Format

Send to

Choose Destination
Proc Natl Acad Sci U S A. 2015 Sep 29;112(39):12093-8. doi: 10.1073/pnas.1515623112. Epub 2015 Sep 11.

Proteomic mapping in live Drosophila tissues using an engineered ascorbate peroxidase.

Author information

1
Department of Genetics, Harvard Medical School, Boston, MA 02115; clchen@genetics.med.harvard.edu perrimon@receptor.med.harvard.edu.
2
Department of Genetics, Harvard Medical School, Boston, MA 02115;
3
Broad Institute, Cambridge, MA 02142;
4
Broad Institute, Cambridge, MA 02142; Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA 02139;
5
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.

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.

KEYWORDS:

APEX; Drosophila; proteomics

PMID:
26362788
PMCID:
PMC4593093
DOI:
10.1073/pnas.1515623112
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for HighWire Icon for PubMed Central
Loading ...
Support Center