Global analysis of cellular protein translation by pulsed SILAC.
Current methods for system-wide gene expression analysis detect changes in mRNA abundance, but neglect regulation at the level of translation. Pulse labeling with stable isotopes has been used to measure protein turnover rates, but this does not directly provide information about translation rates. Here, we developed pulsed stable isotope labeling by amino acids in cell culture (pSILAC) with two heavy isotope labels to directly quantify protein translation on a proteome-wide scale. We applied the method to cellular iron homeostasis as a model system and demonstrate that it can confidently identify proteins that are translationally regulated by iron availability.
Schwanhäusser
Björn
B
Max Delbrück Center for Molecular Medicine, Berlin, Germany.
Gossen
Manfred
M
Dittmar
Gunnar
G
Selbach
Matthias
M
D000596:Amino Acids / Q000032:analysis / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D005656:Fungal Proteins / Q000032:analysis / Q000378:metabolism; D005786:Gene Expression Regulation; D015966:Gene Expression Regulation, Fungal; D006367:HeLa Cells; D006801:Humans; D007501:Iron / Q000378:metabolism; D007553:Isotope Labeling / Q000379:methods*; D008156:Luciferases / Q000032:analysis / Q000378:metabolism; D014176:Protein Biosynthesis; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods; D015003:Yeasts
D000596:Amino Acids; D005656:Fungal Proteins; D020543:Proteome; D007501:Iron; D008156:Luciferases