryo-electron microscopy (cryo-EM) structure of the SADS-CoV S trimer in the prefusion conformation at a 3.55-Å resolution. Our structure reveals that the SADS-CoV S trimer assumes an intrasubunit quaternary packing mode in which the S1 subunit N-terminal domain (S1-NTD) and the S1 subunit C-terminal domain (S1-CTD) of the same protomer pack together by facing each other in the lying-down state. SADS-CoV S has several distinctive structural features that may facilitate immune escape, such as a relatively compact architecture of the S trimer and epitope masking by glycan shielding. Comparison of SADS-CoV S with the spike proteins of the other coronavirus genera suggested that the structural features of SADS-CoV S are evolutionarily related to those of the spike proteins of the other genera rather than to the spike protein of a typical alphacoronavirus. These data provide new insights into the evolutionary relationship between spike glycoproteins of SADS-CoV and those of other coronaviruses and extend our understanding of their structural and functional diversity.IMPORTANCE In this article, we report the atomic-resolution prefusion structure of the spike protein from swine acute diarrhea syndrome coronavirus (SADS-CoV). SADS-CoV is a pathogenic alphacoronavirus that was responsible for a large-scale outbreak of fatal disease in pigs and that was reported to be capable of interspecies transmission. We describe the overall structure of the SADS-CoV spike protein and conducted a detailed analysis of its main structural elements. Our results and analyses are consistent with those of previous phylogenetic studies and suggest that the SADS-CoV spike protein is evolutionarily related to the spike proteins of betacoronaviruses, with a strong similarity in S1-NTDs and a marked divergence in S1-CTDs. Moreover, we discuss the possible immune evasion strategies used by the SADS-CoV spike protein. Our study provides insights into the structure and immune evasion strategies of the SADS-CoV spike protein and broadens the understanding of the evolutionary relationships between coronavirus spike proteins of different genera.
Asunto(s)
Alphacoronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/ultraestructura , Alphacoronavirus/genética , Secuencia de Aminoácidos , Microscopía por Crioelectrón , Evolución Molecular , Evasión Inmune , Modelos Moleculares , Alineación de Secuencia , Glicoproteína de la Espiga del Coronavirus/química , Homología Estructural de ProteínaRESUMEN
Poliovirus (PV) 2CATPase is the most studied 2C protein in the Picornaviridae family. It is involved in RNA replication, encapsidation and uncoating and many inhibitors have been found that target PV 2CATPase. Despite numerous investigations to characterize its functions, a high-resolution structure of PV 2C has not yet been determined. We report here the crystal structure of a soluble fragment of PV 2CATPase to 2.55Å, containing an ATPase domain, a zinc finger and a C-terminal helical domain but missing the N-terminal domain. The ATPase domain shares the common structural features with EV71 2C and other Superfamily 3 helicases. The C-terminal cysteine-rich motif folds into a CCCC type zinc finger in which four cysteine ligands and several auxiliary residues assist in zinc binding. By comparing with the known zinc finger fold groups, we found the zinc finger of 2C proteins belong to a new fold group, which we denote the "Enterovirus 2C-like" group. The C-terminus of PV 2CATPase forms an amphipathic helix that occupies a hydrophobic pocket located on an adjacent PV 2CATPase in the crystal lattice. The C-terminus mediated PV 2C-2C interaction promotes self-oligomerization, most likely hexamerization, which is fundamental to the ATPase activity of 2C. The zinc finger is the most structurally diverse feature in 2C proteins. Available structural and virological data suggest that the zinc finger of 2C might confer the specificity of interaction with other proteins. We built a hexameric ring model of PV 2CATPase and visualized the previously identified functional motifs and drug-resistant sites, thus providing a structure framework for antiviral drug development.
