Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.17 (lysozyme)
 21,489 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The protease activities responsible for the cotranslational processing of the Semliki Forest virus structural polyprotein were investigated by using an in vitro transcription-translation system. Three cleavages released the individual chains from the nascent polyprotein in the order capsid, p62, 6K (a nonstructural peptide), and E1. We showed directly that the protease activity responsible for the release of the capsid protein resides in the capsid itself: by progressive truncation of the cDNA used for the SP6 transcription, we showed that a precursor containing as few as 38 residues of the p62 protein left at the C terminus of the capsid was still very efficiently cleaved in vitro. We further tested the possibility that serine-219 of the capsid is involved in autoproteolysis by site-directed in vitro mutagenesis. A change in the sequence Gly-Asp-Ser(219)-Gly, a tetrapeptide conserved among several animal serine proteases, to Gly-Asp-Arg-Ser-Thr was shown to completely abolish in vitro cleavage. This supports the notion that the capsid is a serine protease. The role of the capsid protease in the processing of the 6K junctions was then investigated by translations of a hybrid polyprotein in which the capsid and most of the p62 sequences are replaced by those of the secretory protein lysozyme. The cleavages and concomitant appearance of the 6K peptide occurred efficiently and were shown to require the presence of membranes. This demonstrates that the capsid protease is not required for those cleavages and suggests that a membrane-associated host protease is responsible for the cleavage.
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PMID:Processing of the Semliki Forest virus structural polyprotein: role of the capsid protease. 355 12

Astrocytes perform a variety of functions that are important for normal neuronal activity and recovery after brain injury. Because astrocytes are very vulnerable to H2O2, protection of astrocytes from oxidative damage in various neurological diseases is important in maintaining brain function and preventing brain damage. In this study, we investigated the characteristics and mechanisms of a specific imidazoline I2 receptor agonist 2-BFI-mediated cytoprotection using a rat astrocyte cultures of H2O2-exposed oxidative stress. Here we show that 2-BFI in H2O2-exposed astrocytes protects cell death through increased lysosomal membrane stability, LC3-II conversion, and subsequently suppresses accumulation of p62. These effects of 2-BFI were significantly reversed after treatment with the lysozyme activity inhibitor Bafilomycin A1. These results suggest that the cytoprotective effects of 2-BFI, which increases lysosomal stability in oxidative stress, may involve regulation of lysosomal-associated membrane protein-dependent autophagy and autolysosome degradation in astrocytes.
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PMID:Protective effect of the imidazoline I2 receptor agonist 2-BFI on oxidative cytotoxicity in astrocytes. 3014 55

CORM-3 is a water-soluble carbon monoxide (CO)-releasing molecule developed for possible therapeutic use of CO. CORM-3 belongs to a group of metal carbonyl compounds that contain transition metals and carbonyls as the central scaffold and coordinated ligands, respectively. CORM-3 has been reported to be reactive with many proteins in eukaryotes including mammals. Among them, several extracellular proteins, such as lysozyme, as well as plasma albumin and fibronectin, have been shown to interact directly with CORM-3. p62 is an intracellular adaptor protein required for targeting ubiquitinated (Ub) proteins to lysosomal degradation through autophagy. p62 has been shown to undergo self-oligomerization via covalent crosslinking in response to treatment with verteporfin, a benzoporphyrin derivative used for photodynamic therapy. Here we show that CORM-3 also interacts directly with p62. When applied to mouse embryonic fibroblasts (MEFs) at a high concentration (1 mM), CORM-3 causes the formation of reduction- and detergent-resistant high molecular weight (HMW)-p62. HMW-p62 accumulates more in atg5-/- MEFs than in wild type (WT) MEFs, showing the elimination of HMW-p62 through autophagy. HMW-p62 is also generated in H9c2 rat cardiomyoblastoma as well as A549 human alveolar epithelial cells, suggesting that HMW-p62 formation is not specific to MEFs, but, rather, is a general event in mammalian cells. HMW-p62 formation by CORM-3 can be reproduced using purified p62 in vitro, demonstrating the direct interaction between CORM-3 and p62. These results show that p62 is a CORM-3-interactive intracellular protein.
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PMID:Formation of high molecular weight p62 by CORM-3. 3062 Jul 62

