Publications by authors named "Songying Ouyang"

67 Publications

A non-canonical cGAS-STING-PERK pathway facilitates the translational program critical for senescence and organ fibrosis.

Nat Cell Biol 2022 May 2;24(5):766-782. Epub 2022 May 2.

The MOE Key Laboratory of Biosystems Homeostasis & Protection and Zhejiang Provincial Key Laboratory of Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou, China.

Innate DNA sensing via the cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) mechanism surveys microbial invasion and cellular damage and thus participates in various human infectious diseases, autoimmune diseases and cancers. However, how DNA sensing rapidly and adaptively shapes cellular physiology is incompletely known. Here we identify the STING-PKR-like endoplasmic reticulum kinase (PERK)-eIF2α pathway, a previously unknown cGAS-STING mechanism, enabling an innate immunity control of cap-dependent messenger RNA translation. Upon cGAMP binding, STING at the ER binds and directly activates the ER-located kinase PERK via their intracellular domains, which precedes TBK1-IRF3 activation and is irrelevant to the unfolded protein response. The activated PERK phosphorylates eIF2α, forming an inflammatory- and survival-preferred translation program. Notably, this STING-PERK-eIF2α pathway is evolutionarily primitive and physiologically critical to cellular senescence and organ fibrosis. Pharmacologically or genetically targeting this non-canonical cGAS-STING pathway attenuated lung and kidney fibrosis. Collectively, the findings identify an alternative innate immune pathway and its critical role in organ fibrosis, report an innate immunity-directed translation program and suggest the therapeutic potential for targeting the STING-PERK pathway in treating fibrotic diseases.
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http://dx.doi.org/10.1038/s41556-022-00894-zDOI Listing
May 2022

Metabolome and Whole-Transcriptome Analyses Reveal the Molecular Mechanisms Underlying Hypoglycemic Nutrient Metabolites Biosynthesis in Leaves During Different Harvest Stages.

Front Nutr 2022 28;9:851569. Epub 2022 Feb 28.

The Public Service Platform for Industrialization Development Technology of Marine Biological Medicine and Products of the State Oceanic Administration, Fujian Key Laboratory of Special Marine Bioresource Sustainable Utilization, Southern Institute of Oceanography, Key Laboratory of Developmental and Neural Biology, College of Life Sciences, Fujian Normal University, Fuzhou, China.

, a well-known nutrient and beverage plant, is under development for use in functional health care products best and natural and organic foods. We hypothesis that the composition and metabolic accumulation of hypoglycemic nutrient metabolites exhibit significant differences depending on harvest time. Therefore, it is of great significance to establish the best harvest time for leaves for the further development of healthy teas and other products. However, the detail compositions and molecular mechanisms of nutrients biosynthesis in leaves during different harvest stages remain largely unclear. Metabolome analysis showed that a suitable leaf-harvesting strategy for could be in September or October each year due to the high content of hypoglycemic nutrient metabolites. We found that two of the seven differentially accumulated phenolic acid metabolites have a relatively good inhibitory effect on α-amylase, indicating that they may play a role in the hypoglycemic function. Combined analysis of coexpression, ceRNA network, and weighted gene correlation network analysis (WGCNA) showed that several genes or transcription factors (TFs) in three modules correlated highly with hypoglycemic nutrient metabolites, including , and . These findings help in the understanding of the molecular mechanisms and regulatory networks of the hypoglycemic nutrient metabolites in leaves which are dependent on harvest time and provide theoretical guidance in the development of functional health care products and foods from .
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http://dx.doi.org/10.3389/fnut.2022.851569DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8919051PMC
February 2022

Rifapentine is an entry and replication inhibitor against yellow fever virus both in vitro and in vivo.

Emerg Microbes Infect 2022 Dec;11(1):873-884

Department of Microbiology, Faculty of Naval Medicine, Naval Medical University, Shanghai, People's Republic of China.

Yellow fever virus (YFV) infection is a major public concern that threatens a large population in South America and Africa. No specific anti-YFV drugs are available till now. Here, we report that rifapentine is a potent YFV inhibitor in various cell lines by high-throughput drugs screening, acting at both cell entry and replication steps. Kinetic test and binding assay suggest that rifapentine interferes the viral attachment to the target cells. The application of YFV replicon and surface plasmon resonance assay indicates that rifapentine suppresses viral replication by binding to the RNA-dependent RNA polymerase (RdRp) domain of viral nonstructural protein NS5. Further molecular docking suggests that it might interact with the active centre of RdRp. Rifapentine significantly improves the survival rate, alleviates clinical signs, and reduces virus load and injury in targeted organs both in YFV-infected type I interferon receptor knockout A129 and wild-type C57 mice. The antiviral effect in vivo is robust during both prophylactic intervention and therapeutic treatment, and the activity is superior to sofosbuvir, a previously reported YFV inhibitor in mice. Our data show that rifapentine may serve as an effective anti-YFV agent, providing promising prospects in the development of YFV pharmacotherapy.
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http://dx.doi.org/10.1080/22221751.2022.2049983DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8942558PMC
December 2022

Induced phase separation of mutant NF2 imprisons the cGAS-STING machinery to abrogate antitumor immunity.

