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Histo-blood group antigens of glycosphingolipids predict susceptibility of human intestinal enteroids to norovirus infection
Department of Laboratory Medicine, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden; Department of Microbiology and Immunology, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi. Umeå universitet, Medicinska fakulteten, Wallenberg centrum för molekylär medicin vid Umeå universitet (WCMM). (Marta Bally)
Wallenberg Laboratory, Department of Molecular and Clinical Medicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas, USA.
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2020 (Engelska)Ingår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 295, nr 47, s. 15974-15987Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

The molecular mechanisms behind infection and propagation of human restricted pathogens such as human norovirus (HuNoV) have defied interrogation because they were previously unculturable. However, human intestinal enteroids (HIEs) have emerged to offer unique ex vivo models for targeted studies of intestinal biology, including inflammatory and infectious diseases. Carbohydrate-dependent histo-blood group antigens (HBGAs) are known to be critical for clinical infection. To explore whether HBGAs of glycosphingolipids contribute to HuNoV infection, we obtained HIE cultures established from stem cells isolated from jejunal biopsies of six individuals with different ABO, Lewis, and secretor genotypes. We analyzed their glycerolipid and sphingolipid compositions and quantified interaction kinetics and the affinity of HuNoV virus-like particles (VLPs) to lipid vesicles produced from the individual HIE-lipid extracts. All HIEs had a similar lipid and glycerolipid composition. Sphingolipids included HBGA-related type 1 chain glycosphingolipids (GSLs), with HBGA epitopes corresponding to the geno- and phenotypes of the different HIEs. As revealed by single-particle interaction studies of Sydney GII.4 VLPs with glycosphingolipid-containing HIE membranes, both binding kinetics and affinities explain the patterns of susceptibility toward GII.4 infection for individual HIEs. This is the first time norovirus VLPs have been shown to interact specifically with secretor gene-dependent GSLs embedded in lipid membranes of HIEs that propagate GII.4 HuNoV ex vivo, highlighting the potential of HIEs for advanced future studies of intestinal glycobiology and host-pathogen interactions.

Ort, förlag, år, upplaga, sidor
Elsevier, 2020. Vol. 295, nr 47, s. 15974-15987
Nyckelord [en]
ceramide, fucosyltransferase, glycerophospholipid, glycolipid structure, glycosphingolipid, glycosylation, host-pathogen interaction, human norovirus, intestinal epithelium, intestine, plus-stranded RNA virus, sphingolipid, sphingomyelin, viral replication, virus entry
Nationell ämneskategori
Mikrobiologi inom det medicinska området
Identifikatorer
URN: urn:nbn:se:umu:diva-176980DOI: 10.1074/jbc.RA120.014855ISI: 000595657900016PubMedID: 32913124Scopus ID: 2-s2.0-85096814513OAI: oai:DiVA.org:umu-176980DiVA, id: diva2:1503036
Forskningsfinansiär
Vetenskapsrådet, K2014-68X-08266-27-4Vetenskapsrådet, 2017-00955Vetenskapsrådet, 2017-04029Stiftelsen för strategisk forskning (SSF), SB12/KF10-0088WallenbergstiftelsernaTillgänglig från: 2020-11-23 Skapad: 2020-11-23 Senast uppdaterad: 2023-08-21Bibliografiskt granskad
Ingår i avhandling
1. Probing and elucidating the dynamics of virus-membrane interaction via plasma membrane mimics
Öppna denna publikation i ny flik eller fönster >>Probing and elucidating the dynamics of virus-membrane interaction via plasma membrane mimics
2023 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Alternativ titel[sv]
Undersökande av dynamiken i interaktioner mellan virus och membran genom en modell för plasmamembra
Abstract [en]

Virus infection is initiated by the attachment of a virion to a susceptible cell’s plasma membrane, in a highly dynamic and well-orchestrated process that encompasses various steps and engages numerous viral and cellular factors. These dynamic steps may include initial non-specific binding to ubiquitous cell-membrane ligands, diffusion across the membrane to a suitable entry site and virus engagement with various receptors and co-receptors on the cell surface. Molecules and processes involved may vary across virus species, but it is likely that in all cases the dynamics of virus-membrane interactions need to be carefully fine-tuned to optimize the entry process. Nevertheless, the investigation and characterization of the involved biomolecular interactions are often oversimplified to isolated virus-receptor pairs engagement, ignoring the complexity of the membrane and the dynamic behaviors of the interaction. This doctoral thesis aims to shed light on this critical sphere of virus-membrane interactions, focusing on how viruses dynamically engage with plasma membrane molecules to successfully infect the cell. To facilitate this research, we utilized an innovative approach of using plasma membrane mimics to explore how viruses dynamically interact with the plasma membrane across several chosen contexts.

