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1.
PLoS Pathog ; 18(7): e1010660, 2022 07.
Article in English | MEDLINE | ID: covidwho-1993526

ABSTRACT

Coxiella burnetii is the etiological agent of the zoonotic disease Q fever, which is featured by its ability to replicate in acid vacuoles resembling the lysosomal network. One key virulence determinant of C. burnetii is the Dot/Icm system that transfers more than 150 effector proteins into host cells. These effectors function to construct the lysosome-like compartment permissive for bacterial replication, but the functions of most of these effectors remain elusive. In this study, we used an affinity tag purification mass spectrometry (AP-MS) approach to generate a C. burnetii-human protein-protein interaction (PPI) map involving 53 C. burnetii effectors and 3480 host proteins. This PPI map revealed that the C. burnetii effector CBU0425 (designated CirB) interacts with most subunits of the 20S core proteasome. We found that ectopically expressed CirB inhibits hydrolytic activity of the proteasome. In addition, overexpression of CirB in C. burnetii caused dramatic inhibition of proteasome activity in host cells, while knocking down CirB expression alleviated such inhibitory effects. Moreover, we showed that a region of CirB that spans residues 91-120 binds to the proteasome subunit PSMB5 (beta 5). Finally, PSMB5 knockdown promotes C. burnetii virulence, highlighting the importance of proteasome activity modulation during the course of C. burnetii infection.


Subject(s)
Coxiella burnetii , Q Fever , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Host-Pathogen Interactions , Humans , Proteasome Endopeptidase Complex/genetics , Proteasome Endopeptidase Complex/metabolism , Protein Interaction Maps , Q Fever/metabolism , Vacuoles/metabolism
2.
Biomolecules ; 12(7)2022 07 01.
Article in English | MEDLINE | ID: covidwho-1917275

ABSTRACT

Ubiquitin is a small protein that is conjugated to target proteins to signal a great number of critical biological processes. Impaired ubiquitin signaling and defects in the ubiquitin proteasome system (UPS) surveillance are implicated in many human diseases, including cancer. Characterization of the physiological roles of UPS components and their regulatory mechanisms is therefore vital for the identification of therapeutic targets and the development of tools and paradigms to better understand and treat human diseases. In this Special Issue, we assembled seven original research and review articles to provide insights on the multifaceted role of the UPS in pathogenesis and disease, covering the areas of molecular and cellular mechanisms of UPS enzymes, biochemical and biophysical characterization strategies, drug development, and targeted protein degradation.


Subject(s)
Neoplasms , Ubiquitin , Humans , Neoplasms/genetics , Proteasome Endopeptidase Complex/metabolism , Signal Transduction , Ubiquitin/metabolism , Ubiquitin-Protein Ligases/metabolism , Ubiquitination
3.
Nat Commun ; 13(1): 2442, 2022 05 04.
Article in English | MEDLINE | ID: covidwho-1890176

ABSTRACT

Interferon restricts SARS-CoV-2 replication in cell culture, but only a handful of Interferon Stimulated Genes with antiviral activity against SARS-CoV-2 have been identified. Here, we describe a functional CRISPR/Cas9 screen aiming at identifying SARS-CoV-2 restriction factors. We identify DAXX, a scaffold protein residing in PML nuclear bodies known to limit the replication of DNA viruses and retroviruses, as a potent inhibitor of SARS-CoV-2 and SARS-CoV replication in human cells. Basal expression of DAXX is sufficient to limit the replication of SARS-CoV-2, and DAXX over-expression further restricts infection. DAXX restricts an early, post-entry step of the SARS-CoV-2 life cycle. DAXX-mediated restriction of SARS-CoV-2 is independent of the SUMOylation pathway but dependent on its D/E domain, also necessary for its protein-folding activity. SARS-CoV-2 infection triggers the re-localization of DAXX to cytoplasmic sites and promotes its degradation. Mechanistically, this process is mediated by the viral papain-like protease (PLpro) and the proteasome. Together, these results demonstrate that DAXX restricts SARS-CoV-2, which in turn has evolved a mechanism to counteract its action.


