title: Identification of novel protein functions and signaling mechanisms by genetics and quantitative phosphoproteomics in Caenorhabditis elegans.
abstract: Stable isotope labeling by amino acids combined with mass spectrometry is a widely used methodology for measuring relative changes in protein and phosphorylation levels at a global level. We have applied this method to the model organism Caenorhabditis elegans in combination with RNAi-mediated gene knockdown by feeding the nematode on pre-labeled lysine auxotroph Escherichia coli. In this chapter, we describe in details the generation of the E. coli strain, incorporation of heavy isotope-labeled lysine in C. elegans, and the procedure for a comprehensive global phosphoproteomic experiment.
authors: Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, 5230, Odense M, Denmark.
authors: Julius
authors: J
authors: Fredens
authors: Kasper
authors: K
authors: Engholm-Keller
authors: Jakob
authors: J
authors: Møller-Jensen
authors: Martin Røssel
authors: MR
authors: Larsen
authors: Nils J
authors: NJ
authors: Færgeman
mesh_terms: D000818:Animals; D017173:Caenorhabditis elegans / Q000166:cytology* / Q000235:genetics / Q000378:metabolism*; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D055785:Gene Knockdown Techniques; D007553:Isotope Labeling; D013058:Mass Spectrometry; D010748:Phosphopeptides / Q000737:chemistry / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D059748:Proteolysis; D040901:Proteomics / Q000379:methods*; D034622:RNA Interference; D015398:Signal Transduction; D014025:Titanium / Q000737:chemistry
chemical_list: D029742:Caenorhabditis elegans Proteins; D010748:Phosphopeptides; D010750:Phosphoproteins; C009495:titanium dioxide; D014025:Titanium