SILAC-based quantitative proteomic analysis of Drosophila gastrula stage embryos mutant for fibroblast growth factor signaling

Abstract

AbstractThe application of quantitative proteomics in model organisms has been successful in determining changes in the proteome under distinct physiological conditions. Quantitative mass spectrometry-based proteomic analyses in combination with genetics provide powerful tools in developmental cell signaling research. Drosophila melanogaster is one of the most widely used genetic models for studying development and disease. Here we combined quantitative proteomics with genetic selection to determine global changes in the proteome upon depletion of the Heartless (Htl) Fibroblast-Growth Factor (FGF) receptor signaling in Drosophila embryos at early gastrulation stages. We present a robust, single generation SILAC (stable isotope labeling with amino acids in cell culture) protocol for labeling proteins in early embryos and for selection of homozygously mutant embryos at pre-gastrula stages using an independent genetic marker. Our analyses detected quantitative changes in the global proteome of htl mutant embryos during gastrulation. We identified distinct classes of down-regulated and up-regulated proteins and network analyses indicates functionally related groups of proteins in each class. These data suggest that FGF signaling in the early embryo affects global changes in the abundance of metabolic, nucleoplasmic, cytoskeletal and endomembrane transport proteins.