Ribosome Profiling of Mouse Embryonic Stem Cells Reveals the Complexity and Dynamics of Mammalian Proteomes

• Published in: Cell 2011-11, Vol.147 (4), p.789-802
• Main Authors: Ingolia, Nicholas T., Lareau, Liana F., Weissman, Jonathan S.
• Format: Article
• Language: English
• Subjects: Algorithms; Animals; Artificial Intelligence; Codons; Differentiation; Embryo cells; Embryoid Bodies - cytology; Embryoid Bodies - metabolism; embryonic stem cells; Embryonic Stem Cells - metabolism; Gene mapping; genome; Genomes; Genomics - methods; Harringtonines - pharmacology; high-throughput nucleotide sequencing; High-Throughput Nucleotide Sequencing - methods; Kinetics; Learning algorithms; messenger RNA; Mice; Open Reading Frames; Peptide Chain Initiation, Translational; Protein Biosynthesis; protein products; protein synthesis; Ribosomes; Ribosomes - chemistry; Ribosomes - drug effects; RNA - analysis; Sequence Analysis, RNA - methods; Stem cells; translation (genetics); Translation elongation; Translation initiation
• ISSN: 0092-8674; 1097-4172
• DOI: 10.1016/j.cell.2011.10.002

The ability to sequence genomes has far outstripped approaches for deciphering the information they encode. Here we present a suite of techniques, based on ribosome profiling (the deep sequencing of ribosome-protected mRNA fragments), to provide genome-wide maps of protein synthesis as well as a pulse-chase strategy for determining rates of translation elongation. We exploit the propensity of harringtonine to cause ribosomes to accumulate at sites of translation initiation together with a machine learning algorithm to define protein products systematically. Analysis of translation in mouse embryonic stem cells reveals thousands of strong pause sites and unannotated translation products. These include amino-terminal extensions and truncations and upstream open reading frames with regulatory potential, initiated at both AUG and non-AUG codons, whose translation changes after differentiation. We also define a class of short, polycistronic ribosome-associated coding RNAs (sprcRNAs) that encode small proteins. Our studies reveal an unanticipated complexity to mammalian proteomes. [Display omitted] ► Ribosome-profiling technique reveals complexity of mammalian proteome ► Many transcripts previously characterized as noncoding are in fact translated ► Translation proceeds at 5.6 codons per second and stalls at Pro-Pro-Glu motifs ► mESC differentiation involves global shifts in upstream translation A high-resolution look at mammalian translation reveals unanticipated diversity in the resulting proteome, including peptide products from putative noncoding RNAs.