Asunto(s)
Adenosina Trifosfatasas/química , Poliovirus/enzimología , Proteínas Virales/química , Adenosina Trifosfatasas/genética , Secuencia de Aminoácidos , Animales , Línea Celular , Chlorocebus aethiops , Cristalografía por Rayos X , Humanos , Modelos Moleculares , Fragmentos de Péptidos/química , Fragmentos de Péptidos/genética , Poliovirus/genética , Poliovirus/patogenicidad , Dominios Proteicos , Multimerización de Proteína , Estructura Cuaternaria de Proteína , Homología de Secuencia de Aminoácido , Solubilidad , Electricidad Estática , Células Vero , Proteínas Virales/genéticaRESUMEN
Interferon (IFN) is a vital antiviral factor in host in the early stages after the viral invasion. Meanwhile, viruses have to survive by taking advantage of the cellular machinery and complete their replication. As a result, viruses evolved several immune escape mechanisms to inhibit host IFN expression. However, the mechanisms used to escape the host's IFN system are still unclear for infectious hematopoietic necrosis virus (IHNV). In this study, we report that the N protein of IHNV inhibits IFN1 production in rainbow trout by degrading the MITA. Firstly, the upregulation of IFN1 promoter activity stimulated by poly I:C was suppressed by IHNV infection. Consistent with this result, the overexpression of the N protein of IHNV blocked the IFN1 transcription that was activated by poly I:C and MITA. Secondly, MITA was remarkably decreased by the overexpression of N protein at the protein level. Further analysis demonstrated that the N protein targeted MITA and promoted the ubiquitination of MITA. Taken together, these data suggested that the production of rainbow trout IFN1 could be suppressed by the N protein of IHNV via degrading MITA.
Asunto(s)
Proteínas de Peces/genética , Virus de la Necrosis Hematopoyética Infecciosa/inmunología , Interferones/inmunología , Proteínas de la Membrana/genética , Proteínas de la Nucleocápside/inmunología , Oncorhynchus mykiss/inmunología , Animales , Antivirales/farmacología , Células HEK293 , Interacciones Microbiota-Huesped/inmunología , Humanos , Virus de la Necrosis Hematopoyética Infecciosa/genética , Proteínas de la Nucleocápside/genética , Oncorhynchus mykiss/virología , Poli I-C/farmacología , Infecciones por Rhabdoviridae , UbiquitinaciónRESUMEN
Although the infratentorial superior-lateral cerebellar approach has been traditionally chosen for exposure of the V cranial nerve root in the process of microvascular decompression for treatment of trigeminal neuralgia, those petrosal veins often block this surgical corridor. To detour these petrosal veins, we require a new approach. We provide a via-cerebellar-fissures approach to expose well the trigeminal nerve. With microscopy, cerebrospinal fluid was drained sufficiently to relax the cerebellum. Caudally to petrosal veins, the dissection was started from the cerebellar fissures. With the arachnoid membranes around the petrosal fissure and superior cerebellopontine fissures being opened thoroughly, the root entry zone of V nerve was visualized directly. This new approach was used in 106 patients. Among them, the block veins were encountered in 17 (16.0%). Among the 17 vein-blocked cases, 1 or 2 branches of the veins were finally cut in 2 (1.9%). The postoperative relief rate was 95.3% without complications. This via-cerebellar-fissures approach may access the root entry zone of the V cranial nerve without killing those petrosal veins, which is worth to be recommended and popularized.
