The zebrafish lens proteome during development and aging.

PURPOSE: Changes in lens protein expression during zebrafish development results in a smooth gradient of refractive index necessary for excellent optical function. Age-related changes in crystallin expression have been well documented in mammals but are poorly understood in the zebrafish. METHODS: In the zebrafish lens, a systematic analysis of protein content with age was performed using size exclusion chromatography (SEC) combined with linear trap quadrupole Fourier transform tandem mass spectrometry (LTQ-FT LC-MS/MS; rank-order shotgun) proteomics in lenses of larval, juvenile, and adult zebrafish. RESULTS: alpha-Crystallins, previously shown to have low abundance in the zebrafish lens, were found to increase dramatically with maturation and aging. SEC determined that beta-crystallin was predominant at 4.5 days. With age, the alpha- and gamma-crystallins increased, and a high molecular weight fraction appeared between six weeks and six months to become the dominant component by 2.5 years. Similarly, shotgun proteomics determined that beta-crystallins were the predominant proteins in the young lens. With age, the proportion of alpha- and gamma-crystallins increased dramatically. After crystallins, calpain 3, membrane, and cytoskeletal proteins were most abundant. Five new beta-crystallins and 13 new gamma-crystallins were identified. CONCLUSIONS: As expected, SEC and proteomics demonstrated changing levels of protein expression with age, especially among the crystallins. The results also confirmed the existence of novel crystallins in the zebrafish genome.