Glycosylated extracellular mucin domains protect against SARS-CoV-2 infection at the respiratory surface.

Mucins play an essential role in protecting the respiratory tract against microbial infections while also acting as binding sites for bacterial and viral adhesins. The heavily O-glycosylated gel-forming mucins MUC5AC and MUC5B eliminate pathogens by mucociliary clearance. Transmembrane mucins MUC1, MUC4, and MUC16 can restrict microbial invasion at the apical surface of the epithelium. In this study, we determined the impact of host mucins and mucin glycans on epithelial entry of SARS-CoV-2. Human lung epithelial Calu-3 cells express the SARS-CoV-2 entry receptor ACE2 and high levels of glycosylated MUC1, but not MUC4 and MUC16, on their cell surface. The O-glycan-specific mucinase StcE specifically removed the glycosylated part of the MUC1 extracellular domain while leaving the underlying SEA domain and cytoplasmic tail intact. StcE treatment of Calu-3 cells significantly enhanced infection with SARS-CoV-2 pseudovirus and authentic virus, while removal of terminal mucin glycans sialic acid and fucose from the epithelial surface did not impact viral entry. In Calu-3 cells, the transmembrane mucin MUC1 and ACE2 are located to the apical surface in close proximity and StcE treatment results in enhanced binding of purified spike protein. Both MUC1 and MUC16 are expressed on the surface of human organoid-derived air-liquid interface (ALI) differentiated airway cultures and StcE treatment led to mucin removal and increased levels of SARS-CoV-2 replication. In these cultures, MUC1 was highly expressed in non-ciliated cells while MUC16 was enriched in goblet cells. In conclusion, the glycosylated extracellular domains of different transmembrane mucins might have similar protective functions in different respiratory cell types by restricting SARS-CoV-2 binding and entry.

Campylobacter jejuni permeabilizes the host cell membrane by short chain lysophosphatidylethanolamines.

Lysophospholipids (LPLs) are crucial for regulating epithelial integrity and homeostasis in eukaryotes, however the effects of LPLs produced by bacteria on host cells is largely unknown. The membrane of the human bacterial pathogen Campylobacter jejuni is rich in LPLs. Although C. jejuni possesses several virulence factors, it lacks traditional virulence factors like type III secretion systems, present in most enteropathogens. Here, we provide evidence that membrane lipids lysophosphatidylethanolamines (lysoPEs) of C. jejuni are able to lyse erythrocytes and are toxic for HeLa and Caco-2 cells. Lactate dehydrogenase (LDH) release assays and confocal microscopy revealed that lysoPE permeabilizes the cells. LysoPE toxicity was partially rescued by oxidative stress inhibitors, indicating that intracellular reactive oxygen species may contribute to the cell damage. Our results show that especially the short-chain lysoPEs (C:14) which is abundantly present in the C. jejuni membrane may be considered as a novel virulence factor.

Human-type sialic acid receptors contribute to avian influenza A virus binding and entry by hetero-multivalent interactions.

Establishment of zoonotic viruses, causing pandemics like the Spanish flu and Covid-19, requires adaptation to human receptors. Pandemic influenza A viruses (IAV) that crossed the avian-human species barrier switched from binding avian-type α2-3-linked sialic acid (2-3Sia) to human-type 2-6Sia receptors. Here, we show that this specificity switch is however less dichotomous as generally assumed. Binding and entry specificity were compared using mixed synthetic glycan gradients of 2-3Sia and 2-6Sia and by employing a genetically remodeled Sia repertoire on the surface of a Sia-free cell line and on a sialoglycoprotein secreted from these cells. Expression of a range of (mixed) 2-3Sia and 2-6Sia densities shows that non-binding human-type receptors efficiently enhanced avian IAV binding and entry provided the presence of a low density of high affinity avian-type receptors, and vice versa. Considering the heterogeneity of sialoglycan receptors encountered in vivo, hetero-multivalent binding is physiologically relevant and will impact evolutionary pathways leading to host adaptation.