Asunto(s)
Cerebelo/cirugía , Cirugía para Descompresión Microvascular/métodos , Procedimientos Neuroquirúrgicos/métodos , Nervio Trigémino/cirugía , Neuralgia del Trigémino/cirugía , Aracnoides/cirugía , Seno Cavernoso/anatomía & histología , Ángulo Pontocerebeloso/anatomía & histología , Cerebelo/irrigación sanguínea , Venas Cerebrales/anatomía & histología , HumanosRESUMEN
Le Chatelier's principle is a basic rule in textbook defining the correlations of reaction activities and specific system parameters (like concentrations), serving as the guideline for regulating chemical/catalytic systems. Here we report a model system breaking this constraint in O2 electroreduction in mixed dioxygen. We unravel the central role of creating single-zinc vacancies in a crystal structure that leads to enzyme-like binding of the catalyst with enhanced selectivity to O2, shifting the reaction pathway from Langmuir-Hinshelwood to an upgraded triple-phase Eley-Rideal mechanism. The model system shows minute activity alteration of H2O2 yields (25.89~24.99 mol gcat-1 h-1) and Faradaic efficiencies (92.5%~89.3%) in the O2 levels of 100%~21% at the current density of 50~300 mA cm-2, which apparently violate macroscopic Le Chatelier's reaction kinetics. A standalone prototype device is built for high-rate H2O2 production from atmospheric air, achieving the highest Faradaic efficiencies of 87.8% at 320 mA cm-2, overtaking the state-of-the-art catalysts and approaching the theoretical limit for direct air electrolysis (~345.8 mA cm-2). Further techno-economics analyses display the use of atmospheric air feedstock affording 21.7% better economics as comparison to high-purity O2, achieving the lowest H2O2 capital cost of 0.3 $ Kg-1. Given the recent surge of demonstrations on tailoring chemical/catalytic systems based on the Le Chatelier's principle, the present finding would have general implications, allowing for leveraging systems "beyond" this classical rule.
RESUMEN
The reversal of ubiquitination induced by members of the SidE effector family of Legionella pneumophila produces phosphoribosyl ubiquitin (PR-Ub) that is potentially detrimental to host cells. Here we show that the effector LnaB functions to transfer the AMP moiety from ATP to the phosphoryl moiety of PR-Ub to convert it into ADP-ribosylated ubiquitin (ADPR-Ub), which is further processed to ADP-ribose and functional ubiquitin by the (ADP-ribosyl)hydrolase MavL, thus maintaining ubiquitin homeostasis in infected cells. Upon being activated by Actin, LnaB also undergoes self-AMPylation on tyrosine residues. The activity of LnaB requires a motif consisting of Ser, His and Glu (S-HxxxE) present in a large family of toxins from diverse bacterial pathogens. Our study not only reveals intricate mechanisms for a pathogen to maintain ubiquitin homeostasis but also identifies a new family of enzymes capable of protein AMPylation, suggesting that this posttranslational modification is widely used in signaling during host-pathogen interactions.
RESUMEN
The intracellular bacterial pathogen Legionella pneumophila modulates host cell functions by secreting multiple effectors with diverse biochemical activities. In particular, effectors of the SidE family interfere with host protein ubiquitination in a process that involves production of phosphoribosyl ubiquitin (PR-Ub). Here, we show that effector LnaB converts PR-Ub into ADP-ribosylated ubiquitin, which is further processed to ADP-ribose and functional ubiquitin by the (ADP-ribosyl)hydrolase MavL, thus maintaining ubiquitin homeostasis in infected cells. Upon being activated by actin, LnaB also undergoes self-AMPylation on tyrosine residues. The activity of LnaB requires a motif consisting of Ser, His and Glu (SHxxxE) present in a large family of toxins from diverse bacterial pathogens. Thus, our study sheds light on the mechanisms by which a pathogen maintains ubiquitin homeostasis and identifies a family of enzymes capable of protein AMPylation.
