Comparative Proteomic Analysis Identifies Age-Dependent Increases in the Abundance of Specific Proteins after Deletion of the Small Heat Shock Proteins alpha A- and alpha B-Crystallin

Mice with deletion of genes for small heat shock proteins alpha A- and alpha B-crystallin (alpha A/alpha B-/-) develop cataracts. We used proteomic analysis to identify lens proteins that change in abundance after deletion of these alpha-crystallin genes. Wild-type (WT) and alpha A/alpha B-1- knockout (DKO) mice were compared using two-dimensional difference gel electrophoresis and mass spectrometric analysis, and protein identifications were validated by Mascot proteomic software. The abundance of histones H2A, H4, and H2B fragment, and a low molecular weight beta 1-catenin increased 2-3-fold in postnatal day 2 lenses of DKO lenses compared with WT lenses. Additional major increases were observed in abundance of beta B2-crystallin and vimentin in 30-day-old lenses of DKO animals compared with WT animals. Lenses of DKO mice were comprised of nine protein spots containing beta B2-crystallin at 10-40-fold higher abundance and three protein spots containing vimentin at >= 2-fold higher abundance than in WT lenses. Gel permeation chromatography identified a unique 328 kDa protein in DKO lenses, containing beta-crystallin, demonstrating aggregation of beta-crystallin in the absence of alpha-crystallins. Together, these changes provide biochemical evidence for possible functions of specific cell adhesion proteins, cytoskeletal proteins, and crystallins in lens opacities caused by the absence of the. major chaperones, alpha A- and alpha B-crystallins.