title: Imaging protein synthesis in cells and tissues with an alkyne analog of puromycin.
abstract: Synthesis of many proteins is tightly controlled at the level of translation, and plays an essential role in fundamental processes such as cell growth and proliferation, signaling, differentiation, or death. Methods that allow imaging and identification of nascent proteins are critical for dissecting regulation of translation, both spatially and temporally, particularly in whole organisms. We introduce a simple and robust chemical method to image and affinity-purify nascent proteins in cells and in animals, based on an alkyne analog of puromycin, O-propargyl-puromycin (OP-puro). OP-puro forms covalent conjugates with nascent polypeptide chains, which are rapidly turned over by the proteasome and can be visualized or captured by copper(I)-catalyzed azide-alkyne cycloaddition. Unlike methionine analogs, OP-puro does not require methionine-free conditions and, uniquely, can be used to label and assay nascent proteins in whole organisms. This strategy should have broad applicability for imaging protein synthesis and for identifying proteins synthesized under various physiological and pathological conditions in vivo.
authors.affiliation: Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA.
authors.forename: Jing
authors.initials: J
authors.lastname: Liu
authors.forename: Yangqing
authors.initials: Y
authors.lastname: Xu
authors.forename: Dan
authors.initials: D
authors.lastname: Stoleru
authors.forename: Adrian
authors.initials: A
authors.lastname: Salic
mesh_terms: D000480:Alkynes / Q000737:chemistry; D001386:Azides / Q000737:chemistry; D003300:Copper; D003952:Diagnostic Imaging / Q000379:methods*; D009682:Magnetic Resonance Spectroscopy; D008856:Microscopy, Fluorescence; D015394:Molecular Structure; D014176:Protein Biosynthesis / Q000502:physiology*; D011691:Puromycin / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry
chemical_list: D000480:Alkynes; D001386:Azides; C570448:O-propargyl-puromycin; D011691:Puromycin; D003300:Copper