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1.
Microb Biotechnol ; 15(8): 2145-2159, 2022 08.
Article in English | MEDLINE | ID: covidwho-1961453

ABSTRACT

The growing world needs commodity amino acids such as L-glutamate and L-lysine for use as food and feed, and specialty amino acids for dedicated applications. To meet the supply a paradigm shift regarding their production is required. On the one hand, the use of sustainable and cheap raw materials is necessary to sustain low production cost and decrease detrimental effects of sugar-based feedstock on soil health and food security caused by competing uses of crops in the feed and food industries. On the other hand, the biotechnological methods to produce functionalized amino acids need to be developed further, and titres enhanced to become competitive with chemical synthesis methods. In the current review, we present successful strain mutagenesis and rational metabolic engineering examples leading to the construction of recombinant bacterial strains for the production of amino acids such as L-glutamate, L-lysine, L-threonine and their derivatives from methanol as sole carbon source. In addition, the fermentative routes for bioproduction of N-methylated amino acids are highlighted, with focus on three strategies: partial transfer of methylamine catabolism, S-adenosyl-L-methionine dependent alkylation and reductive methylamination of 2-oxoacids.


Subject(s)
Amino Acids , Corynebacterium glutamicum , Amino Acids/metabolism , Corynebacterium glutamicum/genetics , Glutamic Acid/metabolism , Lysine/metabolism , Metabolic Engineering , Methanol/metabolism
2.
Front Immunol ; 13: 919477, 2022.
Article in English | MEDLINE | ID: covidwho-1938621

ABSTRACT

The interferon-induced transmembrane protein 3 (IFITM3), a small molecule transmembrane protein induced by interferon, is generally conserved in vertebrates, which can inhibit infection by a diverse range of pathogenic viruses such as influenza virus. However, the precise antiviral mechanisms of IFITM3 remain unclear. At least four post-translational modifications (PTMs) were found to modulate the antiviral effect of IFITM3. These include positive regulation provided by S-palmitoylation of cysteine and negative regulation provided by lysine ubiquitination, lysine methylation, and tyrosine phosphorylation. IFITM3 S-palmitoylation is an enzymatic addition of a 16-carbon fatty acid on the three cysteine residues within or adjacent to its two hydrophobic domains at positions 71, 72, and 105, that is essential for its proper targeting, stability, and function. As S-palmitoylation is the only PTM known to enhance the antiviral activity of IFITM3, enzymes that add this modification may play important roles in IFN-induced immune responses. This study mainly reviews the research progresses on the antiviral mechanism of IFITM3, the regulation mechanism of S-palmitoylation modification on its subcellular localization, stability, and function, and the enzymes that mediate the S-palmitoylation modification of IFITM3, which may help elucidate the mechanism by which this IFN effector restrict virus replication and thus aid in the design of therapeutics targeted at pathogenic viruses.


Subject(s)
Antiviral Agents , Lipoylation , Animals , Antiviral Agents/metabolism , Antiviral Agents/pharmacology , Cysteine , Interferons/metabolism , Lysine/metabolism , RNA-Binding Proteins/metabolism
3.
Trends Biochem Sci ; 47(5): 372-374, 2022 05.
Article in English | MEDLINE | ID: covidwho-1821500

ABSTRACT

Modifications of cysteine residues in redox-sensitive proteins are key to redox signaling and stress response in all organisms. A novel type of redox switch was recently discovered that comprises lysine and cysteine residues covalently linked by an nitrogen-oxygen-sulfur (NOS) bridge. Here, we discuss chemical and biological implications of this discovery.


Subject(s)
Cysteine , Lysine , Cysteine/chemistry , Lysine/metabolism , Oxidation-Reduction , Oxidative Stress , Protein Processing, Post-Translational , Proteins/chemistry
4.
Viruses ; 14(4)2022 04 10.
Article in English | MEDLINE | ID: covidwho-1786079