Mol Cell 2021 10 27;81(20):4147-4164.e7. Epub 2021 Aug 27.

MOE Laboratory of Biosystems Homeostasis & Protection and Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou 310058, China; Department of Hepatobiliary and Pancreatic Surgery and Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310058, China; ZJU-Hangzhou Global Scientific and Technological Innovation Center (HIC-ZJU), Hangzhou 310058, China; Cancer Center, Zhejiang University, Hangzhou 310058, China. Electronic address:

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.
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http://dx.doi.org/10.1016/j.molcel.2021.07.040DOI Listing
October 2021

Functional Features and Current Applications of the RNA-Targeting Type VI CRISPR-Cas Systems.

Adv Sci (Weinh) 2021 07 5;8(13):2004685. Epub 2021 May 5.

The Key Laboratory of Innate Immune Biology of Fujian Province Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation Biomedical Research Center of South China Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education College of Life Sciences Fujian Normal University Fuzhou 350117 China.

CRISPR-Cas systems are a form of prokaryotic adaptive immunity that employs RNA-guided endonucleases (Cas effectors) to cleave foreign genetic elements. Due to their simplicity, targeting programmability, and efficiency, single-effector CRISPR-Cas systems have great potential for application in research, biotechnology, and therapeutics. While DNA-targeting Cas effectors such as Cas9 and Cas12a have become indispensable tools for genome editing in the past decade, the more recent discovery of RNA-targeting CRISPR-Cas systems has opened the door for implementation of CRISPR-Cas technology in RNA manipulation. With an increasing number of studies reporting their application in transcriptome engineering, viral interference, nucleic acid detection, and RNA imaging, type VI CRISPR-Cas systems and the associated Cas13 effectors particularly hold promise as RNA-targeting or RNA-binding tools. However, even though previous structural and biochemical characterization provided a firm basis for leveraging type VI CRISPR-Cas systems into such tools, the lack of comprehension of certain mechanisms underlying their functions hinders more sophisticated and conventional use. This review will summarize current knowledge on structural and mechanistic properties of type VI CRISPR-Cas systems, give an overview on the reported applications, and discuss functional features that need further investigation in order to improve performance of Cas13-based tools.
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http://dx.doi.org/10.1002/advs.202004685DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8209922PMC
July 2021

Mechanistic insights into the R-loop formation and cleavage in CRISPR-Cas12i1.

Nat Commun 2021 06 9;12(1):3476. Epub 2021 Jun 9.

The Key Laboratory of Innate Immune Biology of Fujian Province, Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou, China.

Cas12i is a newly identified member of the functionally diverse type V CRISPR-Cas effectors. Although Cas12i has the potential to serve as genome-editing tool, its structural and functional characteristics need to be investigated in more detail before effective application. Here we report the crystal structures of the Cas12i1 R-loop complexes before and after target DNA cleavage to elucidate the mechanisms underlying target DNA duplex unwinding, R-loop formation and cis cleavage. The structure of the R-loop complex after target DNA cleavage also provides information regarding trans cleavage. Besides, we report a crystal structure of the Cas12i1 binary complex interacting with a pseudo target oligonucleotide, which mimics target interrogation. Upon target DNA duplex binding, the Cas12i1 PAM-interacting cleft undergoes a remarkable open-to-closed adjustment. Notably, a zipper motif in the Helical-I domain facilitates unzipping of the target DNA duplex. Formation of the 19-bp crRNA-target DNA strand heteroduplex in the R-loop complexes triggers a conformational rearrangement and unleashes the DNase activity. This study provides valuable insights for developing Cas12i1 into a reliable genome-editing tool.
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http://dx.doi.org/10.1038/s41467-021-23876-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8190297PMC
June 2021

Biochemical and structural characterization of the BioZ enzyme engaged in bacterial biotin synthesis pathway.

Nat Commun 2021 04 6;12(1):2056. Epub 2021 Apr 6.

Department of Pathogen Biology & Microbiology and General Intensive Care Unit of Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, China.

Biotin is an essential micro-nutrient across the three domains of life. The paradigm earlier step of biotin synthesis denotes "BioC-BioH" pathway in Escherichia coli. Here we report that BioZ bypasses the canonical route to begin biotin synthesis. In addition to its origin of Rhizobiales, protein phylogeny infers that BioZ is domesticated to gain an atypical role of β-ketoacyl-ACP synthase III. Genetic and biochemical characterization demonstrates that BioZ catalyzes the condensation of glutaryl-CoA (or ACP) with malonyl-ACP to give 5'-keto-pimeloyl ACP. This intermediate proceeds via type II fatty acid synthesis (FAS II) pathway, to initiate the formation of pimeloyl-ACP, a precursor of biotin synthesis. To further explore molecular basis of BioZ activity, we determine the crystal structure of Agrobacterium tumefaciens BioZ at 1.99 Å, of which the catalytic triad and the substrate-loading tunnel are functionally defined. In particular, we localize that three residues (S84, R147, and S287) at the distant bottom of the tunnel might neutralize the charge of free C-carboxyl group of the primer glutaryl-CoA. Taken together, this study provides molecular insights into the BioZ biotin synthesis pathway.
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http://dx.doi.org/10.1038/s41467-021-22360-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8024396PMC
April 2021

Structural Insights into gp16 ATPase in the Bacteriophage ϕ29 DNA Packaging Motor.

Biochemistry 2021 03 9;60(11):886-897. Epub 2021 Mar 9.

The Key Laboratory of Innate Immune Biology of Fujian Province, Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou 350117, China.

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 ϕ29 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 ϕ29 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 ϕ29 and suggested how the individual domains of gp16 work together to power such translocation.
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http://dx.doi.org/10.1021/acs.biochem.0c00935DOI Listing
March 2021

Structural and biochemical analyses of the tetrameric cell binding domain of Lys170 from enterococcal phage F170/08.

Eur Biophys J 2021 Jul 20;50(5):721-729. Epub 2021 Feb 20.

The Key Laboratory of Innate Immune Biology of Fujian Province, Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou, China.

Lysins are a class of hydrolytic enzymes used by bacteriophages to target and cleave the peptidoglycan of bacterial cell walls during their lytic cycle. The lysins from bacteriophages that infect Gram-positive bacteria are typically monomeric and consist of one or two catalytic domains (CD) and a cell binding domain (CBD). However, multimeric lysins encoded by a single gene have also been reported, among which Lys170 from enterococcal phage F170/08 was one of the first identified. Here, we determined the crystal structure of Lys170 CBD at 1.40 Å resolution. The structure reveals that Lys170 CBDs assemble into a tetrameric functional unit and that each monomer folds into a three-stranded β-sheet core capped on each side by an α-helix. In addition, we identified key residues of Lys170 CBD involved in host cell binding. Our work provides a basis for designing highly efficient lysins targeting Enterococcus faecalis.
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http://dx.doi.org/10.1007/s00249-021-01511-xDOI Listing
July 2021

Marine-derived drugs: Recent advances in cancer therapy and immune signaling.

Biomed Pharmacother 2021 Feb 16;134:111091. Epub 2020 Dec 16.

The Key Laboratory of Innate Immune Biology of Fujian Province, Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou, 350117, China; Laboratory for Marine Biology and Biotechnology, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao, 266237, China. Electronic address:

The marine environment is an enormous source of marine-derived natural products (MNPs), and future investigation into anticancer drug discovery. Current progress in anticancer drugs offers a rise in isolation and clinical validation of numerous innovative developments and advances in anticancer therapy. However, only a limited number of FDA-approved marine-derived anticancer drugs are available due to several challenges and limitations highlighted here. The use of chitosan in developing marine-derived drugs is promising in the nanotech sector projected for a prolific anticancer drug delivery system (DDS). The cGAS-STING-mediated immune signaling pathway is crucial, which has not been significantly investigated in anticancer therapy and needs further attention. Additionally, a small range of anticancer mediators is currently involved in regulating various JAK/STAT signaling pathways, such as immunity, cell death, and tumor formation. This review addressed critical features associated with MNPs, origin, and development of anticancer drugs. Moreover, recent advances in the nanotech delivery of anticancer drugs and understanding into cancer immunity are detailed for improved human health.
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http://dx.doi.org/10.1016/j.biopha.2020.111091DOI Listing
February 2021

TBK1-Mediated DRP1 Targeting Confers Nucleic Acid Sensing to Reprogram Mitochondrial Dynamics and Physiology.

Mol Cell 2020 12 9;80(5):810-827.e7. Epub 2020 Nov 9.

MOE Laboratory of Biosystems Homeostasis & Protection and Innovation Center for Cell Signaling Network, Life Sciences Institute, Zhejiang University, Hangzhou 310058, China; Department of Hepatobiliary and Pancreatic Surgery and Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310058, China. Electronic address:

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.
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http://dx.doi.org/10.1016/j.molcel.2020.10.018DOI Listing
December 2020

Interplay between bacterial deubiquitinase and ubiquitin E3 ligase regulates ubiquitin dynamics on Legionella phagosomes.

Elife 2020 11 2;9. Epub 2020 Nov 2.

Department of Respiratory Medicine and Center of Infection and Immunity, Key Laboratory of Organ Regeneration and Transplantation of the Ministry of Education, The First Hospital, Jilin University, Changchun, China.

extensively modulates the host ubiquitin network to create the Legionella-containing vacuole (LCV) for its replication. Many of its virulence factors function as ubiquitin ligases or deubiquitinases (DUBs). Here, we identify Lem27 as a DUB that displays a preference for diubiquitin formed by K6, K11, or K48. Lem27 is associated with the LCV where it regulates Rab10 ubiquitination in concert with SidC and SdcA, two bacterial E3 ubiquitin ligases. Structural analysis of the complex formed by an active fragment of Lem27 and the substrate-based suicide inhibitor ubiquitin-propargylamide (PA) reveals that it harbors a fold resembling those in the OTU1 DUB subfamily with a Cys-His catalytic dyad and that it recognizes ubiquitin via extensive hydrogen bonding at six contact sites. Our results establish Lem27 as a DUB that functions to regulate protein ubiquitination on phagosomes by counteracting the activity of bacterial ubiquitin E3 ligases.
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http://dx.doi.org/10.7554/eLife.58114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7669269PMC
November 2020

Novel polyadenylylation-dependent neutralization mechanism of the HEPN/MNT toxin/antitoxin system.

