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Identification of major histocompatibility complex class I C molecule as an attachment factor that facilitates coronavirus HKU1 spike-mediated infection.

ABSTRACT: Human coronavirus HKU1 (HCoV-HKU1) is a recently discovered human coronavirus associated with respiratory tract infections worldwide. In this study, we have identified the major histocompatibility complex class I C molecule (HLA-C) as an attachment factor in facilitating HCoV-HKU1 spike (S)-mediated infection. HCoV-HKU1 S pseudotyped virus was assembled using a human immunodeficiency virus type 1-derived reporter virus harboring the human codon-optimized spike of HCoV-HKU1. We identified human alveolar epithelial A549 cells as the most susceptible cell line among those tested to infection by HCoV-HKU1 S pseudotypes. A549 cells were shown to bind purified soluble HCoV-HKU1 S(1-600) glycopeptide. To search for the functional receptor for HCoV-HKU1, an A549 cDNA expression library was constructed and transduced into the nonpermissive, baby hamster kidney cells line BHK-21. Transduced cells that bind soluble HCoV-HKU1 S(1-600) glycoprotein with C-terminal FLAG were sorted. Sequencing of two independent clones revealed cDNA inserts encoding HLA-C. Inhibition of HLA-C expression or function by RNAi silencing and anti-HLA-C antibody decreased HCoV-HKU1 S pseudotyped virus infection of A549 cells by 62 to 65%, whereas pretreatment of cells with neuraminidase decreased such infection by only 13%. When HLA-C was constitutively expressed in another nonpermissive cell line, NIH-3T3, quantitative PCR showed that the binding of HCoV-HKU1 S pseudotyped virus to cell surfaces was increased by 200-fold, but the cells remained nonsusceptible to HCoV-HKU1 S pseudotyped virus infection. Our data suggest that HLA-C is involved in the attachment of HCoV-HKU1 to A549 cells and is a potential candidate to facilitate cell entry. However, other unknown surface proteins on A549 cells may be concomitantly utilized by S glycoprotein of HCoV-HKU1 during viral entry. Further studies are required to elucidate other putative receptors or coreceptors for HCoV-HKU1 and the mechanism of HCoV-HKU1 S-mediated cell entry.

SUBMITTER: Chan CM

PROVIDER: S-EPMC2612401 | biostudies-literature | 2009 Jan

REPOSITORIES: biostudies-literature

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Identification of major histocompatibility complex class I C molecule as an attachment factor that facilitates coronavirus HKU1 spike-mediated infection.

Chan Che Man CM Lau Susanna K P SK Woo Patrick C Y PC Tse Herman H Zheng Bo-Jian BJ Chen Ling L Huang Jian-Dong JD Yuen Kwok-Yung KY

Journal of virology 20081105 2

Human coronavirus HKU1 (HCoV-HKU1) is a recently discovered human coronavirus associated with respiratory tract infections worldwide. In this study, we have identified the major histocompatibility complex class I C molecule (HLA-C) as an attachment factor in facilitating HCoV-HKU1 spike (S)-mediated infection. HCoV-HKU1 S pseudotyped virus was assembled using a human immunodeficiency virus type 1-derived reporter virus harboring the human codon-optimized spike of HCoV-HKU1. We identified human a ...[more]

PMID: 18987136

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Project description:Sirenians share with cetaceans and pinnipeds several convergent traits selected for the aquatic lifestyle. Living in water poses new challenges not only for locomotion and feeding but also for combating new pathogens, which may render the immune system one of the best tools aquatic mammals have for dealing with aquatic microbial threats. So far, only cetaceans have had their class II Major Histocompatibility Complex (MHC) organization characterized, despite the importance of MHC genes for adaptive immune responses. This study aims to characterize the organization of the marine mammal class II MHC using publicly available genomes. We located class II sequences in the genomes of one sirenian, four pinnipeds and eight cetaceans using NCBI-BLAST and reannotated the sequences using local BLAST search with exon and intron libraries. Scaffolds containing class II sequences were compared using dotplot analysis and introns were used for phylogenetic analysis. The manatee class II region shares overall synteny with other mammals, however most DR loci were translocated from the canonical location, past the extended class II region. Detailed analysis of the genomes of closely related taxa revealed that this presumed translocation is shared with all other living afrotherians. Other presumptive chromosome rearrangements in Afrotheria are the deletion of DQ loci in Afrosoricida and deletion of DP in E. telfairi. Pinnipeds share the main features of dog MHC: lack of a functional pair of DPA/DPB genes and inverted DRB locus between DQ and DO subregions. All cetaceans share the Cetartiodactyla inversion separating class II genes into two subregions: class IIa, with DR and DQ genes, and class IIb, with non-classic genes and a DRB pseudogene. These results point to three distinct and unheralded class II MHC structures in marine mammals: one canonical organization but lacking DP genes in pinnipeds; one bearing an inversion separating IIa and IIb subregions lacking DP genes found in cetaceans; and one with a translocation separating the most diverse class II gene from the MHC found in afrotherians and presumptive functional DR, DQ, and DP genes. Future functional research will reveal how these aquatic mammals cope with pathogen pressures with these divergent MHC organizations.

Project description:BACKGROUND:CD4+ T cells play an important role in atherosclerosis, but their antigen specificity is poorly understood. Immunization with apolipoprotein B (ApoB, core protein of low density lipoprotein) is known to be atheroprotective in animal models. Here, we report on a human APOB peptide, p18, that is sequence-identical in mouse ApoB and binds to both mouse and human major histocompatibility complex class II molecules. METHODS:We constructed p18 tetramers to detect human and mouse APOB-specific T cells and assayed their phenotype by flow cytometry including CD4 lineage transcription factors, intracellular cytokines, and T cell receptor activation. Apolipoprotein E-deficient ( Apoe-/-) mice were vaccinated with p18 peptide or adjuvants alone, and atherosclerotic burden in the aorta was determined. RESULTS:In human peripheral blood mononuclear cells from donors without cardiovascular disease, p18 specific CD4+ T cells detected by a new human leukocyte antigen-antigen D related-p18 tetramers were mostly Foxp3+ regulatory T cells (Tregs). Donors with subclinical cardiovascular disease as detected by carotid artery ultrasound had Tregs coexpressing retinoic acid-related orphan receptor gamma t or T-bet, which were both almost absent in donors without cardiovascular disease. In Apoe-/- mice, immunization with p18 induced Tregs and reduced atherosclerotic lesions. After peptide restimulation, responding CD4+ T cells identified by Nur77-GFP (green fluorescent protein) were highly enriched in Tregs. A new mouse I-Ab-p18 tetramer identified the expansion of p18-specific CD4+ T cells on vaccination, which were enriched for interleukin-10-producing Tregs. CONCLUSIONS:These findings show that APOB p18-specific CD4+ T cells are mainly Tregs in healthy donors, but coexpress other CD4 lineage transcription factors in donors with subclinical cardiovascular disease. This study identifies ApoB peptide 18 as the first Treg epitope in human and mouse atherosclerosis.

Dataset Information

Identification of major histocompatibility complex class I C molecule as an attachment factor that facilitates coronavirus HKU1 spike-mediated infection.

Publications

Identification of major histocompatibility complex class I C molecule as an attachment factor that facilitates coronavirus HKU1 spike-mediated infection.

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