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In horses, the CXCL16 gene is located on equine chromosome 11 (ECA11) and encodes a glycosylated, type I transmembrane protein with 247 amino acids. Stable transfection of HEK-293T cells with plasmid DNA carrying EqCXCL16 (HEK-EqCXCL16 cells) increased the proportion of the cell population permissive to EAV infection from &lt;3% to almost 100%. The increase in permissiveness was blocked either by transfection of HEK-EqCXCL16 cells with small interfering RNAs (siRNAs) directed against EqCXCL16 or by pretreatment with guinea pig polyclonal antibody against EqCXCL16 protein (Gp anti-EqCXCL16 pAb). Furthermore, using a virus overlay protein-binding assay (VOPBA) in combination with far-Western blotting, gradient-purified EAV particles were shown to bind directly to the EqCXCL16 protein<jats:italic>in vitro<\/jats:italic>. The binding of biotinylated virulent EAV strain Bucyrus at 4\u00b0C was significantly higher in HEK-EqCXCL16 cells than nontransfected HEK-293T cells. Finally, the results demonstrated that EAV preferentially infects subpopulations of horse CD14<jats:sup>+<\/jats:sup>monocytes expressing EqCXCL16 and that infection of these cells is significantly reduced by pretreatment with Gp anti-EqCXCL16 pAb. The collective data from this study provide confirmatory evidence that the transmembrane form of EqCXCL16 likely plays a major role in EAV host cell entry processes, possibly acting as a primary receptor molecule for this virus.<\/jats:p><jats:p><jats:bold>IMPORTANCE<\/jats:bold>Outbreaks of EVA can be a source of significant economic loss for the equine industry from high rates of abortion in pregnant mares, death in young foals, establishment of the carrier state in stallions, and trade restrictions imposed by various countries. Similar to other arteriviruses, EAV primarily targets cells of the monocyte\/macrophage lineage, which, when infected, are believed to play a critical role in EVA pathogenesis. To this point, however, the host-specified molecules involved in EAV binding and entry into monocytes\/macrophages have not been identified. Identification of the cellular receptors for EAV may provide insights to design antivirals and better prophylactic reagents. In this study, we have demonstrated that EqCXCL16 acts as an EAV entry receptor in EAV-susceptible cells, equine monocytes. These findings represent a significant advance in our understanding of the fundamental mechanisms associated with the entry of EAV into susceptible cells.<\/jats:p>","DOI":"10.1128\/jvi.02455-15","type":"journal-article","created":{"date-parts":[[2016,1,14]],"date-time":"2016-01-14T09:42:57Z","timestamp":1452764577000},"page":"3366-3384","update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":30,"title":["Equine Arteritis Virus Uses Equine CXCL16 as an Entry Receptor"],"prefix":"10.1128","volume":"90","author":[{"given":"Sanjay","family":"Sarkar","sequence":"first","affiliation":[{"name":"Department of Veterinary Science, Maxwell H. Gluck Equine Research Center, University of Kentucky, Lexington, Kentucky, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lakshman","family":"Chelvarajan","sequence":"additional","affiliation":[{"name":"Department of Veterinary Science, Maxwell H. 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