Asunto(s)
Proteínas Bacterianas , Homeostasis , Legionella pneumophila , Ubiquitina , Ubiquitinación , Ubiquitina/metabolismo , Legionella pneumophila/metabolismo , Legionella pneumophila/patogenicidad , Humanos , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/genética , ADP-Ribosilación , Interacciones Huésped-Patógeno , Adenosina Difosfato Ribosa/metabolismo , Enfermedad de los Legionarios/metabolismo , Enfermedad de los Legionarios/microbiología , Células HEK293 , Actinas/metabolismo , Células HeLaRESUMEN
BACKGROUND: Due to its anatomical features, the vertebrobasilar artery complex (VBA) seldom contributes to the neurovascular conflict in patient with trigeminal neuralgia (TN). However, once it offends the trigeminal root, this large artery is really difficult to manipulate during microvascular decompression (MVD) surgery. Therefore, the surgical strategy for such cases needs to be detailed in order to obtain a satisfactory outcome. METHODS: From 2009 through 2011, 475 consecutive TN patients underwent MVDs in our department. Among them, ten were found in which an ipsilateral deviating ectatic vertebrobasilar artery complex (VBA) offended the trigeminal nerve. Those cases were focused on in this study and each operation was analyzed retrospectively. RESULTS: During the operation, the vertebral artery was regarded as the direct culprit in six (60 %) patients, while the basilar artery in four (40 %). As companions, some smaller vessels were also observed to be close to the nerve, including the superior cerebellar artery (SCA) in five, veins in two and anterior inferior cerebellar artery (AICA) in two. The neurovascular conflict was discovered in the cisternal segment of the trigeminal root in eight, while in the root entry zone (REZ) in two. In six out of the ten cases, the affected nerves were demonstrated to be squeezed towards the tentorium by the ectatic VBA. Postoperatively, the symptom of pain totally disappeared immediately in eight (80 %) patients, while it was relieved apparently in two (20 %). During the follow-up period of 3-30 months, no recurrence or complication was found, except for one patient who had numbness of the face. CONCLUSION: With a proper strategy, MVD is probably the most effective therapy for the TN cases caused by ectatic vertebrobasilar artery complex. The substance of the surgery is to withdraw the proximal vertebral artery caudally via a lateroinferior cerebellar approach.
Asunto(s)
Cirugía para Descompresión Microvascular/métodos , Neuralgia del Trigémino/cirugía , Anciano , Anciano de 80 o más Años , Arteria Basilar/cirugía , Femenino , Estudios de Seguimiento , Humanos , Masculino , Cirugía para Descompresión Microvascular/efectos adversos , Persona de Mediana Edad , Resultado del Tratamiento , Neuralgia del Trigémino/etiología , Arteria Vertebral/cirugíaRESUMEN
OBJECTIVE: Although microvascular decompression (MVD) has been accepted as effective therapy for hemifacial spasm, failed surgery has been reported frequently. For a sophisticated neurosurgeon, an apparent offending artery is seldom missed. However, it is still an embarrassed situation when the neurovascular conflict site could not be approached. METHODS: Clinical data were collected from consecutive 211 MVDs in 2010. Intraoperative abnormal muscle response was recorded. Among them, the neurovascular conflict was not finally discovered in 3 patients, whom were then focused on. All patients were followed up for 6 to 15 months. RESULTS: In 17 of the 211 MVDs, the cerebellum was hard to be retracted because of adhesions. After careful dissection, a working space was finally created in the cerebellopontine angle. However, there still were 3 cases, whose neurovascular conflict site was unable to be discovered at last because of a branch of an artery embedded in the petrous bone and made the cerebellum unmovable. With navigation of real-time abnormal muscle response, the offending artery was moved away eventually even without exposing the conflict site. Postoperatively, all the patients were completely spasm-free immediately. No recurrence was noticed in the last follow-up period. CONCLUSIONS: The most important thing for a successful MVD operation is to remove the offending artery off the nerve. However, if the conflict site failed to be approached after endeavors, a successful MVD can still be achieved by relocating the offending artery with the guidance of real-time electromyography even without visualization of the confliction.
