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Mason Posner, Phillip A. Wages, Joseph Horwitz, Linlin Ding; Developmental Changes In Zebrafish (Danio rerio) Lens Crystallin Expression. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2773.
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© ARVO (1962-2015); The Authors (2016-present)
The zebrafish is attracting increased attention as a model species for investigations of lens function and development. In this study we detail the ontogenetic changes in zebrafish lens crystallin content to facilitate the use of this model species for the study of crystallin function and lens aging.
Lens homogenates from 2, 7 and 13.5 month old zebrafish were fractionated by FPLC on a Superose HR6 column (50 mM NaPO4, 0.1 M NaCl, pH 7.0) and the resulting peaks analyzed by SDS-PAGE to identify crystallin families. Lens homogenates from 10 day, 6 week and 15 month old zebrafish were separated by 2-D electrophoresis using 11 cm pH 3-10 nonlinear IPG strips and 12.5% SDS-PAGE gels. The resulting 2-D gels were stained with SYPRO Ruby and the pixel density of each resulting spot was measured to calculate the relative abundance of each lens crystallin.
Analysis of the first major chromatographic peak indicated the presence of both α- and β-crystallins. This combined α-/β-crystallin peak eluted earlier in the 7 and 13.5 month adult samples, likely due to an increased proportion of the larger α-crystallins. Using physiological buffers we failed to detect high molecular weight aggregated crystallins in adult lenses. The crystallin profiles from our 2-DE gel analysis showed significant changes in the total amount and relative abundance of individual crystallins during development. α-crystallin contributed less than 1% of the total protein content in 10 day old larval lenses, rising to 6.5% of the adult 15 month old lens. αA-crystallin was always the most abundant while αBb-crystallin was least abundant. β-crystallins were abundant at all stages and maintained similar relative abundance with the exception of βB3, which did not occur at noticeable levels until adulthood. γ-crystallins showed diverse ontogenetic profiles, with γN1, γN2 and γS4 found throughout development, while γS1, γS2 and γS3 abundance increased significantly in adulthood.
Our results show dynamic changes in lens crystallin abundance during zebrafish development. The increased expression of specific β- and γ-crystallins in adult lenses suggests that individual crystallins may have age-specific functions. We confirmed the co-occurrence of zebrafish α- and β-crystallins in the typical "α-crystallin peak" that makes using size exclusion chromatography as the sole approach a poor method for estimating teleost fish lens α-crystallin abundance.
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