Subject(s)
COVID-19 , SARS-CoV-2 , CRISPR-Cas Systems , Co-Repressor Proteins/genetics , Co-Repressor Proteins/metabolism , Humans , Interferons/metabolism , Molecular Chaperones/genetics , Molecular Chaperones/metabolism , Proteasome Endopeptidase Complex/metabolism
4.
mBio ; 13(3): e0130022, 2022 06 28.
Article in English | MEDLINE | ID: covidwho-1874506

ABSTRACT

Ubiquitin signaling is essential for immunity to restrict pathogen proliferation. Due to its enormous impact on human health and the global economy, intensive efforts have been invested in studying severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its interactions with hosts. However, the role of the ubiquitin network in pathogenicity has not yet been explored. Here, we found that ORF9b of SARS-CoV-2 is ubiquitinated on Lys-4 and Lys-40 by unknown E3 ubiquitin ligases and is degraded by the ubiquitin proteasomal system. Importantly, we identified USP29 as a host factor that prevents ORF9b ubiquitination and subsequent degradation. USP29 interacts with the carboxyl end of ORF9b and removes ubiquitin chains from the protein, thereby inhibiting type I interferon (IFN) induction and NF-κB activation. We also found that ORF9b stabilization by USP29 enhanced the virulence of VSV-eGFP and transcription and replication-competent SARS-CoV-2 virus-like-particles (trVLP). Moreover, we observed that the mRNA level of USP29 in SARS-CoV-2 patients was higher than that in healthy people. Our findings provide important evidence indicating that targeting USP29 may effectively combat SARS-CoV-2 infection. IMPORTANCE Coronavirus disease 2019 (COVID-19) is a current global health threat caused by SARS-CoV-2. The innate immune response such as type I IFN (IFN-I) is the first line of host defense against viral infections, whereas SARS-CoV-2 proteins antagonize IFN-I production through distinct mechanisms. Among them, ORF9b inhibits the canonical IκB kinase alpha (IKKɑ)/ß/γ-NF-κB signaling and subsequent IFN production; therefore, discovering the regulation of ORF9b by the host might help develop a novel antiviral strategy. Posttranslational modification of proteins by ubiquitination regulates many biological processes, including viral infections. Here, we report that ORF9b is ubiquitinated and degraded through the proteasome pathway, whereas deubiquitinase USP29 deubiquitinates ORF9b and prevents its degradation, resulting in the enhancement of ORF9b-mediated inhibition of IFN-I and NF-κB activation and the enhancement of virulence of VSV-eGFP and SARS-CoV-2 trVLP.


Subject(s)
Biological Phenomena , COVID-19 , Coronavirus Nucleocapsid Proteins/metabolism , Deubiquitinating Enzymes , Humans , Immunity, Innate , NF-kappa B , Phosphoproteins/metabolism , Proteasome Endopeptidase Complex , SARS-CoV-2/genetics , Ubiquitin-Specific Proteases , Ubiquitins , Virulence
5.
Phytomedicine ; 103: 154215, 2022 Aug.
Article in English | MEDLINE | ID: covidwho-1867672