Central to this project was the development and optimization of cell membrane mimics consisting of supported lipid bilayers (SLBs) reflecting the compositional complexity of the membrane. In the first paper, we developed a method to quantify the membrane material in a lipid vesicle sample in terms of total membrane surface area. This method is vital for the full utilization of our membrane mimics, which in many cases require the mixing of different vesicles in specific ratios.

Subsequently, cell membrane mimics of complex composition were used to investigate the molecular mechanisms modulating virus-membrane interactions in several chosen contexts. These investigations relied primarily on total internal reflection fluorescent microscopy to characterize the attachment and detachment behavior of individual virus particles from the membrane. Firstly, studies focused on norovirus, a pathogen that infects cells in the gastrointestinal tract. The virus is known to bind to histo-blood group antigens (HGBA), specific glycans found on intestinal epithelial cells membranes. However, susceptibility to norovirus infection varies between individuals, and the difference is correlated to the specific glycan expression of the intestinal cells. This suggests that the differences in susceptibility might be related to differences in virus-membrane interaction dynamics. Using membranes constructed from lipids extracted from human intestinal enteroids (HIE) derived from susceptible and non-susceptible individuals, it was determined that norovirus associates similarly to both susceptible and non-susceptible membranes but dissociates slower from susceptible membranes. Using native supported lipid bilayers (nSLBs), bilayers derived from plasma membrane extracts of the HIEs, we then investigated the contribution of different carbohydrate moieties to interaction kinetics. This investigation revealed that virus binding to fucose residues on HBGAs is only part of the interaction, and that the virus also binds to sialic acid to a similar degree. It was also found that binding occurs primarily to membrane glycoproteins, and not membrane glycolipids.

nSLBs were further found to be highly useful complements to virological investigations in a number of contexts. First, we studied the effect of isoform 4 of apolipoprotein E (ApoE4) on herpes simplex virus 1 (HSV-1) binding kinetics to plasma membrane SLBs. ApoE4 is a lipid binding protein that has been found, in conjunction with HSV-1, to be a risk factor for Alzheimer's disease. We showed that membrane-bound ApoE4 does not affect HSV-1 binding kinetics, but that viruses coated with ApoE4 demonstrate faster dissociation from susceptible membranes than non-coated viruses, indicating that the protein facilitates the release of new virions from the infected cell. This provides a mechanistic understanding of the overall pro-viral effect of ApoE, observed in infection experiments.

Second, nSLBs from respiratory epithelial cells were used to quantify binding of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to the plasma membrane. SARS-CoV-2 has caused one of the largest pandemic in modern history and the virus has shown the ability to rapidly mutate, causing periodic surges of new cases, with newer strains spreading more easily. These mutations have often been linked to the viral spike glycoprotein, responsible for viral attachment and entry. Investigations using spike-decorated liposomes as virus-mimetics, revealed that virus-membrane interaction dynamics vary for different variants of concerns. Specifically, the late Omicron variant, a highly transmissible variant, shows significantly increased affinity to susceptible membranes. This increased affinity was primarily due to increased association to the membrane. Experiments also showed that membrane-bound heparan sulfate has an inhibiting effect on virus binding to ACE2, for earlier variants.

In summary, we have successfully implemented membrane mimics with different levels of complexity to investigate virus-membrane interactions. This thesis demonstrates their potential in virology research in several contexts, including measuring the avidity of viruses to membranes, evaluating the relative contributions of different attachment factors to kinetics, and the influence of viral and cellular factors on binding.

Ort, förlag, år, upplaga, sidor
Umeå: Umeå University, 2023. s. 67
Serie
Umeå University medical dissertations, ISSN 0346-6612 ; 2255
Nyckelord
Plasma membrane, supported lipid bilayer, binding kinetics, equilibrium fluctuation analysis, TIRF, FRET, norovirus, HSV, SARS-CoV-2, histo-blood group antigens, ApoE, heparan sulfate
Nationell ämneskategori
Mikrobiologi
Forskningsämne
mikrobiologi
Identifikatorer
urn:nbn:se:umu:diva-213026 (URN)978-91-8070-148-8 (ISBN)978-91-8070-149-5 (ISBN)
Disputation
2023-09-08, 6E-E04, byggnad 6E, 13:00 (Engelska)
Opponent
Handledare
Anmärkning

In the digital version wrongly stated UmU Print Service, Umeå University as printer. The right information is: "Printed by: Cityprint i Norr AB". 

Tillgänglig från: 2023-08-23 Skapad: 2023-08-18 Senast uppdaterad: 2023-11-28Bibliografiskt granskad

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