ABSTRACT

The spread of SARS-CoV-2 variants in the population depends on their ability to anchor the ACE2 receptor in the host cells. Differences in the electrostatic potentials of the spike protein RBD (electropositive/basic) and ACE2 receptor (electronegative/acidic) play a key role in both the rapprochement and the recognition of the coronavirus by the cell receptors. Accordingly, point mutations that result in an increase in electropositively charged residues, e.g., arginine and lysine, especially in the RBD of spike proteins in the SARS-CoV-2 variants, could contribute to their spreading capacity by favoring their recognition by the electronegatively charged ACE2 receptors. All SARS-CoV-2 variants that have been recognized as being highly transmissible, such as the kappa (κ), delta (δ) and omicron (o) variants, which display an enhanced electropositive character in their RBDs associated with a higher number of lysine- or arginine-generating point mutations. Lysine and arginine residues also participate in the enhanced RBD-ACE2 binding affinity of the omicron variant, by creating additional salt bridges with aspartic and glutamic acid residues from ACE2. However, the effects of lysine- and arginine-generating point mutations on infectivity is more contrasted, since the overall binding affinity of omicron RBD for ACE2 apparently results from some epistasis among the whole set of point mutations.


Subject(s)
COVID-19 , SARS-CoV-2 , Angiotensin-Converting Enzyme 2/genetics , Arginine/genetics , Humans , Lysine/metabolism , Mutation , Point Mutation , Protein Binding , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/metabolism
5.
Brief Bioinform ; 23(2)2022 03 10.
Article in English | MEDLINE | ID: covidwho-1704326

ABSTRACT

Protein lysine crotonylation (Kcr) is an important type of posttranslational modification that is associated with a wide range of biological processes. The identification of Kcr sites is critical to better understanding their functional mechanisms. However, the existing experimental techniques for detecting Kcr sites are cost-ineffective, to a great need for new computational methods to address this problem. We here describe Adapt-Kcr, an advanced deep learning model that utilizes adaptive embedding and is based on a convolutional neural network together with a bidirectional long short-term memory network and attention architecture. On the independent testing set, Adapt-Kcr outperformed the current state-of-the-art Kcr prediction model, with an improvement of 3.2% in accuracy and 1.9% in the area under the receiver operating characteristic curve. Compared to other Kcr models, Adapt-Kcr additionally had a more robust ability to distinguish between crotonylation and other lysine modifications. Another model (Adapt-ST) was trained to predict phosphorylation sites in SARS-CoV-2, and outperformed the equivalent state-of-the-art phosphorylation site prediction model. These results indicate that self-adaptive embedding features perform better than handcrafted features in capturing discriminative information; when used in attention architecture, this could be an effective way of identifying protein Kcr sites. Together, our Adapt framework (including learning embedding features and attention architecture) has a strong potential for prediction of other protein posttranslational modification sites.


Subject(s)
Computational Biology , Deep Learning , Lysine/metabolism , Protein Processing, Post-Translational , Software , Algorithms , Benchmarking , Computational Biology/methods , Computational Biology/standards , Databases, Factual , Neural Networks, Computer , Phosphorylation , ROC Curve , Reproducibility of Results , User-Computer Interface
6.
Proc Natl Acad Sci U S A ; 119(9)2022 03 01.
Article in English | MEDLINE | ID: covidwho-1684239

ABSTRACT

High-fidelity replication of the large RNA genome of coronaviruses (CoVs) is mediated by a 3'-to-5' exoribonuclease (ExoN) in nonstructural protein 14 (nsp14), which excises nucleotides including antiviral drugs misincorporated by the low-fidelity viral RNA-dependent RNA polymerase (RdRp) and has also been implicated in viral RNA recombination and resistance to innate immunity. Here, we determined a 1.6-Å resolution crystal structure of severe acute respiratory syndrome CoV 2 (SARS-CoV-2) ExoN in complex with its essential cofactor, nsp10. The structure shows a highly basic and concave surface flanking the active site, comprising several Lys residues of nsp14 and the N-terminal amino group of nsp10. Modeling suggests that this basic patch binds to the template strand of double-stranded RNA substrates to position the 3' end of the nascent strand in the ExoN active site, which is corroborated by mutational and computational analyses. We also show that the ExoN activity can rescue a stalled RNA primer poisoned with sofosbuvir and allow RdRp to continue its extension in the presence of the chain-terminating drug, biochemically recapitulating proofreading in SARS-CoV-2 replication. Molecular dynamics simulations further show remarkable flexibility of multidomain nsp14 and suggest that nsp10 stabilizes ExoN for substrate RNA binding to support its exonuclease activity. Our high-resolution structure of the SARS-CoV-2 ExoN-nsp10 complex serves as a platform for future development of anticoronaviral drugs or strategies to attenuate the viral virulence.


Subject(s)
Exoribonucleases/chemistry , Molecular Dynamics Simulation , Nucleic Acid Conformation , Protein Domains , RNA, Viral/chemistry , SARS-CoV-2/enzymology , Viral Nonstructural Proteins/chemistry , Binding Sites/genetics , COVID-19/virology , Catalytic Domain , Crystallography, X-Ray , Exoribonucleases/genetics , Exoribonucleases/metabolism , Humans , Lysine/chemistry , Lysine/genetics , Lysine/metabolism , Mutation, Missense , Protein Binding , RNA, Viral/genetics , RNA, Viral/metabolism , SARS-CoV-2/physiology , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolism
7.
Nat Chem Biol ; 18(4): 368-375, 2022 04.
Article in English | MEDLINE | ID: covidwho-1684084

ABSTRACT

We recently reported the discovery of a lysine-cysteine redox switch in proteins with a covalent nitrogen-oxygen-sulfur (NOS) bridge. Here, a systematic survey of the whole protein structure database discloses that NOS bridges are ubiquitous redox switches in proteins of all domains of life and are found in diverse structural motifs and chemical variants. In several instances, lysines are observed in simultaneous linkage with two cysteines, forming a sulfur-oxygen-nitrogen-oxygen-sulfur (SONOS) bridge with a trivalent nitrogen, which constitutes an unusual native branching cross-link. In many proteins, the NOS switch contains a functionally essential lysine with direct roles in enzyme catalysis or binding of substrates, DNA or effectors, linking lysine chemistry and redox biology as a regulatory principle. NOS/SONOS switches are frequently found in proteins from human and plant pathogens, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and also in many human proteins with established roles in gene expression, redox signaling and homeostasis in physiological and pathophysiological conditions.


Subject(s)
COVID-19 , Cysteine , Cysteine/chemistry , Humans , Lysine/metabolism , Oxidation-Reduction , SARS-CoV-2
8.
Trends Immunol ; 43(2): 148-162, 2022 02.
Article in English | MEDLINE | ID: covidwho-1634995

ABSTRACT

Ubc13-catalyzed K63 ubiquitination is a major control point for immune signaling. Recent evidence has shown that the control of multiple immune functions, including chronic inflammation, pathogen responses, lymphocyte activation, and regulatory signaling, is altered by K63 ubiquitination. In this review, we detail the novel cellular sensors that are dependent on K63 ubiquitination for their function in the immune signaling network. Many pathogens, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), can target K63 ubiquitination to inhibit pathogen immune responses; we describe novel details of the pathways involved and summarize recent clinically relevant SARS-CoV-2-specific responses. We also discuss recent evidence that regulatory T cell (Treg) versus T helper (TH) 1 and TH17 cell subset regulation might involve K63 ubiquitination. Knowledge gaps that merit future investigation and clinically relevant pathways are also addressed.


Subject(s)
COVID-19 , Lysine , Humans , Lysine/metabolism , SARS-CoV-2 , Signal Transduction , Ubiquitin-Protein Ligases/metabolism , Ubiquitination
9.
Microbiol Spectr ; 10(1): e0061821, 2022 02 23.
Article in English | MEDLINE | ID: covidwho-1622002

ABSTRACT

The host transmembrane protein MARCH8 is a RING finger E3 ubiquitin ligase that downregulates various host transmembrane proteins, such as MHC-II. We have recently reported that MARCH8 expression in virus-producing cells impairs viral infectivity by reducing virion incorporation of not only HIV-1 envelope glycoprotein but also vesicular stomatitis virus G-glycoprotein through two different pathways. However, the MARCH8 inhibition spectrum remains largely unknown. Here, we show the antiviral spectrum of MARCH8 using viruses pseudotyped with a variety of viral envelope glycoproteins. Infection experiments revealed that viral envelope glycoproteins derived from the rhabdovirus, arenavirus, coronavirus, and togavirus (alphavirus) families were sensitive to MARCH8-mediated inhibition. Lysine mutations at the cytoplasmic tails of rabies virus-G, lymphocytic choriomeningitis virus glycoproteins, SARS-CoV and SARS-CoV-2 spike proteins, and Chikungunya virus and Ross River virus E2 proteins conferred resistance to MARCH8. Immunofluorescence showed impaired downregulation of the mutants of these viral envelope glycoproteins by MARCH8, followed by lysosomal degradation, suggesting that MARCH8-mediated ubiquitination leads to intracellular degradation of these envelopes. Indeed, rabies virus-G and Chikungunya virus E2 proteins proved to be clearly ubiquitinated. We conclude that MARCH8 has inhibitory activity on a variety of viral envelope glycoproteins whose cytoplasmic lysine residues are targeted by this antiviral factor. IMPORTANCE A member of the MARCH E3 ubiquitin ligase family, MARCH8, downregulates many different kinds of host transmembrane proteins, resulting in the regulation of cellular homeostasis. On the other hands, MARCH8 acts as an antiviral factor when it binds to and downregulates HIV-1 envelope glycoprotein and vesicular stomatitis virus G-glycoprotein that are viral transmembrane proteins. This study reveals that, as in the case of cellular membrane proteins, MARCH8 shows broad-spectrum inhibition against various viral envelope glycoproteins by recognizing their cytoplasmic lysine residues, resulting in lysosomal degradation.