Nucleic Acids Res 2020 11;48(19):11054-11067

Key Laboratory of Tropical Marine Bio-resources and Ecology, Guangdong Key Laboratory of Marine Materia Medica, Innovation Academy of South China Sea Ecology and Environmental Engineering, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.

The two-gene module HEPN/MNT is predicted to be the most abundant toxin/antitoxin (TA) system in prokaryotes. However, its physiological function and neutralization mechanism remains obscure. Here, we discovered that the MntA antitoxin (MNT-domain protein) acts as an adenylyltransferase and chemically modifies the HepT toxin (HEPN-domain protein) to block its toxicity as an RNase. Biochemical and structural studies revealed that MntA mediates the transfer of three AMPs to a tyrosine residue next to the RNase domain of HepT in Shewanella oneidensis. Furthermore, in vitro enzymatic assays showed that the three AMPs are transferred to HepT by MntA consecutively with ATP serving as the substrate, and this polyadenylylation is crucial for reducing HepT toxicity. Additionally, the GSX10DXD motif, which is conserved among MntA proteins, is the key active motif for polyadenylylating and neutralizing HepT. Thus, HepT/MntA represents a new type of TA system, and the polyadenylylation-dependent TA neutralization mechanism is prevalent in bacteria and archaea.
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http://dx.doi.org/10.1093/nar/gkaa855DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641770PMC
November 2020

Cryo-electron Microscopy Structure of the Swine Acute Diarrhea Syndrome Coronavirus Spike Glycoprotein Provides Insights into Evolution of Unique Coronavirus Spike Proteins.

J Virol 2020 10 27;94(22). Epub 2020 Oct 27.

Key Laboratory of Innate Immune Biology of Fujian Province, Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou, China

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. 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.
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http://dx.doi.org/10.1128/JVI.01301-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7592232PMC
October 2020

Evolutionary Arms Race between Virus and Host Drives Genetic Diversity in Bat Severe Acute Respiratory Syndrome-Related Coronavirus Spike Genes.

J Virol 2020 09 29;94(20). Epub 2020 Sep 29.

CAS Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China

The Chinese horseshoe bat (), reservoir host of severe acute respiratory syndrome coronavirus (SARS-CoV), carries many bat SARS-related CoVs (SARSr-CoVs) with high genetic diversity, particularly in the spike gene. Despite these variations, some bat SARSr-CoVs can utilize the orthologs of the human SARS-CoV receptor, angiotensin-converting enzyme 2 (ACE2), for entry. It is speculated that the interaction between bat ACE2 and SARSr-CoV spike proteins drives diversity. Here, we identified a series of ACE2 variants with some polymorphic sites involved in the interaction with the SARS-CoV spike protein. Pseudoviruses or SARSr-CoVs carrying different spike proteins showed different infection efficiencies in cells transiently expressing bat ACE2 variants. Consistent results were observed by binding affinity assays between SARS-CoV and SARSr-CoV spike proteins and receptor molecules from bats and humans. All tested bat SARSr-CoV spike proteins had a higher binding affinity to human ACE2 than to bat ACE2, although they showed a 10-fold lower binding affinity to human ACE2 compared with that of their SARS-CoV counterpart. Structure modeling revealed that the difference in binding affinity between spike and ACE2 might be caused by the alteration of some key residues in the interface of these two molecules. Molecular evolution analysis indicates that some key residues were under positive selection. These results suggest that the SARSr-CoV spike protein and ACE2 may have coevolved over time and experienced selection pressure from each other, triggering the evolutionary arms race dynamics. Evolutionary arms race dynamics shape the diversity of viruses and their receptors. Identification of key residues which are involved in interspecies transmission is important to predict potential pathogen spillover from wildlife to humans. Previously, we have identified genetically diverse SARSr-CoVs in Chinese horseshoe bats. Here, we show the highly polymorphic ACE2 in Chinese horseshoe bat populations. These ACE2 variants support SARS-CoV and SARSr-CoV infection but with different binding affinities to different spike proteins. The higher binding affinity of SARSr-CoV spike to human ACE2 suggests that these viruses have the capacity for spillover to humans. The positive selection of residues at the interface between ACE2 and SARSr-CoV spike protein suggests long-term and ongoing coevolutionary dynamics between them. Continued surveillance of this group of viruses in bats is necessary for the prevention of the next SARS-like disease.
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http://dx.doi.org/10.1128/JVI.00902-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7527062PMC
September 2020

Molecular Basis of Ubiquitination Catalyzed by the Bacterial Transglutaminase MavC.

Adv Sci (Weinh) 2020 Jun 30;7(12):2000871. Epub 2020 Apr 30.

The Key Laboratory of Innate Immune Biology of Fujian Province Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation Biomedical Research Center of South China Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education College of Life Sciences Fujian Normal University Fuzhou 350117 China.

