title: Comparative proteomic analysis identifies age-dependent increases in the abundance of specific proteins after deletion of the small heat shock proteins αA- and αB-crystallin.
abstract: Mice with deletion of genes for small heat shock proteins αA- and αB-crystallin (αA/αB(-/-)) develop cataracts. We used proteomic analysis to identify lens proteins that change in abundance after deletion of these α-crystallin genes. Wild-type (WT) and αA/αB(-/-) 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 β1-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 βB2-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 βB2-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 β-crystallin, demonstrating aggregation of β-crystallin in the absence of α-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, αA- and αB-crystallins.
authors.affiliation: Department of Ophthalmology and Visual Sciences, Washington University, School of Medicine, St. Louis, MO 63110, USA. andley@vision.wustl.edu
authors.forename: Usha P
authors.initials: UP
authors.lastname: Andley
authors.forename: James P
authors.initials: JP
authors.lastname: Malone
authors.forename: Paul D
authors.initials: PD
authors.lastname: Hamilton
authors.forename: Nathan
authors.initials: N
authors.lastname: Ravi
authors.forename: R Reid
authors.initials: RR
authors.lastname: Townsend
mesh_terms: D000367:Age Factors; D000818:Animals; D015153:Blotting, Western; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003459:Crystallins / Q000235:genetics* / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D006360:Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D013058:Mass Spectrometry; D051379:Mice; D018345:Mice, Knockout; D040901:Proteomics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014746:Vimentin / Q000378:metabolism; D051176:beta Catenin / Q000235:genetics* / Q000378:metabolism
chemical_list: D003459:Crystallins; D006360:Heat-Shock Proteins; D006657:Histones; D014746:Vimentin; D051176:beta Catenin