title: Simian hemorrhagic fever virus cell entry is dependent on CD163 and uses a clathrin-mediated endocytosis-like pathway.
abstract: Simian hemorrhagic fever virus (SHFV) causes a severe and almost uniformly fatal viral hemorrhagic fever in Asian macaques but is thought to be nonpathogenic for humans. To date, the SHFV life cycle is almost completely uncharacterized on the molecular level. Here, we describe the first steps of the SHFV life cycle. Our experiments indicate that SHFV enters target cells by low-pH-dependent endocytosis. Dynamin inhibitors, chlorpromazine, methyl-β-cyclodextrin, chloroquine, and concanamycin A dramatically reduced SHFV entry efficiency, whereas the macropinocytosis inhibitors EIPA, blebbistatin, and wortmannin and the caveolin-mediated endocytosis inhibitors nystatin and filipin III had no effect. Furthermore, overexpression and knockout study and electron microscopy results indicate that SHFV entry occurs by a dynamin-dependent clathrin-mediated endocytosis-like pathway. Experiments utilizing latrunculin B, cytochalasin B, and cytochalasin D indicate that SHFV does not hijack the actin polymerization pathway. Treatment of target cells with proteases (proteinase K, papain, α-chymotrypsin, and trypsin) abrogated entry, indicating that the SHFV cell surface receptor is a protein. Phospholipases A2 and D had no effect on SHFV entry. Finally, treatment of cells with antibodies targeting CD163, a cell surface molecule identified as an entry factor for the SHFV-related porcine reproductive and respiratory syndrome virus, diminished SHFV replication, identifying CD163 as an important SHFV entry component.
  
  
  OBJECTIVE
  Simian hemorrhagic fever virus (SHFV) causes highly lethal disease in Asian macaques resembling human illness caused by Ebola or Lassa virus. However, little is known about SHFV's ecology and molecular biology and the mechanism by which it causes disease. The results of this study shed light on how SHFV enters its target cells. Using electron microscopy and inhibitors for various cellular pathways, we demonstrate that SHFV invades cells by low-pH-dependent, actin-independent endocytosis, likely with the help of a cellular surface protein.
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA.
authors.forename: Yíngyún
authors.identifier: http://orcid.org/0000-0002-1101-0920
authors.initials: Y
authors.lastname: Caì
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA.
authors.forename: Elena N
authors.identifier: http://orcid.org/0000-0002-0275-2148
authors.initials: EN
authors.lastname: Postnikova
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA.
authors.forename: John G
authors.identifier: http://orcid.org/0000-0002-1529-8191
authors.initials: JG
authors.lastname: Bernbaum
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA.
authors.forename: Shu Qìng
authors.identifier: http://orcid.org/0000-0002-9473-566X
authors.initials: SQ
authors.lastname: Yú
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA.
authors.forename: Steven
authors.identifier: http://orcid.org/0000-0002-9349-338X
authors.initials: S
authors.lastname: Mazur
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA.
authors.forename: Nicole M
authors.identifier: http://orcid.org/0000-0002-8923-0288
authors.initials: NM
authors.lastname: Deiuliis
authors.affiliation: United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, Frederick, Maryland, USA.
authors.forename: Sheli R
authors.identifier: http://orcid.org/0000-0001-8976-8056
authors.initials: SR
authors.lastname: Radoshitzky
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA.
authors.forename: Matthew G
authors.identifier: http://orcid.org/0000-0002-2174-9630
authors.initials: MG
authors.lastname: Lackemeyer
authors.affiliation: Department of Chemistry, Centre for Chemical Biology, School of Environmental and Life Sciences, University of Newcastle, Callaghan, New South Wales, Australia.
authors.forename: Adam
authors.initials: A
authors.lastname: McCluskey
authors.affiliation: Cell Signaling Unit, Children's Medical Research Institute, The University of Sydney, Sydney, New South Wales, Australia.
authors.forename: Phillip J
authors.initials: PJ
authors.lastname: Robinson
authors.affiliation: Leibniz Institut für Molekulare Pharmakologie, Berlin, Germany.
authors.forename: Volker
authors.initials: V
authors.lastname: Haucke
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA.
authors.forename: Victoria
authors.initials: V
authors.lastname: Wahl-Jensen
authors.affiliation: Wisconsin National Primate Research Center, Madison, Wisconsin, USA.
authors.forename: Adam L
authors.initials: AL
authors.lastname: Bailey
authors.affiliation: Wisconsin National Primate Research Center, Madison, Wisconsin, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Lauck
authors.affiliation: Wisconsin National Primate Research Center, Madison, Wisconsin, USA.
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Friedrich
authors.affiliation: Wisconsin National Primate Research Center, Madison, Wisconsin, USA.
authors.forename: David H
authors.identifier: http://orcid.org/0000-0003-2139-470X
authors.initials: DH
authors.lastname: O'Connor
authors.affiliation: Wisconsin National Primate Research Center, Madison, Wisconsin, USA.
authors.forename: Tony L
authors.initials: TL
authors.lastname: Goldberg
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA.
authors.forename: Peter B
authors.identifier: http://orcid.org/0000-0002-9775-3724
authors.initials: PB
authors.lastname: Jahrling
authors.affiliation: Integrated Research Facility at Fort Detrick, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Frederick, Maryland, USA kuhnjens@mail.nih.gov.
authors.forename: Jens H
authors.identifier: http://orcid.org/0000-0002-7800-6045
authors.initials: JH
authors.lastname: Kuhn
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism*; D015214:Antigens, Differentiation, Myelomonocytic / Q000378:metabolism*; D018018:Arterivirus / Q000502:physiology*; D002460:Cell Line; D002522:Chlorocebus aethiops; D004705:Endocytosis; D054884:Host-Pathogen Interactions; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011991:Receptors, Virus / Q000378:metabolism*; D053586:Virus Internalization; D000098902:CD163 Antigen
chemical_list: D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D000098902:CD163 Antigen; D011956:Receptors, Cell Surface; D011991:Receptors, Virus