Developmental Proteomics of Mouse Retina.

Abstract

Abstract: : Purpose: Our goal is to understand ocular physiology and pathology through proteomics. Toward this goal we plan to identify and quantify the retinal proteins displayed on two–dimensional (2–D) gels along the developmental axis. Methods: Retinas were obtained from C57/B6 mice at postnatal (P) days 1, 3, 5, 7, 9, 14, 21 and at the adult stages (P>35). The proteins were extracted and separated by 2–D gel electrophoresis. Protein spots were visualized with Coomassie brilliant blue–R250 and the 2–D gels were dried using cellophane sheets. The gels were scanned by a 16 bit transmission scanner (PowerLookIII, UMAX) and the images were analyzed by Progenesis Workstation (Nonlinear Dynamics). Some of the proteins that exhibited over 3–folds changes in their expression levels during the postnatal development were excised from the gel, digested in gel with trypsin, and analyzed by matrix–assisted laser desorption/ionization time–of–flight mass spectrometer (MALDI–TOF–MS; Voyager Elite, Applied Biosystems). The peptide mass fingerprint search was performed through MS–Fit and/or MASCOT. Identities of some of the proteins were confirmed by MS/MS using MALDI–quadrupole ion trap TOF MS (Axima QIT, Shimadzu/Kratos). Results: Approximately 1,000 protein spots were detected on each gel. Nearly half of them showed significant changes in their expression levels with various kinetic profiles during the postnatal development. Among the proteins showing over 3–folds changes, the levels of glutamine synthetase and neuron–specific enolase increased, and those of proliferation–associated protein 1 and heterogeneous nuclear ribonucleoprotein K decreased. In contrast, the level of phosphoglycerate kinase 1 remained constant during the entire developmental stages. Conclusions: Our developmental proteomics of mouse retina demonstrates that approximately 500 retinal proteins detected on a gel underwent significant changes in their expression levels during maturation. Their expression profiles clearly correlate with the crucial molecular events underlying the development of retinal neurons. Our proteome maps of mouse retina at various postnatal developmental stages would be a crucial tool to understand the visual pathophysiology.

Keywords: proteomics • retina • retinal development