The effector MavC is a transglutaminase that carries out atypical ubiquitination of the host ubiquitin (Ub)-conjugation enzyme UBE2N by catalyzing the formation of an isopeptide bond between Gln40 and Lys92, which leads to inhibition of signaling in the NF-κB pathway. In the absence of UBE2N, MavC deamidates Ub at Gln40 or catalyzes self-ubiquitination. However, the mechanisms underlying these enzymatic activities of MavC are poorly understood at the molecular level. This study reports the structure of the MavC-UBE2N-Ub ternary complex representing a snapshot of MavC-catalyzed crosslinking of UBE2N and Ub, which reveals the way by which UBE2N-Ub binds to the Insertion and Tail domains of MavC. Based on the structural and experimental data, the catalytic mechanism for the deamidase and transglutaminase activities of MavC is proposed. Finally, by comparing the structures of MavC and MvcA, the homologous protein that reverses MavC-induced UBE2N ubiquitination, several essential regions and two key residues (Trp255 and Phe268) responsible for their respective enzymatic activities are identified. The results provide insights into the mechanisms for substrate recognition and ubiquitination mediated by MavC as well as explanations for the opposite activity of MavC and MvcA in terms of regulation of UBE2N ubiquitination.
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http://dx.doi.org/10.1002/advs.202000871DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7312448PMC
June 2020

Structural and Functional Characterization of the Phosphoprotein Central Domain of Spring Viremia of Carp Virus.

J Virol 2020 07 16;94(15). Epub 2020 Jul 16.

State Key Laboratory of Agricultural Microbiology, College of Fisheries, Huazhong Agricultural University, Wuhan, China

Spring viremia of carp virus (SVCV) is a highly pathogenic 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 (P) of SVCV (SVCV). The P monomer consists of two β sheets, an α helix, and another two β sheets. Two P monomers pack together through their hydrophobic surfaces to form a dimer. The mutations of residues on the hydrophobic surfaces of P 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. 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 P in immunosuppression. Here, we present the crystal structure of SVCV A new mechanism of immune evasion is clarified by exploring the relationship between SVCV 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 NSV.
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http://dx.doi.org/10.1128/JVI.00855-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7375382PMC
July 2020

Regulation of cGAS-Mediated Immune Responses and Immunotherapy.

Adv Sci (Weinh) 2020 Mar 6;7(6):1902599. Epub 2020 Feb 6.

The Key Laboratory of Innate Immune Biology of Fujian Province Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation Biomedical Research Center of South China Key Laboratory of Optoelectronic Science and Technology for Medicine of Ministry of Education College of Life Sciences Fujian Normal University Fuzhou 350117 China.

Early detection of infectious nucleic acids released from invading pathogens by the innate immune system is critical for immune defense. Detection of these nucleic acids by host immune sensors and regulation of DNA sensing pathways have been significant interests in the past years. Here, current understandings of evolutionarily conserved DNA sensing cyclic GMP-AMP (cGAMP) synthase (cGAS) are highlighted. Precise activation and tight regulation of cGAS are vital in appropriate innate immune responses, senescence, tumorigenesis and immunotherapy, and autoimmunity. Hence, substantial insights into cytosolic DNA sensing and immunotherapy of indispensable cytosolic sensors have been detailed to extend limited knowledge available thus far. This Review offers a critical, in-depth understanding of cGAS regulation, cytosolic DNA sensing, and currently established therapeutic approaches of essential cytosolic immune agents for improved human health.
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http://dx.doi.org/10.1002/advs.201902599DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7080523PMC
March 2020

Structural and functional insights into a novel two-component endolysin encoded by a single gene in Enterococcus faecalis phage.

PLoS Pathog 2020 03 16;16(3):e1008394. Epub 2020 Mar 16.

The Key Laboratory of Innate Immune Biology of Fujian Province, Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou, China.

Using bacteriophage-derived endolysins as an alternative strategy for fighting drug-resistant bacteria has recently been garnering renewed interest. However, their application is still hindered by their narrow spectra of activity. In our previous work, we demonstrated that the endolysin LysIME-EF1 possesses efficient bactericidal activity against multiple strains of Enterococcus faecalis (E. faecalis). Herein, we observed an 8 kDa fragment and hypothesized that it contributes to LysIME-EF1 lytic activity. To examine our hypothesis, we determined the structure of LysIME-EF1 at 1.75 Å resolution. LysIME-EF1 exhibits a unique architecture in which one full-length LysIME-EF1 forms a tetramer with three additional C-terminal cell-wall binding domains (CBDs) that correspond to the abovementioned 8 kDa fragment. Furthermore, we identified an internal ribosomal binding site (RBS) and alternative start codon within LysIME-EF1 gene, which are demonstrated to be responsible for the translation of the truncated CBD. To elucidate the molecular mechanism for the lytic activity of LysIME-EF1, we combined mutagenesis, lytic activity assays and in vivo animal infection experiments. The results confirmed that the additional LysIME-EF1 CBDs are important for LysIME-EF1 architecture and its lytic activity. To our knowledge, this is the first determined structure of multimeric endolysin encoded by a single gene in E. faecalis phages. As such, it may provide valuable insights into designing potent endolysins against the opportunistic pathogen E. faecalis.
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http://dx.doi.org/10.1371/journal.ppat.1008394DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7098653PMC
March 2020

NOD1 Promotes Antiviral Signaling by Binding Viral RNA and Regulating the Interaction of MDA5 and MAVS.