ABSTRACT

BACKGROUND: Severe Acute Respiratory Syndrome Coronavirus Type 2 (SARS-CoV-2) induces a global serious pandemic and is responsible for over 4 million human deaths. Currently, although various vaccines have been developed, humans can still get SARS-CoV-2 infection after being vaccinated. Therefore, the blocking of SARS-CoV-2 infection may be potential therapeutic strategies. Ganoderma microsporum immunomodulatory protein (GMI), a small fungal protein, is cloned from Ganoderma microsporum. It exhibits anti-cancer and immunomodulatory functions. Currently, it is still unclear whether GMI involves in interfering with viral infection. PURPOSE: This study aimed to examine the potential functions and mechanisms of GMI on inhibiting SARS-CoV-2 pseudovirus infection. METHODS: The effects of GMI were examined in vitro on ACE2 overexpressing HEK293T (HEK293T/ACE2) cells exposed to SARS-CoV-2 Spike lentiviral pseudovirus encoding a green fluorescent protein (GFP) gene. The infection efficacy was determined using fluorescence microscopy and flow cytometry. The protein level of ACE2 was verified by Western blot. The effects of GMI on cell viability of HEK293T/ACE2 and lung epithelial WI38-2RA cells were determined by MTT assay. Mice received GMI via nebulizer. RESULTS: GMI did not affect the cell viability of HEK293T/ACE2, WI38-2RA and macrophages. Functional studies showed that GMI inhibited GFP expressing SARS-CoV-2 pseudovirus from infecting HEK293T/ACE2 cells. GMI slightly interfered the interaction between ACE2 and Spike protein. GMI interacted with S2 domain of Spike protein. Specifically, GMI dramatically reduced ACE2 expression in HEK293T/ACE2 and WI38-2RA cells. Mechanistically, GMI induced ACE2 degradation via activating protein degradation system, including proteasome and lysosome. Abolishing proteasome and lysosome by MG132 and bafilomycin A1, respectively, rescued GMI-reduced ACE2 levels. In addition, GMI triggered dynamin and lipid raft-mediated ACE2 endocytosis. ACE2 levels were downregulated in the lung tissue after the mice inhaling GMI. CONCLUSIONS: GMI prevents SARS-CoV-2 pseudovirus infection via induction of ACE2 degradation in host cells. Our findings suggest that GMI will be a potential prevention agent to alleviate SARS-CoV-2 infection.


Subject(s)
COVID-19 , SARS-CoV-2 , Angiotensin-Converting Enzyme 2 , Animals , COVID-19/drug therapy , Ganoderma , HEK293 Cells , Humans , Mice , Proteasome Endopeptidase Complex/metabolism , Protein Binding , Spike Glycoprotein, Coronavirus/metabolism , Viral Pseudotyping
6.
Aging (Albany NY) ; 14(10): 4195-4210, 2022 05 23.
Article in English | MEDLINE | ID: covidwho-1863474

ABSTRACT

Previous studies have shown that the polyamine spermidine increased the maximum life span in C. elegans and the median life span in mice. Since spermidine increases autophagy, we asked if treatment with chloroquine, an inhibitor of autophagy, would shorten the lifespan of mice. Recently, chloroquine has intensively been discussed as a treatment option for COVID-19 patients. To rule out unfavorable long-term effects on longevity, we examined the effect of chronic treatment with chloroquine given in the drinking water on the lifespan and organ pathology of male middle-aged NMRI mice. We report that, surprisingly, daily treatment with chloroquine extended the median life span by 11.4% and the maximum life span of the middle-aged male NMRI mice by 11.8%. Subsequent experiments show that the chloroquine-induced lifespan elevation is associated with dose-dependent increase in LC3B-II, a marker of autophagosomes, in the liver and heart that was confirmed by transmission electron microscopy. Quite intriguingly, chloroquine treatment was also associated with a decrease in glycogenolysis in the liver suggesting a compensatory mechanism to provide energy to the cell. Accumulation of autophagosomes was paralleled by an inhibition of proteasome-dependent proteolysis in the liver and the heart as well as with decreased serum levels of insulin growth factor binding protein-3 (IGFBP3), a protein associated with longevity. We propose that inhibition of proteasome activity in conjunction with an increased number of autophagosomes and decreased levels of IGFBP3 might play a central role in lifespan extension by chloroquine in male NMRI mice.


