Description

Ribosome profiling (ribo-seq) is a technique that takes advantage of NGS technology to sequence ribosome-protected mRNA fragments and consequently allows the locations of translating ribosomes to be determined at the entire transcriptome level (Ingolia et al., 2009).

For a more detailed description of the protocol, see Ingolia et al. (2012). For reviews on this technique and its applications, please refer to Ingolia (2014) and Michel et al. (2013).

This track displays cumulative ribo-seq data obtained from human cells under different conditions and can be used for the exploration of human genomic loci that are being translated. The values on the y-axis represent the number of ribosome footprint sequence reads at a given position. As of December 2014, the track contains data from 12 studies (see References section for details). Further details about the aggregated track and additional ribo-seq data from these and other studies including data obtained from other organisms can be found at the specialized ribo-seq browser GWIPS-viz.

Methods

For each study used to generate this track, raw fastq files were downloaded from a repository (e.g., NCBI GEO datasets). Cutadapt was used to trim the relevant adapter sequence from the reads, after which reads below 25 nt in length were discarded. The trimmed reads were aligned to ribosomal RNA using Bowtie and aligning reads were discarded. The remaining reads were then aligned to the hg19 (GRCh37) genome assembly using RUM. An offset of 15 nt (to infer the position of the A-site) was added to the most 5' nucleotide coordinate of each uniquely-mapped read.

The alignment files from each of the included studies were merged to generate this aggregate track.

See individual studies at GWIPS-viz for a full description of the methods of data acquisition and processing.

Credits

Thanks to Audrey Michel and GWIPS-viz for providing the data for this track. If you wish to cite this track, please reference:

Michel AM, Fox G, M Kiran A, De Bo C, O'Connor PB, Heaphy SM, Mullan JP, Donohue CA, Higgins DG, Baranov PV. GWIPS-viz: development of a ribo-seq genome browser. Nucleic Acids Res. 2014 Jan;42(Database issue):D859-64. PMID: 24185699; PMC: PMC3965066

References

Data

Fritsch C, Herrmann A, Nothnagel M, Szafranski K, Huse K, Schumann F, Schreiber S, Platzer M, Krawczak M, Hampe J et al. Genome-wide search for novel human uORFs and N-terminal protein extensions using ribosomal footprinting. Genome Res. 2012 Nov;22(11):2208-18. PMID: 22879431; PMC: PMC3483550

Gonzalez C, Sims JS, Hornstein N, Mela A, Garcia F, Lei L, Gass DA, Amendolara B, Bruce JN, Canoll P et al. Ribosome profiling reveals a cell-type-specific translational landscape in brain tumors. J Neurosci. 2014 Aug 13;34(33):10924-36. PMID: 25122893; PMC: PMC4131009

Guo H, Ingolia NT, Weissman JS, Bartel DP. Mammalian microRNAs predominantly act to decrease target mRNA levels. Nature. 2010 Aug 12;466(7308):835-40. PMID: 20703300; PMC: PMC2990499

Hsieh AC, Liu Y, Edlind MP, Ingolia NT, Janes MR, Sher A, Shi EY, Stumpf CR, Christensen C, Bonham MJ et al. The translational landscape of mTOR signalling steers cancer initiation and metastasis. Nature. 2012 Feb 22;485(7396):55-61. PMID: 22367541; PMC: PMC3663483

Lee S, Liu B, Lee S, Huang SX, Shen B, Qian SB. Global mapping of translation initiation sites in mammalian cells at single-nucleotide resolution. Proc Natl Acad Sci U S A. 2012 Sep 11;109(37):E2424-32. PMID: 22927429; PMC: PMC3443142

Liu B, Han Y, Qian SB. Cotranslational response to proteotoxic stress by elongation pausing of ribosomes. Mol Cell. 2013 Feb 7;49(3):453-63. PMID: 23290916; PMC: PMC3570626

Loayza-Puch F, Drost J, Rooijers K, Lopes R, Elkon R, Agami R. p53 induces transcriptional and translational programs to suppress cell proliferation and growth. Genome Biol. 2013 Apr 17;14(4):R32. PMID: 23594524; PMC: PMC4053767

Reid DW, Nicchitta CV. Primary role for endoplasmic reticulum-bound ribosomes in cellular translation identified by ribosome profiling. J Biol Chem. 2012 Feb 17;287(8):5518-27. PMID: 22199352; PMC: PMC3285328

Stadler M, Fire A. Wobble base-pairing slows in vivo translation elongation in metazoans. RNA. 2011 Dec;17(12):2063-73. PMID: 22045228; PMC: PMC3222120

Stern-Ginossar N, Weisburd B, Michalski A, Le VT, Hein MY, Huang SX, Ma M, Shen B, Qian SB, Hengel H et al. Decoding human cytomegalovirus. Science. 2012 Nov 23;338(6110):1088-93. PMID: 23180859; PMC: PMC3817102

Stumpf CR, Moreno MV, Olshen AB, Taylor BS, Ruggero D. The translational landscape of the mammalian cell cycle. Mol Cell. 2013 Nov 21;52(4):574-82. PMID: 24120665; PMC: PMC3959127

Wein N, Vulin A, Falzarano MS, Szigyarto CA, Maiti B, Findlay A, Heller KN, Uhlén M, Bakthavachalu B, Messina S et al. Translation from a DMD exon 5 IRES results in a functional dystrophin isoform that attenuates dystrophinopathy in humans and mice. Nat Med. 2014 Sep;20(9):992-1000. PMID: 25108525; PMC: PMC4165597

Protocol/Technique

Ingolia NT. Ribosome profiling: new views of translation, from single codons to genome scale. Nat Rev Genet. 2014 Mar;15(3):205-13. PMID: 24468696

Ingolia NT, Brar GA, Rouskin S, McGeachy AM, Weissman JS. The ribosome profiling strategy for monitoring translation in vivo by deep sequencing of ribosome- protected mRNA fragments. Nat Protoc. 2012 Jul 26;7(8):1534-50. PMID: 22836135; PMC: PMC3535016

Ingolia NT, Ghaemmaghami S, Newman JR, Weissman JS. Genome-wide analysis in vivo of translation with nucleotide resolution using ribosome profiling. Science. 2009 Apr 10;324(5924):218-23. PMID: 19213877; PMC: PMC2746483

Michel AM, Baranov PV. Ribosome profiling: a Hi-Def monitor for protein synthesis at the genome-wide scale. Wiley Interdiscip Rev RNA. 2013 Sep-Oct;4(5):473-90. PMID: 23696005; PMC: PMC3823065