J Immunol 2020 04 13;204(8):2216-2231. Epub 2020 Mar 13.

State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, Hubei Province, China;

Nucleotide oligomerization domain-like receptors (NLRs) and RIG-I-like receptors (RLRs) detect diverse pathogen-associated molecular patterns to activate the innate immune response. The role of mammalian NLR NOD1 in sensing bacteria is well established. Although several studies suggest NOD1 also plays a role in sensing viruses, the mechanisms behind this are still largely unknown. In this study, we report on the synergism and antagonism between NOD1 and MDA5 isoforms in teleost. In zebrafish, the overexpression of NOD1 enhances the antiviral response and mRNA abundances of key antiviral genes involved in RLR-mediated signaling, whereas the loss of NOD1 has the opposite effect. Notably, spring viremia of carp virus-infected zebrafish exhibit reduced survival compared with wild-type counterparts. Mechanistically, NOD1 targets MDA5 isoforms and TRAF3 to modulate the formation of MDA5-MAVS and TRAF3-MAVS complexes. The cumulative effects of NOD1 and MDA5a (MDA5 normal form) were observed for the binding with poly(I:C) and the formation of the MDA5a-MAVS complex, which led to increased transcription of type I IFNs and ISGs. However, the antagonism between NOD1 and MDA5b (MDA5 truncated form) was clearly observed during proteasomal degradation of NOD1 by MDA5b. In humans, the interactions between NOD1-MDA5 and NOD1-TRAF3 were confirmed. Furthermore, the roles that NOD1 plays in enhancing the binding of MDA5 to MAVS and poly(I:C) are also evolutionarily conserved across species. Taken together, our findings suggest that mutual regulation between NOD1 and MDA5 isoforms may play a crucial role in the innate immune response and that NOD1 acts as a positive regulator of MDA5/MAVS normal form-mediated immune signaling in vertebrates.
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http://dx.doi.org/10.4049/jimmunol.1900667DOI Listing
April 2020

Cryo-EM structures of the human PA200 and PA200-20S complex reveal regulation of proteasome gate opening and two PA200 apertures.

PLoS Biol 2020 03 5;18(3):e3000654. Epub 2020 Mar 5.

The Key Laboratory of Innate Immune Biology of Fujian Province, Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou, China.

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.
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http://dx.doi.org/10.1371/journal.pbio.3000654DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7077846PMC
March 2020

The bacterial deubiquitinase Ceg23 regulates the association of Lys-63-linked polyubiquitin molecules on the phagosome.

J Biol Chem 2020 02 6;295(6):1646-1657. Epub 2020 Jan 6.

Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, Changchun, China. Electronic address:

is the causative agent of the lung malady Legionnaires' disease, it modulates host function to create a niche termed the -containing vacuole (LCV) that permits intracellular replication. One important aspect of such modulation is the co-option of the host ubiquitin network with a panel of effector proteins. Here, using recombinantly expressed and purified proteins, analytic ultracentrifugation, structural analysis, and computational modeling, along with deubiquitinase (DUB), and bacterial infection assays, we found that the bacterial defective in organelle trafficking/intracellular multiplication effector Ceg23 is a member of the ovarian tumor (OTU) DUB family. We found that Ceg23 displays high specificity toward Lys-63-linked polyubiquitin chains and is localized on the LCV, where it removes ubiquitin moieties from proteins ubiquitinated by the Lys-63-chain type. Analysis of the crystal structure of a Ceg23 variant lacking two putative transmembrane domains at 2.80 Å resolution revealed that despite very limited homology to established members of the OTU family at the primary sequence level, Ceg23 harbors a catalytic motif resembling those associated with typical OTU-type DUBs. deletion increased the association of Lys-63-linked polyubiquitin with the bacterial phagosome, indicating that Ceg23 regulates Lys-63-linked ubiquitin signaling on the LCV. In summary, our findings indicate that Ceg23 contributes to the regulation of the association of Lys-63 type polyubiquitin with the phagosome. Future identification of host substrates targeted by Ceg23 could clarify the roles of these polyubiquitin chains in the intracellular life cycle of and Ceg23's role in bacterial virulence.
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http://dx.doi.org/10.1074/jbc.RA119.011758DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7008378PMC
February 2020

Infectious hematopoietic necrosis virus N protein suppresses fish IFN1 production by targeting the MITA.

Fish Shellfish Immunol 2020 Feb 24;97:523-530. Epub 2019 Dec 24.

Laboratory for Marine Biology and Biotechnology, Pilot National Laboratory for Marine Science and Technology (Qingdao), 266337, China; Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, The Key Laboratory of Innate Immune Biology of Fujian Province, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of the Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou, 350117, China; The Public Service Platform for Industrialization Development Technology of Marine Biological Medicine and Product of State Oceanic Administration, College of Life Sciences, Fujian Normal University, Fuzhou, 350117, China; Fujian Key Laboratory of Special Marine Bio-resources Sustainable Utilization, Fujian Normal University, Fuzhou, 350117, China.

