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K. Varadaraj, S. S. Kumari, T. Okamura, N. Kasai, A. Shiels, R. Patil, M. B. Wax, R. T. Mathias; Congenital Cataract and Microopthalmia Result from Cytotoxicity and Necrosis in AQP0 Mutant Mouse, CatTohm. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5859.
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© ARVO (1962-2015); The Authors (2016-present)
A deletion mutation in the 2nd transmembrane domain of AQP0 resulted in a dominant cataract, and smaller lens and eye in CatTohm mice. The present study was conducted to determine the cause of the cataract and microopthalmia.
Transgenic mice that expresses mutant AQP0 (Tg-ΔAQP0), CatTohm mice and AQP0-/- mice were used to generate mouse models expressing ΔAQP0 protein in the presence or absence of wild type (WT) AQP0 in the lens fiber cells. WT AQP0 and ΔAQP0 were expressed in Xenopus oocytes and in MDCK cells. Trafficking and dominant negative effects were studied using FRET. Water permeability (Pw) was determined from the rate of volume change. Cytotoxicity due to ΔAQP0 protein was determined using necrosis- and apoptosis-specific staining and cytotoxicity assays.
Co-localization studies demonstrated that ΔAQP0 protein trapped WT AQP0 in the ER. Microscopic observations and functional studies showed that lens fiber cell morphology and membrane Pw were severely affected; the severity of cataract and reduction in Pw depended on the quantity of mutant protein present in the fiber cells. WT AQP0 protein localized in the plasma membranes of oocytes and cultured cells whereas ΔAQP0 was mostly in the cytoplasmic organelles. FRET studies showed that WT AQP0 localized with the co-expressed ΔAQP0. Necrosis and apoptosis were significantly higher in the ΔAQP0 transfected cells than in the WT AQP0 transfected cells. However, necrosis was higher than apoptosis in the ΔAQP0 transfected cells. The cultured lenses of CatTohm showed higher levels of LDH in the medium indicating necrosis.
The deletion mutation significantly affects the trafficking of AQP0 to the membrane resulting in reduced fiber cell Pw. Interactions of mutant and wild type AQP0 proteins cause the dominant negative effect. Cytotoxicity due to mutant protein accumulation leads to fiber cell disintegration and manifests as cataract, smaller lens and eye.
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