A CRISPR Screen Using Subtilase Cytotoxin Identifies SLC39A9 as a Glycan-Regulating Factor
Toshiyuki Yamaji,
Hisatoshi Hanamatsu,
Tsuyoshi Sekizuka,
Makoto Kuroda,
Norimasa Iwasaki,
Makoto Ohnishi,
Jun-ichi Furukawa,
Kinnosuke Yahiro,
Kentaro Hanada
Affiliations
Toshiyuki Yamaji
Department of Biochemistry and Cell Biology, National Institute of Infectious Diseases, Shinjuku-ku, Tokyo 162-8640, Japan; Corresponding author
Hisatoshi Hanamatsu
Department of Advanced Clinical Glycobiology, Faculty of Medicine and Graduate School of Medicine, Hokkaido University, Sapporo 001-0021, Japan; Department of Gastroenterology and Hepatology, Graduate School of Medicine, Hokkaido University, Sapporo 060-8638, Japan
Tsuyoshi Sekizuka
Pathogen Genomics Center, National Institute of Infectious Diseases, Shinjuku-ku, Tokyo 162-8640, Japan
Makoto Kuroda
Pathogen Genomics Center, National Institute of Infectious Diseases, Shinjuku-ku, Tokyo 162-8640, Japan
Norimasa Iwasaki
Department of Advanced Clinical Glycobiology, Faculty of Medicine and Graduate School of Medicine, Hokkaido University, Sapporo 001-0021, Japan; Department of Orthopaedic Surgery, Graduate School of Medicine, Hokkaido University, Sapporo 060-8638, Japan
Makoto Ohnishi
Department of Bacteriology I, National Institute of Infectious Diseases, Shinjuku-ku, Tokyo 162-8640, Japan
Jun-ichi Furukawa
Department of Advanced Clinical Glycobiology, Faculty of Medicine and Graduate School of Medicine, Hokkaido University, Sapporo 001-0021, Japan
Kinnosuke Yahiro
Department of Molecular Infectiology, Graduate School of Medicine, Chiba University, Chiba 260-8670, Japan; Corresponding author
Kentaro Hanada
Department of Biochemistry and Cell Biology, National Institute of Infectious Diseases, Shinjuku-ku, Tokyo 162-8640, Japan
Summary: Subtilase cytotoxin (SubAB) is a virulence factor produced by locus of enterocyte effacement-negative Shiga-toxigenic Escherichia coli strains. The toxin recognizes sialoglycans for entry and cleaves an endoplasmic reticulum chaperon, binding immunoglobulin protein, to cause cell death. However, no systematic screening has yet been performed to identify critical host factors. Here, we performed a genome-wide CRISPR/Cas9 knockout screen for SubAB-induced cell death and identified various sialoglycan-related and membrane-trafficking genes. Analysis of glycan-deficient cells demonstrated that not only N-glycans but also O-glycans serve as SubAB receptors. In addition, SLC39A9, which is a predicted zinc transporter, as well as KDELRs and JTB, were required for SubAB to induce maximal cell death. Disruption of the SLC39A9 gene markedly reduced both complex-type N-glycans and core 1 O-glycans, and the O-glycan reduction was attributed to the reduction of core 1 synthase (C1GalT1). These results provide insights into the post-transcriptional regulation of glycosyltransferases by SLC39A9, as well as sialoglycan species as SubAB receptors. : Biological Sciences; Biochemistry; Molecular Biology; Microbiology; Cell Biology Subject Areas: Biological Sciences, Biochemistry, Molecular Biology, Microbiology, Cell Biology