Subject(s)
Antiviral Agents/pharmacology , Lysine/drug effects , Ubiquitin-Protein Ligases/pharmacology , Viral Envelope Proteins/chemistry , Blotting, Western , Down-Regulation , HEK293 Cells , HeLa Cells , Humans , Immunoprecipitation , Lysine/metabolism , Ubiquitination/physiology , Viral Envelope Proteins/drug effects
10.
Commun Biol ; 4(1): 1076, 2021 09 14.
Article in English | MEDLINE | ID: covidwho-1550352

ABSTRACT

Lysine-selective molecular tweezers are promising drug candidates against proteinopathies, viral infection, and bacterial biofilm. Despite demonstration of their efficacy in multiple cellular and animal models, important questions regarding their mechanism of action, including cell penetrance and intracellular distribution, have not been answered to date. The main impediment to answering these questions has been the low intrinsic fluorescence of the main compound tested to date, called CLR01. Here, we address these questions using new fluorescently labeled molecular tweezers derivatives. We show that these compounds are internalized in neurons and astrocytes, at least partially through dynamin-dependent endocytosis. In addition, we demonstrate that the molecular tweezers concentrate rapidly in acidic compartments, primarily lysosomes. Accumulation of molecular tweezers in lysosomes may occur both through the endosomal-lysosomal pathway and via the autophagy-lysosome pathway. Moreover, by visualizing colocalization of molecular tweezers, lysosomes, and tau aggregates we show that lysosomes likely are the main site for the intracellular anti-amyloid activity of molecular tweezers. These findings have important implications for the mechanism of action of molecular tweezers in vivo, explaining how administration of low doses of the compounds achieves high effective concentrations where they are needed, and supporting the development of these compounds as drugs for currently cureless proteinopathies.


Subject(s)
Astrocytes/metabolism , Bridged-Ring Compounds/metabolism , Endosomes/metabolism , Lysine/metabolism , Lysosomes/metabolism , Neurons/metabolism , Organophosphates/metabolism , Animals , Autophagy/drug effects , Cell Line, Tumor , Humans , Mice , Mice, Inbred C57BL
11.
Biochem J ; 478(14): 2789-2791, 2021 07 30.
Article in English | MEDLINE | ID: covidwho-1526112

ABSTRACT

Post-translational modifications (PTMs) on histone proteins are known as epigenetic marks that demarcate the status of chromatin. These modifications are 'read' by specific reader proteins, which in turn recruit additional factors to modulate chromatin accessibility and the activity of the underlying DNA. Accumulating evidence suggests that these modifications are not restricted solely to histones, many non-histone proteins may function in a similar way through mimicking the histones. In this commentary, we briefly discuss a systematic study of the discovery of histone H3 N-terminal mimicry proteins (H3TMs), and their implications in chromatin regulation and drug discoveries.


Subject(s)
Chromatin/metabolism , DNA/metabolism , Histones/metabolism , Protein Processing, Post-Translational , Animals , Chromatin/genetics , Chromatin Assembly and Disassembly , DNA/genetics , Humans , Lysine/metabolism , Methylation , Models, Biological
12.
Front Immunol ; 12: 651656, 2021.
Article in English | MEDLINE | ID: covidwho-1211812

