Identification of regions and residues in feline junctional adhesion molecule required for feline calicivirus binding and infection.
The feline junctional adhesion molecule A (fJAM-A) is a functional receptor for feline calicivirus (FCV). fJAM-A is a member of the immunoglobulin superfamily (IgSF) and consists of two Ig-like extracellular domains (D1 and D2), a membrane-spanning domain, and a short cytoplasmic tail. To identify regions of fJAM-A that interact with FCV, we purified recombinant fJAM-A ectodomain and D1 and D2 domains. We found that preincubation of FCV with the ectodomain or D1 was sufficient to inhibit FCV infection in plaque reduction assays. In enzyme-linked immunosorbent assays, FCV binding to fJAM-A ectodomain was concentration dependent and saturable; however, FCV bound D1 alone weakly and was unable to bind D2. To characterize FCV binding to surface-expressed fJAM-A, we transfected truncated and chimeric forms of fJAM-A into a nonpermissive cell line and assayed binding by flow cytometry. Only D1 was necessary for FCV binding to cells; all other domains could be replaced. Using a structure-guided mutational approach, we identified three mutants of fJAM-A within D1 (D42N, K43N, and S97A) that exhibited significantly decreased capacities to bind FCV. In contrast to our finding that D1 mediated FCV binding, we found that all domains of fJAM-A were necessary to confer susceptibility to FCV infection. Furthermore, surface expression of fJAM-A was not sufficient to permit FCV infection by all of the isolates we investigated. This indicates that (i) other cellular factors are required to permit productive FCV infection and (ii) individual FCV isolates differ in the factors they require.
Ossiboff
Robert J
RJ
Baker Institute for Animal Health, College of Veterinary Medicine, Cornell University, Hungerford Hill Road, Ithaca, NY 14853, USA.
Parker
John S L
JS
D000818:Animals; D016466:CHO Cells; D017927:Calicivirus, Feline / Q000378:metabolism* / Q000472:pathogenicity*; D002415:Cats; D015815:Cell Adhesion Molecules / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006367:HeLa Cells; D006801:Humans; D062726:Junctional Adhesion Molecules; D008958:Models, Molecular; D009154:Mutation; D014709:Vero Cells; D010948:Viral Plaque Assay
D015815:Cell Adhesion Molecules; D062726:Junctional Adhesion Molecules