title: Human coronavirus NL63 utilizes heparan sulfate proteoglycans for attachment to target cells.
abstract: Human coronavirus NL63 (HCoV-NL63) is an alphacoronavirus that was first identified in 2004 in the nasopharyngeal aspirate from a 7-month-old patient with a respiratory tract infection. Previous studies showed that HCoV-NL63 and the genetically distant severe acute respiratory syndrome (SARS)-CoV employ the same receptor for host cell entry, angiotensin-converting enzyme 2 (ACE2), but it is largely unclear whether ACE2 interactions are sufficient to allow HCoV-NL63 binding to cells. The present study showed that directed expression of angiotensin-converting enzyme 2 (ACE2) on cells previously resistant to HCoV-NL63 renders them susceptible, showing that ACE2 protein acts as a functional receptor and that its expression is required for infection. However, comparative analysis showed that directed expression or selective scission of the ACE2 protein had no measurable effect on virus adhesion. In contrast, binding of HCoV-NL63 to heparan sulfates was required for viral attachment and infection of target cells, showing that these molecules serve as attachment receptors for HCoV-NL63.
  
  
  OBJECTIVE
  ACE2 protein was proposed as a receptor for HCoV-NL63 already in 2005, but an in-depth analysis of early events during virus infection had not been performed thus far. Here, we show that the ACE2 protein is required for viral entry but that it is not the primary binding site on the cell surface. Conducted research showed that heparan sulfate proteoglycans function as adhesion molecules, increasing the virus density on cell surface and possibly facilitating the interaction between HCoV-NL63 and its receptor. Obtained results show that the initial events during HCoV-NL63 infection are more complex than anticipated and that a newly described interaction may be essential for understanding the infection process and, possibly, also assist in drug design.
authors.affiliation: Microbiology Department, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Milewska
authors.affiliation: Division of Cell Biophysics, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland.
authors.forename: Miroslaw
authors.initials: M
authors.lastname: Zarebski
authors.affiliation: Microbiology Department, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland.
authors.forename: Paulina
authors.initials: P
authors.lastname: Nowak
authors.affiliation: Microbiology Department, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland.
authors.forename: Karol
authors.initials: K
authors.lastname: Stozek
authors.affiliation: Microbiology Department, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland Department of Oral Immunology and Infectious Diseases, School of Dentistry, University of Louisville, Louisville, Kentucky, USA.
authors.forename: Jan
authors.initials: J
authors.lastname: Potempa
authors.affiliation: Microbiology Department, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland Laboratory of Virology, Malopolska Centre of Biotechnology, Jagiellonian University, Krakow, Poland k.a.pyrc@uj.edu.pl.
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Pyrc
mesh_terms: D000085962:Angiotensin-Converting Enzyme 2; D000818:Animals; D002460:Cell Line; D058957:Coronavirus NL63, Human / Q000502:physiology*; D019812:Heparan Sulfate Proteoglycans / Q000378:metabolism*; D006801:Humans; D007703:Peptidyl-Dipeptidase A / Q000235:genetics / Q000378:metabolism*; D053585:Virus Attachment
chemical_list: D019812:Heparan Sulfate Proteoglycans; D007703:Peptidyl-Dipeptidase A; C000705307:ACE2 protein, human; D000085962:Angiotensin-Converting Enzyme 2