ABSTRACT

Although immune dysfunction is a key feature of coronavirus disease 2019 (COVID-19), the metabolism-related mechanisms remain elusive. Here, by reanalyzing single-cell RNA sequencing data, we delineated metabolic remodeling in peripheral blood mononuclear cells (PBMCs) to elucidate the metabolic mechanisms that may lead to the progression of severe COVID-19. After scoring the metabolism-related biological processes and signaling pathways, we found that mono-CD14+ cells expressed higher levels of glycolysis-related genes (PKM, LDHA and PKM) and PPP-related genes (PGD and TKT) in severe patients than in mild patients. These genes may contribute to the hyperinflammation in mono-CD14+ cells of patients with severe COVID-19. The mono-CD16+ cell population in COVID-19 patients showed reduced transcription levels of genes related to lysine degradation (NSD1, KMT2E, and SETD2) and elevated transcription levels of genes involved in OXPHOS (ATP6V1B2, ATP5A1, ATP5E, and ATP5B), which may inhibit M2-like polarization. Plasma cells also expressed higher levels of the OXPHOS gene ATP13A3 in COVID-19 patients, which was positively associated with antibody secretion and survival of PCs. Moreover, enhanced glycolysis or OXPHOS was positively associated with the differentiation of memory B cells into plasmablasts or plasma cells. This study comprehensively investigated the metabolic features of peripheral immune cells and revealed that metabolic changes exacerbated inflammation in monocytes and promoted antibody secretion and cell survival in PCs in COVID-19 patients, especially those with severe disease.


Subject(s)
COVID-19/immunology , Glycolysis/genetics , Lysine/metabolism , Monocytes/metabolism , Single-Cell Analysis/methods , Adenosine Triphosphatases/blood , Adenosine Triphosphatases/genetics , Antibodies/metabolism , COVID-19/metabolism , COVID-19/physiopathology , Databases, Genetic , GPI-Linked Proteins/metabolism , Gene Ontology , Hematopoiesis/genetics , Humans , Inflammation/genetics , Inflammation/immunology , Inflammation/metabolism , Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/metabolism , Leukocytes, Mononuclear/pathology , Lipopolysaccharide Receptors/metabolism , Lysine/genetics , Membrane Transport Proteins/blood , Membrane Transport Proteins/genetics , Metabolic Networks and Pathways/genetics , Metabolic Networks and Pathways/physiology , Monocytes/immunology , Monocytes/pathology , Oxidative Phosphorylation , RNA-Seq , Receptors, IgG/metabolism , Signal Transduction/genetics , Signal Transduction/immunology , Transcriptome/genetics
13.
Cell Rep ; 33(3): 108286, 2020 10 20.
Article in English | MEDLINE | ID: covidwho-880155

ABSTRACT

Diffuse intrinsic pontine glioma (DIPG) is an incurable brain tumor of childhood characterized by histone mutations at lysine 27, which results in epigenomic dysregulation. There has been a failure to develop effective treatment for this tumor. Using a combined RNAi and chemical screen targeting epigenomic regulators, we identify the polycomb repressive complex 1 (PRC1) component BMI1 as a critical factor for DIPG tumor maintenance in vivo. BMI1 chromatin occupancy is enriched at genes associated with differentiation and tumor suppressors in DIPG cells. Inhibition of BMI1 decreases cell self-renewal and attenuates tumor growth due to induction of senescence. Prolonged BMI1 inhibition induces a senescence-associated secretory phenotype, which promotes tumor recurrence. Clearance of senescent cells using BH3 protein mimetics co-operates with BMI1 inhibition to enhance tumor cell killing in vivo.


Subject(s)
Aging/genetics , Diffuse Intrinsic Pontine Glioma/genetics , Polycomb Repressive Complex 1/metabolism , Astrocytoma/genetics , Brain Stem Neoplasms/drug therapy , Brain Stem Neoplasms/genetics , Cell Differentiation/genetics , Cell Line, Tumor , Cell Proliferation/drug effects , Child , Child, Preschool , Chromatin/genetics , Diffuse Intrinsic Pontine Glioma/drug therapy , Diffuse Intrinsic Pontine Glioma/metabolism , Epigenomics , Female , Glioma/drug therapy , Glioma/genetics , Glioma/pathology , Histones/metabolism , Humans , Lysine/metabolism , Male , Mutation , Neoplasm Recurrence, Local/drug therapy , Neoplasm Recurrence, Local/genetics , Polycomb Repressive Complex 1/antagonists & inhibitors , Polycomb Repressive Complex 1/genetics
14.
J Gen Virol ; 102(1)2021 01.
Article in English | MEDLINE | ID: covidwho-910383

ABSTRACT

The emerging pathogen severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused social and economic disruption worldwide, infecting over 9.0 million people and killing over 469 000 by 24 June 2020. Unfortunately, no vaccine or antiviral drug that completely eliminates the transmissible disease coronavirus disease 2019 (COVID-19) has been developed to date. Given that coronavirus nonstructural protein 1 (nsp1) is a good target for attenuated vaccines, it is of great significance to explore the detailed characteristics of SARS-CoV-2 nsp1. Here, we first confirmed that SARS-CoV-2 nsp1 had a conserved function similar to that of SARS-CoV nsp1 in inhibiting host-protein synthesis and showed greater inhibition efficiency, as revealed by ribopuromycylation and Renilla luciferase (Rluc) reporter assays. Specifically, bioinformatics and biochemical experiments showed that by interacting with 40S ribosomal subunit, the lysine located at amino acid 164 (K164) was the key residue that enabled SARS-CoV-2 nsp1 to suppress host gene expression. Furthermore, as an inhibitor of host-protein expression, SARS-CoV-2 nsp1 contributed to cell-cycle arrest in G0/G1 phase, which might provide a favourable environment for virus production. Taken together, this research uncovered the detailed mechanism by which SARS-CoV-2 nsp1 K164 inhibited host gene expression, laying the foundation for the development of attenuated vaccines based on nsp1 modification.


Subject(s)
Host-Pathogen Interactions/genetics , Lysine/genetics , Ribosomal Proteins/genetics , Ribosome Subunits, Small, Eukaryotic/genetics , SARS-CoV-2/genetics , Viral Nonstructural Proteins/genetics , Amino Acid Sequence , Amino Acid Substitution , Computational Biology/methods , G1 Phase Cell Cycle Checkpoints/genetics , Gene Expression Regulation , Genes, Reporter , HEK293 Cells , Humans , Luciferases/genetics , Luciferases/metabolism , Lysine/metabolism , Mutation , Ribosomal Proteins/antagonists & inhibitors , Ribosomal Proteins/metabolism , Ribosome Subunits, Small, Eukaryotic/metabolism , Ribosome Subunits, Small, Eukaryotic/virology , SARS Virus/genetics , SARS Virus/metabolism , SARS-CoV-2/metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Signal Transduction , Viral Nonstructural Proteins/metabolism
15.
Sci Adv ; 6(31)2020 07.
Article in English | MEDLINE | ID: covidwho-725277

ABSTRACT

The outbreak of the highly contagious and deadly severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), also known as coronavirus disease 2019 (COVID-19), has posed a serious threat to public health across the globe, calling for the development of effective diagnostic markers and therapeutics. Here, we report a highly reliable severity diagnostic biomarker, acetylated 676th lysine transforming growth factor-beta-induced protein (TGFBIp K676Ac). TGFBIp K676Ac was consistently elevated in the blood of patients with SARS-CoV-2 pneumonia (n = 113), especially in patients in the intensive care unit (ICU) compared to non-ICU patients. Patients' blood samples showed increased cytokines and lymphopenia, which are exemplary indicators of SARS-CoV-2 pneumonia. Treatment with TGFBIp neutralizing antibodies suppressed the cytokine storm. The increased level of TGFBIp K676Ac in ICU patients suggests the promise of this protein as a reliable severity diagnostic biomarker for severe SARS-CoV-2 disease.


Subject(s)
Betacoronavirus/pathogenicity , Coronavirus Infections/diagnosis , Cytokine Release Syndrome/diagnosis , Extracellular Matrix Proteins/immunology , Leukocytes, Mononuclear/immunology , Pneumonia, Viral/diagnosis , Protein Processing, Post-Translational , Respiratory Insufficiency/diagnosis , Transforming Growth Factor beta/immunology , Acetylation , Antibodies, Neutralizing/pharmacology , Betacoronavirus/immunology , Biomarkers/blood , COVID-19 , Case-Control Studies , Coronavirus Infections/blood , Coronavirus Infections/immunology , Coronavirus Infections/pathology , Cytokine Release Syndrome/blood , Cytokine Release Syndrome/immunology , Cytokine Release Syndrome/pathology , Extracellular Matrix Proteins/antagonists & inhibitors , Extracellular Matrix Proteins/genetics , Gene Expression , Humans , Intensive Care Units , Leukocyte Count , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/pathology , Leukocytes, Mononuclear/virology , Lung/blood supply , Lung/drug effects , Lung/pathology , Lung/virology , Lysine/metabolism , NF-kappa B/genetics , NF-kappa B/immunology , Pandemics , Pneumonia, Viral/blood , Pneumonia, Viral/immunology , Pneumonia, Viral/pathology , Primary Cell Culture , Prognosis , Respiratory Insufficiency/blood , Respiratory Insufficiency/immunology , Respiratory Insufficiency/pathology , SARS-CoV-2 , Severity of Illness Index , Transforming Growth Factor beta/antagonists & inhibitors , Transforming Growth Factor beta/genetics
16.
Mol Cell Proteomics ; 19(8): 1303-1309, 2020 08.
Article in English | MEDLINE | ID: covidwho-360321