Subject(s)
Autophagy , Chloroquine , Glycogenolysis , Longevity , Proteasome Endopeptidase Complex , Proteasome Inhibitors , Animals , Autophagy/drug effects , COVID-19/drug therapy , Chloroquine/pharmacology , Glycogen , Glycogenolysis/drug effects , Longevity/drug effects , Male , Mice , Proteasome Inhibitors/pharmacology , Spermidine/pharmacology
7.
Ann N Y Acad Sci ; 1510(1): 79-99, 2022 04.
Article in English | MEDLINE | ID: covidwho-1822055

ABSTRACT

Targeted protein degradation is critical for proper cellular function and development. Protein degradation pathways, such as the ubiquitin proteasomes system, autophagy, and endosome-lysosome pathway, must be tightly regulated to ensure proper elimination of misfolded and aggregated proteins and regulate changing protein levels during cellular differentiation, while ensuring that normal proteins remain unscathed. Protein degradation pathways have also garnered interest as a means to selectively eliminate target proteins that may be difficult to inhibit via other mechanisms. On June 7 and 8, 2021, several experts in protein degradation pathways met virtually for the Keystone eSymposium "Targeting protein degradation: from small molecules to complex organelles." The event brought together researchers working in different protein degradation pathways in an effort to begin to develop a holistic, integrated vision of protein degradation that incorporates all the major pathways to understand how changes in them can lead to disease pathology and, alternatively, how they can be leveraged for novel therapeutics.


Subject(s)
Proteasome Endopeptidase Complex , Ubiquitin , Autophagy/physiology , Humans , Organelles , Proteasome Endopeptidase Complex/metabolism , Proteins/metabolism , Proteolysis , Ubiquitin/metabolism
8.
J Am Chem Soc ; 144(17): 7568-7572, 2022 05 04.
Article in English | MEDLINE | ID: covidwho-1795843

ABSTRACT

The COVID-19 pandemic has been a public health emergency with continuously evolving deadly variants around the globe. Among many preventive and therapeutic strategies, the design of covalent inhibitors targeting the main protease (Mpro) of SARS-CoV-2 that causes COVID-19 has been one of the hotly pursued areas. Currently, about 30% of marketed drugs that target enzymes are covalent inhibitors. Such inhibitors have been shown in recent years to have many advantages that counteract past reservation of their potential off-target activities, which can be minimized by modulation of the electrophilic warhead and simultaneous optimization of nearby noncovalent interactions. This process can be greatly accelerated by exploration of binding affinities using computational models, which are not well-established yet due to the requirement of capturing the chemical nature of covalent bond formation. Here, we present a robust computational method for effective prediction of absolute binding free energies (ABFEs) of covalent inhibitors. This is done by integrating the protein dipoles Langevin dipoles method (in the PDLD/S-LRA/ß version) with quantum mechanical calculations of the energetics of the reaction of the warhead and its amino acid target, in water. This approach evaluates the combined effects of the covalent and noncovalent contributions. The applicability of the method is illustrated by predicting the ABFEs of covalent inhibitors of SARS-CoV-2 Mpro and the 20S proteasome. Our results are found to be reliable in predicting ABFEs for cases where the warheads are significantly different. This computational protocol might be a powerful tool for designing effective covalent inhibitors especially for SARS-CoV-2 Mpro and for targeted protein degradation.


Subject(s)
COVID-19 , SARS-CoV-2 , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , COVID-19/drug therapy , Coronavirus 3C Proteases , Humans , Molecular Docking Simulation , Pandemics , Protease Inhibitors/chemistry , Proteasome Endopeptidase Complex
9.
Molecules ; 27(7)2022 Mar 31.
Article in English | MEDLINE | ID: covidwho-1785838

ABSTRACT

Obesity is the most common nutritional disorder in the developed world and is associated with important comorbidities. Pancreatic lipase (PL) inhibitors play a key role in the metabolism of human fat. A series of novel epoxyketones peptide derivatives were investigated for their pancreatic lipase inhibitory activity. The epoxyketone moiety is a well-known reactive electrophile group that has been used as part of proteasome inhibitors in cancer therapy, and it is widely believed that these are very selective for targeting the proteasome active site. Here we investigated various peptide derivatives with an epoxide warhead for their anti-lipase activity. The assessment of these novel epoxyketones was performed by an in-house method that we developed for rapid screening and identification of lipase inhibitors using GC-FID. Herein, we present a novel anti-lipase pharmacophore based on epoxyketone peptide derivatives that showed potent anti-lipase activity. Many of these derivatives had comparable or more potent activity than the clinically used lipase inhibitors such as orlistat. In addition, the lipase appears to be inhibited by a wide range of epoxyketone analogues regardless of the configuration of the epoxide in the epoxyketone moiety. The presented data in this study shows the first example of the use of epoxyketone peptides as novel lipase inhibitors.