Asunto(s)
Espasmo Hemifacial/cirugía , Cirugía para Descompresión Microvascular/métodos , Ángulo Pontocerebeloso/cirugía , Electromiografía , Músculos Faciales/irrigación sanguínea , Músculos Faciales/cirugía , Femenino , Espasmo Hemifacial/fisiopatología , Humanos , Masculino , Persona de Mediana Edad , Monitoreo Intraoperatorio , Resultado del TratamientoRESUMEN
The mitochondrion is an important signaling hub that governs diverse cellular functions, including metabolism, energy production, and immunity. Among the hundreds of effectors translocated into host cells by the Dot/Icm system of Legionella pneumophila, several are targeted to mitochondria but the function of most of them remains elusive. Our recent study found that the effector Ceg3 inhibits the activity of ADP/ATP translocases (ANTs) by ADP-ribosylation (ADPR). Here, we show that the effect of Ceg3 is antagonized by Larg1, an effector encoded by lpg0081, a gene that is situated next to ceg3. Larg1 functions to reverse Ceg3-mediated ADPR of ANTs by cleaving the N-glycosidic bond between the ADPR moiety and the modified arginine residues in ANTs, leading to restoration of their activity in ADP/ATP exchange. Structural analysis of Larg1 and its complex with ADPR reveals that this ADPR glycohydrolase harbors a unique macrodomain that catalyzes the removal of ADPR modification on ANTs. Our results also demonstrate that together with Ceg3, Larg1 imposes temporal regulation of the activity of ANTs by reversible ADPR during L. pneumophila infection.
RESUMEN
The preparation of highly efficient, stable, and low-cost electrocatalysts for the oxygen evolution reaction (OER) and the hydrogen evolution reaction (HER) is still a challenge for the development of new energy systems. In this work, a NiCo bimetal loaded on porous carbon (NiCo-C/NF) grown on nickel foam (NF) was obtained via the pyrolysis of a NiCo bimetal MOF (NiCo-MOF/NF) under a nitrogen atmosphere at 500 °C. Compared with NiCo-MOF/NF, NiCo-C/NF had a larger specific surface and uniform mesoporous structure. As an electrocatalyst in the OER, this new type of electrode operated with better stability in an alkaline solution (1.0 mol L-1 KOH), the overpotential when the current density reached 10 mA cm-2 was only 260 mV, and the electrode also exhibited long-term durability in a stability test for 10 h without significant changes. The excellent activity and stability toward the OER can be attributed to the synergistic effect of the NiCo bimetal and the abundant active sites exposed after the carbonization of NiCo-MOF, which compensated for the defect of the insufficient conductivity of the material and promoted the evolution of oxygen in the catalytic process.
RESUMEN
The proliferation and migration of vascular smooth muscle cells (VSMCs) are involved in the pathogenesis of intracranial aneurysm (IA) formation and rupture. Interleukin enhancer binding factor 2 (ILF2) is known as the nuclear factor of activated T cells and regulates cell growth. This study was aimed to explore the effects of ILF2 on IA progression. Human brain VSMCs (hBVSMCs) were transfected with pCDNA3.1(+), pCDNA3.1(+)-ILF2, siRNA-negative control, and siRNA-ILF2. The transfection efficiency was then evaluated by determining ILF2 expression. The cell viability and apoptosis were determined using Cell Counting Kit-8 and Annexin V-FITC cell apoptosis assay kit, respectively. Real-time quantification PCR (RT-qPCR) was applied to measure the expression levels of apoptosis-related and inflammation-related genes. Finally, western blot was used to detect the expression level of Fas cell surface death receptor 95 (CD95) and Caspase 8. Overexpression of ILF2 could significantly increase cell viability and decrease cell apoptosis (P < 0.05), while knock-down of ILF2 showed opposite trends for hBVSMCs on cell viability and apoptosis (P < 0.05). RT-qPCR results showed that ILF2 knock-down downregulated the expression levels of BCL2 apoptosis regulator (BCL2), transcriptional regulator Myc-like (c-Myc), and caspase 1 (ICE) whereas upregulated the expression levels of CD95, p21, p53, and interleukin-13 (IL-13). Additionally, the protein expression levels of CD95 and Caspase 8 were significantly decreased after ILF2 overexpression while were significantly increased after ILF2 knock-down (P < 0.05). ILF2 knock-down may inhibit cell viability and promote cell apoptosis of hBVSMCs by regulating the expression levels of apoptosis-related genes and suppressing inflammatory response.