Abstract Title
Surface Glycans as Therapeutic Targets to Prevent Human Norovirus and Rotavirus Infections in a Human Intestinal Enteroid Model
Presenter
Stefan Taube, University of Lübeck
Co-Author(s)
Maryna Chaika1, Alexander Müller1, Sonja Jacobsen2, Sandra Niendorf2, Mario Pieper3, Peter Koenig3, Carmen Mirabelli1, Christiane Wobus4, Stefan Taube1
1Institute of Virology and Cell Biology, University of Luebeck, Germany
2Unit Gastroenteritis and Hepatitis Viruses and Enteroviruses, Department of Infectious Diseases, Robert Koch Institute, Berlin, Germany
3Institute of Anatomy, University of Luebeck, Germany
4Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, MI, USA
Abstract Category
Entry and Replication
Abstract
Human noroviruses (HNoVs) and human rotaviruses (HRoVs) are RNA viruses that cause a significant number of acute gastroenteritis cases worldwide. Both viruses exploit glycans on the surface of susceptible cells, such as histo-blood group antigens (HBGAs) and gangliosides, which serve as entry factors and are essential for infection. Human intestinal enteroids (HIEs), a stem cell dervied 3D-culture system, was described as a model for studying enteric pathogens such as HuNoV and HRoV. This model closely resembles human intestinal lining, it’s luminal polarity, and cell composition, making it highly suitable tool for infection studies. Here, we have adapted an infant ileum-derived HIE model to explore the role of specific surface glycans with fucose, sialic acid, galactose and mannose moieties, for successful infection of HIEs with HNoV, HRoV, and influenza A virus (IAV).
To study the role of glycans, lectins were employed to compete for binding to glycans on the cell surface. HIEs were incubated with increasing concentrations of lectins including UEA-1 (fucose binding), WGA (galactose and sialic acid binding), ConA (mannose binding), or SNA (sialic acid binding) and subsequently infected with HNoV, HRoV or IAV (H1N1). Surface glycan composition was modulated using a broad fucosyltransferase inhibitor (2-fluoro-peracetyl-fucose, 2-FPF) and monitored using fluorescently labeled lectins visualized by 2-photon microscopy. Viral replication was measured 2 h and 2 d post-infection using RT-qPCR or plaque assay.
Fucosyltransferase inhibitor abrogated HNoV and HRoV infection in HIEs, whereas IAV H1N1 infection remained unaffected. UEA-1 and WGA effectively blocked HNoV, while ConA, SNA and WGA reduced IAV H1N1 infection. HRoV infection was not affected by any of the tested lectins in HIEs. To assess the potential competitive effect of fucose, HIEs were infected with HNoV in the presence of 50 mM free fucose. Unexpectedly, fucose supplementation markedly enhanced HNoV infection (3- to 10-fold, depending on tested HNoV stool-isolate) and enabled limited viral passaging in HIEs.
Inhibition of fucosyltransferase activity in HIEs confirmed importance of fucosysalted residues for HNoV and HRoV infection. While lectin competition corroborated significance of H type (UEA-1) and N-acetylgalactosamine-containing (WGA) glycans for HNoV infection and revealed non-significance of above mentioned residues for HRoV infection. Notably, exogenous fucose may function beyond a receptor mimicry, actively enhancing HNoV infection. Our findings highlight the HIE model as robust system for studying role of glycan in virus infection and point to glycan-targeting strategy for a novel therapeutic approach to inhibit HNoV and HRoV infection.
To study the role of glycans, lectins were employed to compete for binding to glycans on the cell surface. HIEs were incubated with increasing concentrations of lectins including UEA-1 (fucose binding), WGA (galactose and sialic acid binding), ConA (mannose binding), or SNA (sialic acid binding) and subsequently infected with HNoV, HRoV or IAV (H1N1). Surface glycan composition was modulated using a broad fucosyltransferase inhibitor (2-fluoro-peracetyl-fucose, 2-FPF) and monitored using fluorescently labeled lectins visualized by 2-photon microscopy. Viral replication was measured 2 h and 2 d post-infection using RT-qPCR or plaque assay.
Fucosyltransferase inhibitor abrogated HNoV and HRoV infection in HIEs, whereas IAV H1N1 infection remained unaffected. UEA-1 and WGA effectively blocked HNoV, while ConA, SNA and WGA reduced IAV H1N1 infection. HRoV infection was not affected by any of the tested lectins in HIEs. To assess the potential competitive effect of fucose, HIEs were infected with HNoV in the presence of 50 mM free fucose. Unexpectedly, fucose supplementation markedly enhanced HNoV infection (3- to 10-fold, depending on tested HNoV stool-isolate) and enabled limited viral passaging in HIEs.
Inhibition of fucosyltransferase activity in HIEs confirmed importance of fucosysalted residues for HNoV and HRoV infection. While lectin competition corroborated significance of H type (UEA-1) and N-acetylgalactosamine-containing (WGA) glycans for HNoV infection and revealed non-significance of above mentioned residues for HRoV infection. Notably, exogenous fucose may function beyond a receptor mimicry, actively enhancing HNoV infection. Our findings highlight the HIE model as robust system for studying role of glycan in virus infection and point to glycan-targeting strategy for a novel therapeutic approach to inhibit HNoV and HRoV infection.