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.
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http://dx.doi.org/10.1016/j.fsi.2019.12.075DOI Listing
February 2020

Legionella pneumophila regulates the activity of UBE2N by deamidase-mediated deubiquitination.

EMBO J 2020 02 11;39(4):e102806. Epub 2019 Dec 11.

Purdue Institute for Inflammation, Immunology and Infectious Disease and Department of Biological Sciences, Purdue University, West Lafayette, IN, USA.

The Legionella pneumophila effector MavC induces ubiquitination of the E2 ubiquitin-conjugating enzyme UBE2N by transglutamination, thereby abolishing its function in the synthesis of K -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.
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http://dx.doi.org/10.15252/embj.2019102806DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7024838PMC
February 2020

The Nuclear Matrix Protein SAFA Surveils Viral RNA and Facilitates Immunity by Activating Antiviral Enhancers and Super-enhancers.

Cell Host Microbe 2019 Sep;26(3):369-384.e8

Institute of Systems Biomedicine, Department of Immunology, Beijing Key Laboratory of Tumor Systems Biology, Peking University Health Science Center, Beijing 100191, China. Electronic address:

Pathogen pattern recognition receptors (PRRs) trigger innate immune responses to invading pathogens. All known PRRs for viral RNA have extranuclear localization. However, for many viruses, replication generates dsRNA in the nucleus. Here, we show that the nuclear matrix protein SAFA (also known as HnRNPU) functions as a nuclear viral dsRNA sensor for both DNA and RNA viruses. Upon recognition of viral dsRNA, SAFA oligomerizes and activates the enhancers of antiviral genes, including IFNB1. Moreover, SAFA is required for the activation of super-enhancers, which direct vigorous immune gene transcription to establish the antiviral state. Myeloid-specific SAFA-deficient mice were more susceptible to lethal HSV-1 and VSV infection, with decreased type I IFNs. Thus, SAFA functions as a nuclear viral RNA sensor and trans-activator to bridge innate sensing with chromatin remodeling and potentiate robust antiviral responses.
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http://dx.doi.org/10.1016/j.chom.2019.08.010DOI Listing
September 2019

Molecular Basis of BioJ, a Unique Gatekeeper in Bacterial Biotin Synthesis.

iScience 2019 Sep 22;19:796-808. Epub 2019 Aug 22.

Department of Pathogen Biology & Microbiology and General Intensive Care Unit of Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310058, China; College of Animal Science, Zhejiang University, Hangzhou 310058, China. Electronic address:

Biotin is an indispensable cofactor in the three domains of life. The unusual virulence factor BioJ of Francisella catalyzes the formation of pimeloyl-ACP, an intermediate in biotin synthesis. Here, we report the 1.58 Å crystal structure of BioJ, the enzymatic activity of which is determined with the in vitro reconstituted reaction and biotin bioassay in vivo. Unlike the paradigm BioH, BioJ displays an atypical α/β-hydrolase fold. A structurally conserved catalytic triad (S151, D248, and H278) of BioJ is functionally defined. A proposed model for BioJ catalysis involves two basic residues-rich cavities, of which cavity-1, rather than cavity-2, binds to the ACP moiety of its physiological substrate, pimeloyl-ACP methyl ester. In summary, this finding provides molecular insights into the BioJ gatekeeper of biotin synthesis.
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http://dx.doi.org/10.1016/j.isci.2019.08.028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6733898PMC
September 2019

HER2 recruits AKT1 to disrupt STING signalling and suppress antiviral defence and antitumour immunity.

Nat Cell Biol 2019 08 22;21(8):1027-1040. Epub 2019 Jul 22.

MOE Laboratory of Biosystems Homeostasis and Protection and Innovation Center for Cell Signaling Network, Life Sciences Institute, Zhejiang University, Hangzhou, China.

Sensing cytosolic DNA through the cGAS-STING pathway constitutes a widespread innate immune mechanism to monitor cellular damage and microbial invasion. Evading this surveillance is crucial in tumorigenesis, but the process remains largely unexplored. Here, we show that the receptor tyrosine kinase HER2 (also known as ErbB-2 or Neu) potently inhibits cGAS-STING signalling and prevents cancer cells from producing cytokines, entering senescence and undergoing apoptosis. HER2, but not EGFR, associates strongly with STING and recruits AKT1 (also known as PKB) to directly phosphorylate TBK1, which prevents the TBK1-STING association and TBK1 K63-linked ubiquitination, thus attenuating STING signalling. Unexpectedly, we observed that DNA sensing robustly activates the HER2-AKT1 axis, resulting in negative feedback. Accordingly, genetic or pharmacological targeting of the HER2-AKT1 cascade augments damage-induced cellular senescence and apoptosis, and enhances STING-mediated antiviral and antitumour immunity. Thus, our findings reveal a critical function of the oncogenic pathway in innate immune regulation and unexpectedly connect HER2-AKT1 signalling to the surveillance of cellular damage and antitumour immunity.
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http://dx.doi.org/10.1038/s41556-019-0352-zDOI Listing
August 2019

Regulation of phosphoribosyl ubiquitination by a calmodulin-dependent glutamylase.

