Human Genetics

, Volume 122, Issue 6, pp 615–623 | Cite as

Moderate reduction of Norrin signaling activity associated with the causative missense mutations identified in patients with familial exudative vitreoretinopathy

  • Minghui Qin
  • Hiroyuki Kondo
  • Tomoko Tahira
  • Kenshi Hayashi
Original Investigation


Mutations in Norrin signaling genes (NDP, FZD4 and LRP5) have been found in patients with familial exudative vitreoretinopathy (FEVR) and the altered signaling is suspected to play a critical role in its pathogenesis. To better understand this relationship, we systematically performed functional analyses on previously identified single nucleotide variants of LRP5, FZD4 and NDP, utilizing the Norrin dependent Topflash reporter assay. Cell surface binding assays and protein electrophoresis analysis of Norrin were also performed. Seven causative mutations and five possibly causative but indecisive variants were examined. We found: (1) a nonsense mutation in FZD4 completely abolished its signaling activity, while single missense mutations in LRP5 and FZD4 caused a moderate level of reduction (ranging from 26 to 48, 36% on average) and a double missense mutation in both genes caused a severe reduction in activity (71%). These observations correlated roughly with clinical phenotypes. (2) A mutational effect is suggested in four of five indecisive variants by signaling reductions comparable to those of missense mutations. (3) Norrin mutants demonstrated variable effects on signal transduction, and no apparent correlation with clinical phenotypes was observed. (4) The Norrin mutants examined demonstrated impaired cell surface binding, and some may have partially lost their ability to form a complex with unknown high molecular weight material(s). Our results illustrate the nature of FEVR in relation to Norrin signaling and further suggest the complexity of its disease causing mechanism.



We thank Dr. Randal Moon for kindly providing the reporter plasmid. Special thanks go to Dr. Jeremy Nathans for his generosity of sharing plasmids and cell line. This project was supported by a Grant-in-aid 15591883 for Scientific Research, Japan to H.K. and a Grant-in-Aid for Research Revolution 2002 from The Ministry of Education, Culture, Sports, Science and Technology, Japan to K.H. in the Division of Genome Analysis, Research Center for Genetic Information, Medical Institute of Bioregulation, Kyushu University, Japan.

Supplementary material

439_2007_438_MOESM1_ESM.tif (283 kb)
Additional Norrin mutants. a Cells transfected with FZD4 were probed with indicated Norrin or its mutants. b Alkaline phosphatase stained native polyacrylamide gel was loaded as indicated. The arrowhead indicates the band specific to the Norrin mutant. c Western blot of conditioned medium, containing either wildtype or mutant Norrin as indicated, under either reducing (left) or non-reducing conditions (right) was performed using mAb anti-c-myc (clone 9E). ESM1 (TIF 283 kb)


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Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Minghui Qin
    • 1
  • Hiroyuki Kondo
    • 2
  • Tomoko Tahira
    • 1
  • Kenshi Hayashi
    • 1
  1. 1.Division of Genome Analysis, Research Center for Genetic Information, Medical Institute of BioregulationKyushu UniversityFukuokaJapan
  2. 2.Department of OphthalmologyFukuoka University School of MedicineFukuokaJapan

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