Subject(s)
Peptides , Proteasome Inhibitors , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Epoxy Compounds/pharmacology , Humans , Lipase , Peptides/chemistry , Peptides/pharmacology , Proteasome Endopeptidase Complex/chemistry , Proteasome Inhibitors/chemistry
10.
Biomolecules ; 12(3)2022 03 13.
Article in English | MEDLINE | ID: covidwho-1742313

ABSTRACT

Severe COVID-19 disease leads to hypoxemia, inflammation and lymphopenia. Viral infection induces cellular stress and causes the activation of the innate immune response. The ubiquitin-proteasome system (UPS) is highly implicated in viral immune response regulation. The main function of the proteasome is protein degradation in its active form, which recognises and binds to ubiquitylated proteins. Some proteasome subunits have been reported to be upregulated under hypoxic and hyperinflammatory conditions. Here, we conducted a prospective cohort study of COVID-19 patients (n = 44) and age-and sex-matched controls (n = 20). In this study, we suggested that hypoxia could induce the overexpression of certain genes encoding for subunits from the α and ß core of the 20S proteasome and from regulatory particles (19S and 11S) in COVID-19 patients. Furthermore, the gene expression of proteasome subunits was associated with lymphocyte count reduction and positively correlated with inflammatory molecular and clinical markers. Given the importance of the proteasome in maintaining cellular homeostasis, including the regulation of the apoptotic and pyroptotic pathways, these results provide a potential link between COVID-19 complications and proteasome gene expression.


Subject(s)
COVID-19 , Lymphopenia , COVID-19/genetics , Humans , Hypoxia , Inflammation/genetics , Lymphopenia/genetics , Prospective Studies , Proteasome Endopeptidase Complex/genetics , Proteasome Endopeptidase Complex/metabolism
11.
Life Sci Alliance ; 5(6)2022 06.
Article in English | MEDLINE | ID: covidwho-1732625

ABSTRACT

Inflammasomes are cytosolic innate immune sensors of pathogen infection and cellular damage that induce caspase-1-mediated inflammation upon activation. Although inflammation is protective, uncontrolled excessive inflammation can cause inflammatory diseases and can be detrimental, such as in coronavirus disease (COVID-19). However, the underlying mechanisms that control inflammasome activation are incompletely understood. Here we report that the leucine-rich repeat (LRR) protein ribonuclease inhibitor (RNH1), which shares homology with LRRs of NLRP (nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain containing) proteins, attenuates inflammasome activation. Deletion of RNH1 in macrophages increases interleukin (IL)-1ß production and caspase-1 activation in response to inflammasome stimulation. Mechanistically, RNH1 decreases pro-IL-1ß expression and induces proteasome-mediated caspase-1 degradation. Corroborating this, mouse models of monosodium urate (MSU)-induced peritonitis and lipopolysaccharide (LPS)-induced endotoxemia, which are dependent on caspase-1, respectively, show increased neutrophil infiltration and lethality in Rnh1 -/- mice compared with wild-type mice. Furthermore, RNH1 protein levels were negatively related with disease severity and inflammation in hospitalized COVID-19 patients. We propose that RNH1 is a new inflammasome regulator with relevance to COVID-19 severity.


