Increased Association of Deamidated αA- with Lens membrane of transgenic αA vs. wild type αA mice: potential effects on intracellular ionic imbalance and membrane disorganization.

Unlabelled: We have generated two mouse models, in one by inserting the human lens αAN101D transgene in CRYαA mice, and in the other by inserting human wild-type αA-transgene in CRYαA mice. The CRYαA mice developed cortical cataract at about 7-months of age relative to CRYαA mice. The objective of the study was to determine the following relative changes in the lenses of CRYαA- vs. CRYαA mice: age-related changes with specific emphasis on protein insolubilization, relative membrane-association of αA vs. WTαA proteins, and changes in intracellular ionic imbalance and membrane organization.

Methods: Lenses of varying ages from CRYαA and CRYαA mice were compared for an age-related protein insolubilization. The relative lens membrane-association of the αAN101D- and WTαA proteins in the two types of mice was determined by immunohistochemical-, immunogold-labeling-, and western blot analyses. The relative levels of membrane-binding of recombinant αA- and WTαA proteins was determined by an in vitro assay, and the levels of intracellular Ca uptake and Na, K-ATPase mRNA were determined in the cultured epithelial cells from lenses of the two types of mice.

Results: Compared to the lenses of CRYαA, the lenses of CRYαA mice exhibited: (A) An increase in age-related protein insolubilization beginning at about 4-months of age. (B) A greater lens membrane-association of αAN101D- relative to WTαA protein during immunogold-labeling- and western blot analyses, including relatively a greater membrane swelling in the CRYαA lenses. (C) During in vitro assay, the greater levels of binding αAN101D- relative to WTαA protein to membranes was observed. (D) The 75% lower level of Na, K-ATPase mRNA but 1.5X greater Ca uptake were observed in cultured lens epithelial cells of CRYαA than those of CRYαA mice.

Conclusions: The results show that an increased lens membrane association of αA-relative WTαA protein in CRYαA mice than CRYαA mice occurs, which causes intracellular ionic imbalance, and in turn, membrane swelling that potentially leads to cortical opacity.