Ehrlichia chaffeensis Uses Its Surface Protein EtpE to Bind GPI-Anchored Protein DNase X and Trigger Entry into Mammalian Cells

PLoS Pathogens

Marek Los¹  Koshiro Miura²  Mingqun Lin²  Dipu Mohan Kumar²  Mamoru Yamaguchi²  Yasuko Rikihisa²  Johannes F. Coy³ 

[1] Department of Clinical & Experimental Medicine, Integrative Regenerative Medical Center Linköping University, Linkoping, Sweden;Department of Veterinary Biosciences, College of Veterinary Medicine, The Ohio State University, Columbus, Ohio, United States of America;Vorstand/CSO TAVARLIN AG, Darmstadt, Germany

关键词: Deoxyribonucleases;    Host cells;    Cell binding;    Macrophages;    Ehrlichia;    Antibodies;    Blood;    Immunofluorescence;   

DOI  :  10.1371/journal.ppat.1003666

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

Ehrlichia chaffeensis, an obligatory intracellular rickettsial pathogen, enters and replicates in monocytes/macrophages and several non-phagocytic cells. E. chaffeensis entry into mammalian cells is essential not only for causing the emerging zoonosis, human monocytic ehrlichiosis, but also for its survival. It remains unclear if E. chaffeensis has evolved a specific surface protein that functions as an ‘invasin’ to mediate its entry. We report a novel entry triggering protein of Ehrlichia, EtpE that functions as an invasin. EtpE is an outer membrane protein and an antibody against EtpE (the C-terminal fragment, EtpE-C) greatly inhibited E. chaffeensis binding, entry and infection of both phagocytes and non-phagocytes. EtpE-C-immunization of mice significantly inhibited E. chaffeensis infection. EtpE-C-coated latex beads, used to investigate whether EtpE-C can mediate cell invasion, entered both phagocytes and non-phagocytes and the entry was blocked by compounds that block E. chaffeensis entry. None of these compounds blocked uptake of non-coated beads by phagocytes. Yeast two-hybrid screening revealed that DNase X, a glycosylphosphatidyl inositol-anchored mammalian cell-surface protein binds EtpE-C. This was confirmed by far-Western blotting, affinity pull-down, co-immunoprecipitation, immunofluorescence labeling, and live-cell image analysis. EtpE-C-coated beads entered bone marrow-derived macrophages (BMDMs) from wild-type mice, whereas they neither bound nor entered BMDMs from DNase X-/- mice. Antibody against DNase X or DNase X knock-down by small interfering RNA impaired E. chaffeensis binding, entry, and infection. E. chaffeensis entry and infection rates of BMDMs from DNase X-/- mice and bacterial load in the peripheral blood in experimentally infected DNase X-/- mice, were significantly lower than those from wild-type mice. Thus this obligatory intracellular pathogen evolved a unique protein EtpE that binds DNase X to enter and infect eukaryotic cells. This study is the first to demonstrate the invasin and its mammalian receptor, and their in vivo relevance in any ehrlichial species.

【 授权许可】

CC BY