ABSTRACT

MERS is a life-threatening disease and MERS-CoV has the potential to cause the next pandemic. Protein acetylation is known to play a crucial role in host response to viral infection. Acetylation of viral proteins encoded by other RNA viruses have been reported to affect viral replication. It is therefore of interest to see whether MERS-CoV proteins are also acetylated. Viral proteins obtained from infected cells were trypsin-digested into peptides. Acetylated peptides were enriched by immunoprecipitation and subject to nano-LC-Orbitrap analysis. Bioinformatic analysis was performed to assess the conservation level of identified acetylation sites and to predict the upstream regulatory factors. A total of 12 acetylation sites were identified from 7 peptides, which all belong to the replicase polyprotein pp1ab. All identified acetylation sites were found to be highly conserved across MERS-CoV sequences in NCBI database. Upstream factors, including deacetylases of the SIRT1 and HDAC families as well as acetyltransferases of the TIP60 family, were predicted to be responsible for regulating the acetylation events identified. Western blotting confirms that acetylation events indeed occur on pp1ab protein by expressing NSP4 in HEK293 cells. Acetylation events on MERS-CoV viral protein pp1ab were identified for the first time, which indicate that MERS-CoV might use the host acetylation machinery to regulate its enzyme activity and to achieve optimal replication. Upstream factors were predicted, which might facilitate further analysis of the regulatory mechanism of MERS-CoV replication.


Subject(s)
Lysine/metabolism , Middle East Respiratory Syndrome Coronavirus/metabolism , Viral Proteins/metabolism , Acetylation , HEK293 Cells , Humans
17.
Biochem Biophys Res Commun ; 526(4): 947-952, 2020 06 11.
Article in English | MEDLINE | ID: covidwho-38740

ABSTRACT

The outbreak of corona virus disease 2019 (COVID-19) caused by SARS-CoV-2 infection is spreading globally and quickly, leading to emerging health issues. SARS-CoV-2 enters into and infects host cells through its spike glycoprotein recognizing the cell receptor Angiotensin-converting enzyme II (ACE2). Here, we noticed that ACE2 was further enhanced by SARS-CoV-2 infection. Human germ cells and early embryos express high level of ACE2. Notably, RNA-seq result showed that reduction of H3K27me3, but not H3K4/9/36me3, led to upregulation of Ace2 expression in mouse germ cell line GC-2. In agreement with this result, we found in human embryonic stem cells that ACE2 expression was significantly increased in absence of EZH2, the major enzyme catalyzing H3K27me3. ChIP-seq analysis further confirmed decrease of H3K27me3 signal and increase of H3K27ac signal at ACE2 promoter upon EZH2 knockout. Therefore, we propose that EZH2-mediated H3K27me3 at ACE2 promoter region inhibits ACE2 expression in mammalian cells. This regulatory pattern may also exist in other human cells and tissues. Our discovery provides clues for pathogenesis and targeted drug therapy towards ACE2 expression for prevention and adjuvant therapy of COVID-19.


Subject(s)
Betacoronavirus/physiology , Coronavirus Infections/virology , Enhancer of Zeste Homolog 2 Protein/metabolism , Epigenesis, Genetic , Gene Expression Regulation , Peptidyl-Dipeptidase A/genetics , Pneumonia, Viral/virology , Angiotensin-Converting Enzyme 2 , Animals , COVID-19 , Embryonic Stem Cells , Gene Knockout Techniques , Histone Code , Histones/chemistry , Histones/metabolism , Humans , Lysine/analysis , Lysine/metabolism , Methylation , Mice , Organ Specificity , Pandemics , Promoter Regions, Genetic , SARS-CoV-2 , Transcription, Genetic , Up-Regulation
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