title: Thy1 associates with the cation channel subunit HCN4 in adult rat retina.
abstract: OBJECTIVE
  The membrane expression and gene promoter of the glycosylphosphatidylinositol (GPI)-anchored protein Thy1 have been widely used to examine the morphology and distribution of retinal ganglion cells in normal eyes and disease models. However, it is not known how adult mammalian retinal neurons use Thy1. Because Thy1 is not a membrane-spanning protein and, instead, complexes with structural and signaling proteins in other tissues, the aim of this study was to find protein partners of retinal Thy1.
  
  
  METHODS
  Coimmunoprecipitation, immunohistochemistry, confocal imaging, and patch-clamp recording were used to test for association of Thy1 and HCN4, a cation channel subunit, in adult rat retina.
  
  
  RESULTS
  Hyperpolarization of cells immunopanned by an anti-Thy1 antibody activated HCN channels. Confocal imaging showed that individual somata in the ganglion cell layer bound antibodies against Thy1 and HCN4, that the majority of these bindings colocalized, and that some of the immunopositive cells also bound antibody against a ganglion cell marker (Brn3a). Consistent with these results, Thy1 and HCN4 were coimmunoprecipitated by magnetic beads coated with either anti-Thy1 antibody or anti-HCN4 antibody. In control experiments, beads coated with these antibodies did not immunoprecipitate a photoreceptor rim protein (ABCR) and uncoated beads did not immunoprecipitate either Thy1 or HCN4.
  
  
  CONCLUSIONS
  This is the first report that Thy1 colocalizes and coimmunoprecipitates with a membrane-spanning protein in retina, that Thy1 complexes with an ion channel protein in any tissue, and that a GPI-anchored protein associates with an HCN channel subunit protein.
authors.affiliation: Section of Neurobiology, Physiology, and Behavior, University ofCalifornia, Davis, California 95616-8519, USA.
authors.forename: Gloria J
authors.initials: GJ
authors.lastname: Partida
authors.forename: Tyler W
authors.initials: TW
authors.lastname: Stradleigh
authors.forename: Genki
authors.initials: G
authors.lastname: Ogata
authors.forename: Iv
authors.initials: I
authors.lastname: Godzdanker
authors.forename: Andrew T
authors.initials: AT
authors.lastname: Ishida
mesh_terms: D000818:Animals; D005260:Female; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D047468:Immunoprecipitation; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000378:metabolism*; D051381:Rats; D020318:Rats, Long-Evans; D012165:Retinal Ganglion Cells / Q000166:cytology / Q000378:metabolism*; D018800:Thy-1 Antigens / Q000378:metabolism*
chemical_list: C513836:HCN4 protein, rat; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D015221:Potassium Channels; D018800:Thy-1 Antigens