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Nat Biotechnol. 2014 May;32(5):465-72. doi: 10.1038/nbt.2860. Epub 2014 Apr 13.

Proteome labeling and protein identification in specific tissues and at specific developmental stages in an animal.

Author information

1
1] Medical Research Council Laboratory of Molecular Biology, Cambridge, England, UK. [2].
2
Medical Research Council Laboratory of Molecular Biology, Cambridge, England, UK.
3
1] Medical Research Council Laboratory of Molecular Biology, Cambridge, England, UK. [2] Department of Chemistry, University of Cambridge, Cambridge, England, UK.
4
Department of Biochemistry, University of Cambridge, Cambridge, England, UK.

Abstract

Identifying the proteins synthesized at specific times in cells of interest in an animal will facilitate the study of cellular functions and dynamic processes. Here we introduce stochastic orthogonal recoding of translation with chemoselective modification (SORT-M) to address this challenge. SORT-M involves modifying cells to express an orthogonal aminoacyl-tRNA synthetase/tRNA pair to enable the incorporation of chemically modifiable analogs of amino acids at diverse sense codons in cells in rich media. We apply SORT-M to Drosophila melanogaster fed standard food to label and image proteins in specific tissues at precise developmental stages with diverse chemistries, including cyclopropene-tetrazine inverse electron demand Diels-Alder cycloaddition reactions. We also use SORT-M to identify proteins synthesized in germ cells of the fly ovary without dissection. SORT-M will facilitate the definition of proteins synthesized in specific sets of cells to study development, and learning and memory in flies, and may be extended to other animals.

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PMID:
24727715
PMCID:
PMC4107302
DOI:
10.1038/nbt.2860
[Indexed for MEDLINE]
Free PMC Article

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