Further insights into GPR179: expression, localization, and associated pathogenic mechanisms leading to complete congenital stationary night blindness.
OBJECTIVE
Mutations in GPR179, which encodes the G protein-coupled receptor 179, lead to autosomal recessive complete (c) congenital stationary night blindness (CSNB), which is characterized by an ON-bipolar retinal cell dysfunction. This study further defined the exact site of Gpr179 expression and its protein localization in human retina and elucidated the pathogenic mechanism of the reported missense and splice site mutations.


METHODS
RNA in situ hybridization was performed with mouse retinal sections. A commercially available antibody was validated with GPR179-overexpressing COS-1 cells and applied to human retinal sections. Live-cell extracellular staining along with subsequent intracellular immunolocalization and ELISA studies were performed using mammalian cells overexpressing wild-type or missense mutated GPR179. Wild-type and splice site-mutated mini-gene constructs were transiently transfected, and RNA was extracted. RT-PCR-amplified products were cloned, and Sanger sequenced.


RESULTS
Mouse Gpr179 transcript was expressed in the upper part of the inner nuclear layer, and the respective human protein localized at the dendritic tips of bipolar cells in human retina. The missense mutations p.Tyr220Cys, p.Gly455Asp, and p.His603Tyr led to severely reduced cell surface localization, whereas p.Asp126His did not. The mutated splice donor site altered GPR179 splicing.


CONCLUSIONS
Our findings indicate that the site of expression and protein localization of human and mouse GPR179 is similar to that of other proteins implicated in cCSNB. For most of the mutations identified so far, loss of the GPR179 protein function seems to be the underlying pathogenic mechanism leading to this form of cCSNB.
Institut National de la Santé et de la Recherche Médicale (INSERM), U968, Paris, France.
Elise
E
Orhan
Laurent
L
Prézeau
Said
S
El Shamieh
Kinga M
KM
Bujakowska
Christelle
C
Michiels
Yvrick
Y
Zagar
Claire
C
Vol
Shomi S
SS
Bhattacharya
José-Alain
JA
Sahel
Florian
F
Sennlaub
Isabelle
I
Audo
Christina
C
Zeitz
D000818:Animals; D004195:Disease Models, Animal; D004596:Electroretinography; D004797:Enzyme-Linked Immunosorbent Assay; D015785:Eye Diseases, Hereditary / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005786:Gene Expression Regulation; D040181:Genetic Diseases, X-Linked / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D017403:In Situ Hybridization; D051379:Mice; D008810:Mice, Inbred C57BL; D009216:Myopia / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism; D009755:Night Blindness / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012313:RNA / Q000235:genetics*; D043562:Receptors, G-Protein-Coupled / Q000096:biosynthesis / Q000235:genetics*; D051245:Retinal Bipolar Cells / Q000378:metabolism / Q000473:pathology*
C575060:GPR179 protein, mouse; D043562:Receptors, G-Protein-Coupled; D012313:RNA
GPR179; cCSNB; expression and localization; mini-gene approach; pathogenicity; trafficking defect