Subject(s)
COVID-19/pathology , Carrier Proteins/metabolism , Inflammasomes/metabolism , /metabolism , Animals , COVID-19/immunology , Caspase 1/metabolism , Humans , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , NF-kappa B/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Patient Acuity , Proteasome Endopeptidase Complex/metabolism
12.
Int J Mol Sci ; 23(1)2022 Jan 01.
Article in English | MEDLINE | ID: covidwho-1615843

ABSTRACT

Virus infection of eukaryotes triggers cellular innate immune response, a major arm of which is the type I interferon (IFN) family of cytokines. Binding of IFN to cell surface receptors triggers a signaling cascade in which the signal transducer and activator of transcription 2 (STAT2) plays a key role, ultimately leading to an antiviral state of the cell. In retaliation, many viruses counteract the immune response, often by the destruction and/or inactivation of STAT2, promoted by specific viral proteins that do not possess protease activities of their own. This review offers a summary of viral mechanisms of STAT2 subversion with emphasis on degradation. Some viruses also destroy STAT1, another major member of the STAT family, but most viruses are selective in targeting either STAT2 or STAT1. Interestingly, degradation of STAT2 by a few viruses requires the presence of both STAT proteins. Available evidence suggests a mechanism in which multiple sites and domains of STAT2 are required for engagement and degradation by a multi-subunit degradative complex, comprising viral and cellular proteins, including the ubiquitin-proteasomal system. However, the exact molecular nature of this complex and the alternative degradation mechanisms remain largely unknown, as critically presented here with prospective directions of future study.


Subject(s)
Proteolysis , STAT2 Transcription Factor/metabolism , Viruses/metabolism , Amino Acid Sequence , Animals , Humans , Models, Biological , Proteasome Endopeptidase Complex/metabolism , STAT2 Transcription Factor/chemistry , STAT2 Transcription Factor/ultrastructure , Ubiquitin/metabolism
13.
Nat Struct Mol Biol ; 28(7): 614-625, 2021 07.
Article in English | MEDLINE | ID: covidwho-1550333

ABSTRACT

p97 processes ubiquitinated substrates and plays a central role in cellular protein homeostasis. Here, we report a series of cryo-EM structures of the substrate-engaged human p97 complex with resolutions ranging from 2.9 to 3.8 Å that captured 'power-stroke'-like motions of both the D1 and D2 ATPase rings of p97. A key feature of these structures is the critical conformational changes of the intersubunit signaling (ISS) motifs, which tighten the binding of nucleotides and neighboring subunits and contribute to the spiral staircase conformation of the D1 and D2 rings. In addition, we determined the cryo-EM structure of human p97 in complex with NMS-873, a potent p97 inhibitor, at a resolution of 2.4 Å. The structures showed that NMS-873 binds at a cryptic groove in the D2 domain and interacts with the ISS motif, preventing its conformational change and thus blocking substrate translocation allosterically.


Subject(s)
Adenosine Triphosphate/chemistry , Protein Folding , Proteostasis/physiology , Signal Transduction/physiology , Valosin Containing Protein/metabolism , Acetanilides/pharmacology , Animals , Benzothiazoles/pharmacology , Cryoelectron Microscopy , Endoplasmic Reticulum/metabolism , Endoplasmic Reticulum-Associated Degradation/physiology , Humans , Intracellular Signaling Peptides and Proteins/metabolism , Nuclear Proteins/metabolism , Proteasome Endopeptidase Complex/metabolism , Protein Conformation , Ubiquitinated Proteins/metabolism , Valosin Containing Protein/antagonists & inhibitors
14.
Cell Mol Immunol ; 19(1): 67-78, 2022 01.
Article in English | MEDLINE | ID: covidwho-1541184

ABSTRACT

The global coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused severe morbidity and mortality in humans. It is urgent to understand the function of viral genes. However, the function of open reading frame 10 (ORF10), which is uniquely expressed by SARS-CoV-2, remains unclear. In this study, we showed that overexpression of ORF10 markedly suppressed the expression of type I interferon (IFN-I) genes and IFN-stimulated genes. Then, mitochondrial antiviral signaling protein (MAVS) was identified as the target via which ORF10 suppresses the IFN-I signaling pathway, and MAVS was found to be degraded through the ORF10-induced autophagy pathway. Furthermore, overexpression of ORF10 promoted the accumulation of LC3 in mitochondria and induced mitophagy. Mechanistically, ORF10 was translocated to mitochondria by interacting with the mitophagy receptor Nip3-like protein X (NIX) and induced mitophagy through its interaction with both NIX and LC3B. Moreover, knockdown of NIX expression blocked mitophagy activation, MAVS degradation, and IFN-I signaling pathway inhibition by ORF10. Consistent with our observations, in the context of SARS-CoV-2 infection, ORF10 inhibited MAVS expression and facilitated viral replication. In brief, our results reveal a novel mechanism by which SARS-CoV-2 inhibits the innate immune response; that is, ORF10 induces mitophagy-mediated MAVS degradation by binding to NIX.