Nature 2019 08 22;572(7769):387-391. Epub 2019 Jul 22.

Purdue Institute for Inflammation, Immunology and Infectious Disease and Department of Biological Sciences, Purdue University, West Lafayette, IN, USA.

The bacterial pathogen Legionella pneumophila creates an intracellular niche permissive for its replication by extensively modulating host-cell functions using hundreds of effector proteins delivered by its Dot/Icm secretion system. Among these, members of the SidE family (SidEs) regulate several cellular processes through a unique phosphoribosyl ubiquitination mechanism that bypasses the canonical ubiquitination machinery. The activity of SidEs is regulated by another Dot/Icm effector known as SidJ; however, the mechanism of this regulation is not completely understood. Here we demonstrate that SidJ inhibits the activity of SidEs by inducing the covalent attachment of glutamate moieties to SdeA-a member of the SidE family-at E860, one of the catalytic residues that is required for the mono-ADP-ribosyltransferase activity involved in ubiquitin activation. This inhibition by SidJ is spatially restricted in host cells because its activity requires the eukaryote-specific protein calmodulin (CaM). We solved a structure of SidJ-CaM in complex with AMP and found that the ATP used in this reaction is cleaved at the α-phosphate position by SidJ, which-in the absence of glutamate or modifiable SdeA-undergoes self-AMPylation. Our results reveal a mechanism of regulation in bacterial pathogenicity in which a glutamylation reaction that inhibits the activity of virulence factors is activated by host-factor-dependent acyl-adenylation.
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http://dx.doi.org/10.1038/s41586-019-1439-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6855250PMC
August 2019

Self-capping of nucleoprotein filaments protects the Newcastle disease virus genome.

Elife 2019 07 10;8. Epub 2019 Jul 10.

Institute of Molecular and Clinical Medicine, Kunming Medical University, Kunming, China.

Non-segmented negative-strand RNA viruses, such as measles, ebola and Newcastle disease viruses (NDV), encapsidate viral genomic RNAs into helical nucleocapsids, which serve as the template for viral replication and transcription. Here, the clam-shaped nucleocapsid structure, where the NDV viral genome is sequestered, was determined at 4.8 Å resolution by cryo-electron microscopy. The clam-shaped structure is composed of two single-turn spirals packed in a back-to-back mode. This tightly packed structure functions as a seed for the assembly of a nucleocapsid from both directions, facilitating the growth of double-headed filaments with two separate RNA strings inside. Disruption of this structure by mutations in its loop interface yielded a single-headed unfunctional filament.
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http://dx.doi.org/10.7554/eLife.45057DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6675542PMC
July 2019

High-throughput sequencing and analysis of microbial communities in the mangrove swamps along the coast of Beibu Gulf in Guangxi, China.

Sci Rep 2019 06 28;9(1):9377. Epub 2019 Jun 28.

The Key Laboratory of Innate Immune Biology of Fujian Province, Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, Biomedical Research Center of South China, Key Laboratory of OptoElectronic Science and Technology for Medicine of Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou, 350117, China.

Mangrove swamp is one of the world's richest and most productive marine ecosystems. This ecosystem also has a great ecological importance, but is highly susceptible to anthropogenic disturbances. The balance of mangrove ecosystem depends largely on the microbial communities in mangrove sediments. Thus, understanding how the mangrove microbial communities respond to spatial differences is essential for more accurate assessment of mangrove ecosystem health. To this end, we performed the first medium-distance (150 km) research on the biogeographic distribution of mangrove microbial communities. The hypervariable regions of 16S rRNA gene was sequenced by Illumina to compare the microbial communities in mangrove sediments collected from six locations (i.e. Zhenzhu harbor, Yuzhouping, Maowei Sea, Qinzhou harbor, Beihai city and Shankou) along the coastline of Beibu Gulf in Guangxi province, China. Collectively, Proteobacteria, Bacteroidetes, Chloroflexi, Actinobacteria, Parvarchaeota, Acidobacteria and Cyanobacteria were the predominant phyla in the mangrove sediments of this area. At genus level, the heat map of microbial communities reflected similarities between study sites and was in agreement with their biogeographic characteristics. Interestingly, the genera Desulfococcus, Arcobacter, Nitrosopumilus and Sulfurimonas showed differences in abundance between study sites. Furthermore, the principal component analysis (PCA) and unweighted UniFrac cluster tree of beta diversity were used to study the biogeographic diversity of the microbial communities. Relatively broader variation of microbial communities was found in Beihai city and Qinzhou harbour, suggesting that environmental condition and historical events may play an important role in shaping the bacterial communities as well. This is the first report on medium-distance range distribution of bacteria in the mangrove swamp ecosystem. Our data is valuable for monitoring and evaluation of the impact of human activity on mangrove habitats from the perspective of microbiome.
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http://dx.doi.org/10.1038/s41598-019-45804-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6599077PMC
June 2019
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