title: Overexpression of the alpha-2,6-sialyltransferase in MDCK cells increases influenza virus sensitivity to neuraminidase inhibitors.
abstract: No reliable cell culture assay is currently available for monitoring human influenza virus sensitivity to neuraminidase inhibitors (NAI). This can be explained by the observation that because of a low concentration of sialyl-alpha2,6-galactose (Sia[alpha2,6]Gal)-containing virus receptors in conventional cell lines, replication of human virus isolates shows little dependency on viral neuraminidase. To test whether overexpression of Sia(alpha2,6)Gal moieties in cultured cells could make them suitable for testing human influenza virus sensitivity to NAI, we stably transfected MDCK cells with cDNA of human 2,6-sialyltransferase (SIAT1). Transfected cells expressed twofold-higher amounts of 6-linked sialic acids and twofold-lower amounts of 3-linked sialic acids than parent MDCK cells as judged by staining with Sambucus nigra agglutinin and Maackia amurensis agglutinin, respectively. After transfection, binding of a clinical human influenza virus isolate was increased, whereas binding of its egg-adapted variant which preferentially bound 3-linked receptors was decreased. The sensitivity of human influenza A and B viruses to the neuraminidase inhibitor oseltamivir carboxylate was substantially improved in the SIAT1-transfected cell line and was consistent with their sensitivity in neuraminidase enzyme assay and with the hemagglutinin (HA) receptor-binding phenotype. MDCK cells stably transfected with SIAT1 may therefore be a suitable system for testing influenza virus sensitivity to NAI.
authors.affiliation: Institute of Virology, Philipps University, 35037 Marburg, Germany. mikhail.matrosovich@med.uni-marburg.de
authors.forename: Mikhail
authors.initials: M
authors.lastname: Matrosovich
authors.forename: Tatyana
authors.initials: T
authors.lastname: Matrosovich
authors.forename: Jackie
authors.initials: J
authors.lastname: Carr
authors.forename: Noel A
authors.initials: NA
authors.lastname: Roberts
authors.forename: Hans-Dieter
authors.initials: HD
authors.lastname: Klenk
mesh_terms: D000081:Acetamides / Q000494:pharmacology*; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D009980:Influenza A virus / Q000187:drug effects* / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D009439:Neuraminidase / Q000037:antagonists & inhibitors*; D053139:Oseltamivir; D012794:Sialic Acids / Q000378:metabolism; D012799:Sialyltransferases / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase
chemical_list: D000081:Acetamides; D000998:Antiviral Agents; D004791:Enzyme Inhibitors; D012794:Sialic Acids; D053139:Oseltamivir; D012799:Sialyltransferases; D009439:Neuraminidase; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase