title: A novel nonsense mutation in the MIP gene linked to congenital posterior polar cataracts in a Chinese family.
abstract: OBJECTIVE
  To detect the causative mutation for congenital posterior polar cataracts in a five-generation Chinese family and further explore the potential pathogenesis of this disease.
  
  
  METHODS
  Coding exons, with flanking sequences of five candidate genes, were screened using direct DNA sequencing. The identified mutations were confirmed by restriction fragment length polymorphism (RFLP) analysis. A full-length wild-type or an Y219* mutant aquaporin0 (AQP0) fused with an N-terminal FLAG tag, was transfected into HEK293T cells. For co-localization studies, FLAG-WT-AQP0 and Myc-Y219*-AQP0 constructs were co-transfected. Quantitative real-time RT-PCR, western blotting and immunofluorescence studies were performed to determine protein expression levels and sub-cellular localization, respectively.
  
  
  RESULTS
  We identified a novel nonsense mutation in MIP (c.657 C>G; p.Y219*) (major intrinsic protein gene) that segregates with congenital posterior polar cataract in a Chinese family. This mutation altered a highly conserved tyrosine to a stop codon (Y219*) within AQP0.When FLAG-WT-AQP0 and FLAG-Y219*-AQP0 expression constructs were singly transfected into HEK 293T cells, mRNA expression showed no significant difference between the wild-type and the mutant, while Y219*-AQP0 protein expression was significantly lower than that of wild-type AQP0. Wild-type AQP0 predominantly localized to the plasma membrane, while the mutated protein was abundant within the cytoplasm of HEK293T cells. However, when FLAG-WT-AQP0 andMyc-MU-AQP0were co-expressed, both proteins showed high fluorescence in the cytoplasm.
  
  
  CONCLUSIONS
  The novel nonsense mutation in the MIP gene (c.657 C>G) identified in a Chinese family may cause posterior polar cataracts. The dominant negative effect of the mutated protein on the wild-type protein interfered with the trafficking of wild-type protein to the cell membrane and both the mutant and wild-type protein were trapped in the cytoplasm. Consequently, both wild-type and mutant protein lost their function as a water channel on the cell membrane, and may result in a cataract phenotype. Our data also expands the spectrum of known MIP mutations.
authors: Department of Ophthalmology, Shengjing Hospital of China Medical University, Shenyang, Liaoning, 110004, China.
authors: Zixun
authors: Z
authors: Song
authors: Department of Medical Genetics, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, P. R. China.
authors: Lianqing
authors: L
authors: Wang
authors: Department of Medical Genetics, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, P. R. China.
authors: Yaping
authors: Y
authors: Liu
authors: Department of Ophthalmology, Shengjing Hospital of China Medical University, Shenyang, Liaoning, 110004, China.
authors: Wei
authors: W
authors: Xiao
mesh_terms: D000328:Adult; D000595:Amino Acid Sequence; D020346:Aquaporins / Q000235:genetics*; D044466:Asian People / Q000235:genetics; D001483:Base Sequence; D016022:Case-Control Studies; D002386:Cataract / Q000151:congenital* / Q000235:genetics*; D002478:Cells, Cultured; D018389:Codon, Nonsense; D005136:Eye Proteins / Q000235:genetics*; D005260:Female; D006801:Humans; D007908:Lens, Crystalline / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D010375:Pedigree; D020641:Polymorphism, Single Nucleotide
chemical_list: D020346:Aquaporins; D018389:Codon, Nonsense; D005136:Eye Proteins; C030328:aquaporin 0