Subject(s)
COVID-19/genetics , COVID-19/virology , Immunity, Innate/immunology , Open Reading Frames , SARS-CoV-2/genetics , Signal Transduction , Adaptor Proteins, Signal Transducing/metabolism , Antiviral Agents/metabolism , Autophagy/immunology , Gene Silencing , HEK293 Cells , HeLa Cells , Humans , Interferon Type I/metabolism , Mitochondria/metabolism , Mitophagy , Proteasome Endopeptidase Complex/metabolism , Ubiquitination , Viral Proteins/metabolism , Virus Replication
15.
Biomolecules ; 11(9)2021 09 08.
Article in English | MEDLINE | ID: covidwho-1438506

ABSTRACT

The majority of critically ill intensive care unit (ICU) patients with severe sepsis develop ICU-acquired weakness (ICUAW) characterized by loss of muscle mass, reduction in myofiber size and decreased muscle strength leading to persisting physical impairment. This phenotype results from a dysregulated protein homeostasis with increased protein degradation and decreased protein synthesis, eventually causing a decrease in muscle structural proteins. The ubiquitin proteasome system (UPS) is the predominant protein-degrading system in muscle that is activated during diverse muscle atrophy conditions, e.g., inflammation. The specificity of UPS-mediated protein degradation is assured by E3 ubiquitin ligases, such as atrogin-1 and MuRF1, which target structural and contractile proteins, proteins involved in energy metabolism and transcription factors for UPS-dependent degradation. Although the regulation of activity and function of E3 ubiquitin ligases in inflammation-induced muscle atrophy is well perceived, the contribution of the proteasome to muscle atrophy during inflammation is still elusive. During inflammation, a shift from standard- to immunoproteasome was described; however, to which extent this contributes to muscle wasting and whether this changes targeting of specific muscular proteins is not well described. This review summarizes the function of the main proinflammatory cytokines and acute phase response proteins and their signaling pathways in inflammation-induced muscle atrophy with a focus on UPS-mediated protein degradation in muscle during sepsis. The regulation and target-specificity of the main E3 ubiquitin ligases in muscle atrophy and their mode of action on myofibrillar proteins will be reported. The function of the standard- and immunoproteasome in inflammation-induced muscle atrophy will be described and the effects of proteasome-inhibitors as treatment strategies will be discussed.


Subject(s)
Inflammation/pathology , Muscle, Skeletal/metabolism , Muscle, Skeletal/pathology , Proteasome Endopeptidase Complex/metabolism , Ubiquitin/metabolism , Animals , Cytokines/metabolism , Humans , Proteolysis
17.
J Cell Biochem ; 123(2): 161-182, 2022 02.
Article in English | MEDLINE | ID: covidwho-1405827

ABSTRACT

Viruses are known to cause various diseases in human and also infect other species such as animal plants, fungi, and bacteria. Replication of viruses depends upon their interaction with hosts. Human cells are prone to such unwanted viral infections. Disintegration and reconstitution require host machinery and various macromolecules like DNA, RNA, and proteins are invaded by viral particles. E3 ubiquitin ligases are known for their specific function, that is, recognition of their respective substrates for intracellular degradation. Still, we do not understand how ubiquitin proteasome system-based enzymes E3 ubiquitin ligases do their functional interaction with different viruses. Whether E3 ubiquitin ligases help in the elimination of viral components or viruses utilize their molecular capabilities in their intracellular propagation is not clear. The first time our current article comprehends fundamental concepts and new insights on the different viruses and their interaction with various E3 Ubiquitin Ligases. In this review, we highlight the molecular pathomechanism of viruses linked with E3 Ubiquitin Ligases dependent mechanisms. An enhanced understanding of E3 Ubiquitin Ligase-mediated removal of viral proteins may open new therapeutic strategies against viral infections.


