title: Emetine optimally facilitates nascent chain puromycylation and potentiates the ribopuromycylation method (RPM) applied to inert cells.
abstract: We previously described the ribopuromyclation method (RPM) to visualize and quantitate translating ribosomes in fixed and permeabilized cells by standard immunofluorescence. RPM is based on puromycylation of nascent chains bound to translating ribosomes followed by detection of puromycylated nascent chains with a puromycin-specific mAb. We now demonstrate that emetine optimally enhances nascent chain puromycylation, and describe a modified RPM protocol for identifying ribosome-bound nascent chains in metabolically inert permeabilized cells.
authors.affiliation: Laboratory of Viral Diseases, NIAID, NIH, Bethesda, MD 20892, USA.
authors.forename: Alexandre
authors.initials: A
authors.lastname: David
authors.forename: Jack R
authors.initials: JR
authors.lastname: Bennink
authors.forename: Jonathan W
authors.initials: JW
authors.lastname: Yewdell
mesh_terms: D002463:Cell Membrane Permeability; D004640:Emetine / Q000494:pharmacology*; D006367:HeLa Cells; D006801:Humans; D046188:Protein Modification, Translational / Q000187:drug effects*; D011691:Puromycin / Q000378:metabolism*; D012270:Ribosomes / Q000187:drug effects / Q000378:metabolism
chemical_list: D011691:Puromycin; D004640:Emetine