title: Proteomic mapping in live Drosophila tissues using an engineered ascorbate peroxidase.
abstract: Characterization of the proteome of organelles and subcellular domains is essential for understanding cellular organization and identifying protein complexes as well as networks of protein interactions. We established a proteomic mapping platform in live Drosophila tissues using an engineered ascorbate peroxidase (APEX). Upon activation, the APEX enzyme catalyzes the biotinylation of neighboring endogenous proteins that can then be isolated and identified by mass spectrometry. We demonstrate that APEX labeling functions effectively in multiple fly tissues for different subcellular compartments and maps the mitochondrial matrix proteome of Drosophila muscle to demonstrate the power of APEX for characterizing subcellular proteomes in live cells. Further, we generate "MitoMax," a database that provides an inventory of Drosophila mitochondrial proteins with subcompartmental annotation. Altogether, APEX labeling in live Drosophila tissues provides an opportunity to characterize the organelle proteome of specific cell types in different physiological conditions.
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, MA 02115; clchen@genetics.med.harvard.edu perrimon@receptor.med.harvard.edu.
authors.forename: Chiao-Lin
authors.initials: CL
authors.lastname: Chen
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, MA 02115;
authors.forename: Yanhui
authors.initials: Y
authors.lastname: Hu
authors.affiliation: Broad Institute, Cambridge, MA 02142;
authors.forename: Namrata D
authors.initials: ND
authors.lastname: Udeshi
authors.affiliation: Broad Institute, Cambridge, MA 02142;
authors.forename: Thomas Y
authors.initials: TY
authors.lastname: Lau
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, MA 02115;
authors.forename: Frederik
authors.initials: F
authors.lastname: Wirtz-Peitz
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, MA 02115;
authors.forename: Li
authors.initials: L
authors.lastname: He
authors.affiliation: Broad Institute, Cambridge, MA 02142; Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA 02139;
authors.forename: Alice Y
authors.initials: AY
authors.lastname: Ting
authors.affiliation: Broad Institute, Cambridge, MA 02142;
authors.forename: Steven A
authors.initials: SA
authors.lastname: Carr
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, MA 02115; Howard Hughes Medical Institute, Boston, MA 02115 clchen@genetics.med.harvard.edu perrimon@receptor.med.harvard.edu.
authors.forename: Norbert
authors.initials: N
authors.lastname: Perrimon
mesh_terms: D000818:Animals; D060387:Ascorbate Peroxidases / Q000235:genetics*; D019295:Computational Biology; D030562:Databases, Protein; D004330:Drosophila / Q000235:genetics / Q000378:metabolism*; D005818:Genetic Engineering / Q000379:methods; D040901:Proteomics / Q000379:methods*; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D060387:Ascorbate Peroxidases
keywords: APEX; Drosophila; proteomics