title: EYS Is a Protein Associated with the Ciliary Axoneme in Rods and Cones.
abstract: OBJECTIVE
  Mutations in the EYS gene are a common cause of autosomal recessive retinitis pigmentosa (arRP), yet the role of the EYS protein in humans is presently unclear. The aim of this study was to investigate the isoform structure, expression and potential function of EYS in the mammalian retina in order to better understand its involvement in the pathogenesis of arRP.
  
  
  METHODS
  To achieve the objective, we examined the expression of mRNA transcripts of EYS isoforms in human tissues and cell lines by RT-PCR. We also investigated the localisation of EYS in cultured cells and retinal cryo-sections by confocal fluorescence microscopy and Western blot analysis.
  
  
  RESULTS
  RT-PCR analysis confirmed that EYS has at least four isoforms. In addition to the previously reported EYS isoforms 1 and 4, we present the experimental validation of two smaller variants referred to as EYS isoforms 2 and 3. All four isoforms are expressed in the human retina and Y79 cells and the short variants were additionally detected in the testis. Immunofluorescent confocal microscopy and Western blot analysis revealed that all EYS isoforms preferentially localise to the cytoplasm of Y79 and HeLa cells. Moreover, an enrichment of the endogenous protein was observed near the centrosomes in Y79 cells. Interestingly, EYS was observed at the ciliary axoneme in Y79 ciliated cells. In macaque retinal cryosections, EYS was found to localise in the region of the photoreceptor ciliary axoneme in both rods and cones as well as in the cytoplasm of the ganglion cells.
  
  
  CONCLUSIONS
  The results obtained in this study lead us to speculate that, in photoreceptor cells, EYS could be a protein involved in maintaining the stability of the ciliary axoneme in both rods and cones. The variability of its isoform structure suggests that other roles are also possible and yet to be established.
authors.affiliation: Institute of Ophthalmology, UCL, London, EC1V 9EL, United Kingdom.
authors.forename: Giovanna
authors.initials: G
authors.lastname: Alfano
authors.affiliation: Institute of Ophthalmology, UCL, London, EC1V 9EL, United Kingdom.
authors.forename: Przemyslaw M
authors.initials: PM
authors.lastname: Kruczek
authors.affiliation: Institute of Ophthalmology, UCL, London, EC1V 9EL, United Kingdom.
authors.forename: Amna Z
authors.initials: AZ
authors.lastname: Shah
authors.affiliation: Institute of Ophthalmology, UCL, London, EC1V 9EL, United Kingdom.
authors.forename: Barbara
authors.initials: B
authors.lastname: Kramarz
authors.affiliation: Institute of Ophthalmology, UCL, London, EC1V 9EL, United Kingdom.
authors.forename: Glen
authors.initials: G
authors.lastname: Jeffery
authors.affiliation: Department of Biology, Indiana University, 1001 East Third St., Bloomington, IN, 47405, United States of America.
authors.forename: Andrew C
authors.initials: AC
authors.lastname: Zelhof
authors.affiliation: Institute of Ophthalmology, UCL, London, EC1V 9EL, United Kingdom.
authors.forename: Shomi S
authors.initials: SS
authors.lastname: Bhattacharya
mesh_terms: D000818:Animals; D054468:Axoneme / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D002923:Cilia / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism; D004252:DNA Mutational Analysis; D004596:Electroretinography; D005136:Eye Proteins / Q000235:genetics* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007211:Indoles; D008251:Macaca / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012160:Retina / Q000378:metabolism / Q000473:pathology; D017949:Retinal Cone Photoreceptor Cells / Q000378:metabolism* / Q000473:pathology; D017948:Retinal Rod Photoreceptor Cells / Q000378:metabolism* / Q000473:pathology; D012174:Retinitis Pigmentosa / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
chemical_list: C533105:EYS protein, human; D005136:Eye Proteins; D007211:Indoles; D020033:Protein Isoforms; C007293:DAPI