Subject(s)
Ubiquitin-Protein Ligases/physiology , Viral Proteins/physiology , Virus Diseases/enzymology , Virus Replication/physiology , COVID-19/drug therapy , Cell Transformation, Viral/physiology , Cullin Proteins/physiology , Endosomes/virology , Host-Pathogen Interactions , Humans , Immunity, Innate , Inflammation/enzymology , Inflammation/virology , Neoplasms/enzymology , Neoplasms/virology , Oncogenic Viruses/physiology , Proteasome Endopeptidase Complex/metabolism , Proteolysis , Tripartite Motif Proteins/physiology , Ubiquitin-Protein Ligases/antagonists & inhibitors , Virus Diseases/immunology , Virus Diseases/virology , Virus Replication/drug effects
18.
Curr Opin Virol ; 50: 183-191, 2021 10.
Article in English | MEDLINE | ID: covidwho-1401390

ABSTRACT

Immunodominance is a complex and highly debated topic of T cell biology. The current SARS-CoV-2 pandemic has provided the opportunity to profile adaptive immune responses and determine molecular factors contributing to emerging responses towards immunodominant viral epitopes. Here, we discuss parameters that alter the dynamics of CD8 viral epitope processing, generation and T-cell responses, and how immunodominance counteracts viral immune escape mechanisms that develop in the context of emerging SARS-CoV-2 variants.


Subject(s)
Immunodominant Epitopes/immunology , SARS-CoV-2/immunology , T-Lymphocytes/immunology , Antigen Presentation , Cytosol/metabolism , Humans , Proteasome Endopeptidase Complex/physiology , Receptors, Antigen, T-Cell/genetics , Receptors, Antigen, T-Cell/physiology , T-Lymphocytes, Cytotoxic/immunology
20.
BMC Bioinformatics ; 21(Suppl 17): 484, 2020 Dec 14.
Article in English | MEDLINE | ID: covidwho-1388725

ABSTRACT

BACKGROUND: We previously introduced PCPS (Proteasome Cleavage Prediction Server), a web-based tool to predict proteasome cleavage sites using n-grams. Here, we evaluated the ability of PCPS immunoproteasome cleavage model to discriminate CD8+ T cell epitopes. RESULTS: We first assembled an epitope dataset consisting of 844 unique virus-specific CD8+ T cell epitopes and their source proteins. We then analyzed cleavage predictions by PCPS immunoproteasome cleavage model on this dataset and compared them with those provided by a related method implemented by NetChop web server. PCPS was clearly superior to NetChop in term of sensitivity (0.89 vs. 0.79) but somewhat inferior with regard to specificity (0.55 vs. 0.60). Judging by the Mathew's Correlation Coefficient, PCPS predictions were overall superior to those provided by NetChop (0.46 vs. 0.39). We next analyzed the power of C-terminal cleavage predictions provided by the same PCPS model to discriminate CD8+ T cell epitopes, finding that they could be discriminated from random peptides with an accuracy of 0.74. Following these results, we tuned the PCPS web server to predict CD8+ T cell epitopes and predicted the entire SARS-CoV-2 epitope space. CONCLUSIONS: We report an improved version of PCPS named iPCPS for predicting proteasome cleavage sites and peptides with CD8+ T cell epitope features. iPCPS is available for free public use at https://imed.med.ucm.es/Tools/pcps/ .


Subject(s)
Epitopes, T-Lymphocyte , Proteasome Endopeptidase Complex/metabolism , Proteomics/methods , SARS-CoV-2 , Viral Proteins , COVID-19/virology , Epitopes, T-Lymphocyte/chemistry , Epitopes, T-Lymphocyte/metabolism , Humans , Peptides/chemistry , Peptides/metabolism , SARS-CoV-2/chemistry , SARS-CoV-2/metabolism , Software , Viral Proteins/chemistry , Viral Proteins/metabolism
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