Paneth cells are key epithelial cells that provide an antimicrobial barrier and maintain integrity of the small-intestinal stem cell niche. Paneth cell abnormalities are unfortunately detrimental to gut health and are often associated with digestive pathologies such as Crohn's disease or infections. Similar alterations are observed in individuals with impaired autophagy, a process that recycles cellular components. The direct effect of autophagy impairment on Paneth cells has not been analysed. To investigate this, we generated a mouse model lacking Atg16l1 specifically in intestinal epithelial cells, making these cells impaired in autophagy. Using three-dimensional intestinal organoids enriched for Paneth cells, we compared the proteomic profiles of wild-type and autophagy-impaired organoids. We used an integrated computational approach combining protein-protein interaction networks, autophagy-targeted proteins and functional information to identify the mechanistic link between autophagy impairment and disrupted pathways. Of the 284 altered proteins, 198 (70%) were more abundant in autophagy-impaired organoids, suggesting reduced protein degradation. Interestingly, these differentially abundant proteins comprised 116 proteins (41%) that are predicted targets of the selective autophagy proteins p62, LC3 and ATG16L1. Our integrative analysis revealed autophagy-mediated mechanisms that degrade key proteins in Paneth cell functions, such as exocytosis, apoptosis and DNA damage repair. Transcriptomic profiling of additional organoids confirmed that 90% of the observed changes upon autophagy alteration have effects at the protein level, not on gene expression. We performed further validation experiments showing differential lysozyme secretion, confirming our computationally inferred downregulation of exocytosis. Our observations could explain how protein-level alterations affect Paneth cell homeostatic functions upon autophagy impairment.This article has an associated First Person interview with the joint first authors of the paper.
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PMID:Integrative analysis of Paneth cell proteomic and transcriptomic data from intestinal organoids reveals functional processes dependent on autophagy. 3081 64

Staphylococcus aureus is a major human pathogen causing multiple pathologies, from cutaneous lesions to life-threatening sepsis. Although neutrophils contribute to immunity against S. aureus, multiple lines of evidence suggest that these phagocytes can provide an intracellular niche for staphylococcal dissemination. However, the mechanism of neutrophil subversion by intracellular S. aureus remains unknown. Targeting of intracellular pathogens by macroautophagy/autophagy is recognized as an important component of host innate immunity, but whether autophagy is beneficial or detrimental to S. aureus-infected hosts remains controversial. Here, using larval zebrafish, we showed that the autophagy marker Lc3 rapidly decorates S. aureus following engulfment by macrophages and neutrophils. Upon phagocytosis by neutrophils, Lc3-positive, non-acidified spacious phagosomes are formed. This response is dependent on phagocyte NADPH oxidase as both cyba/p22phox knockdown and diphenyleneiodonium (DPI) treatment inhibited Lc3 decoration of phagosomes. Importantly, NADPH oxidase inhibition diverted neutrophil S. aureus processing into tight acidified vesicles, which resulted in increased host resistance to the infection. Some intracellular bacteria within neutrophils were also tagged by Sqstm1/p62-GFP fusion protein and loss of Sqstm1 impaired host defense. Together, we have shown that intracellular handling of S. aureus by neutrophils is best explained by Lc3-associated phagocytosis (LAP), which appears to provide an intracellular niche for bacterial pathogenesis, while the selective autophagy receptor Sqstm1 is host-protective. The antagonistic roles of LAP and Sqstm1-mediated pathways in S. aureus-infected neutrophils may explain the conflicting reports relating to anti-staphylococcal autophagy and provide new insights for therapeutic strategies against antimicrobial-resistant Staphylococci.Abbreviations: ATG: autophagy related; CFU: colony-forming units; CMV: cytomegalovirus; Cyba/P22phox: cytochrome b-245, alpha polypeptide; DMSO: dimethyl sulfoxide; DPI: diphenyleneiodonium; EGFP: enhanced green fluorescent protein; GFP: green fluorescent protein; hpf: hours post-fertilization; hpi: hours post-infection; Irf8: interferon regulatory factor 8; LAP: LC3-associated phagocytosis; lyz: lysozyme; LWT: london wild type; Map1lc3/Lc3: microtubule-associated protein 1 light chain 3; NADPH oxidase: nicotinamide adenine dinucleotide phosphate oxidase; RFP: red fluorescent protein; ROS: reactive oxygen species; RT-PCR: reverse transcriptase polymerase chain reaction; Sqstm1/p62: sequestosome 1; Tg: transgenic; TSA: tyramide signal amplification.
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PMID:The autophagic response to Staphylococcus aureus provides an intracellular niche in neutrophils. 3217 46