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Using EMOTE to Map the Exact 5′-Ends of Processed RNA on a Transcriptome-Wide Scale

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RNA Remodeling Proteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1259))

Abstract

The presence or absence of structure in an RNA is often crucial to its function. This is evident for highly structured RNAs such as rRNA, tRNA, or riboswitches, but it is also the case for many mRNAs, where secondary structures in the 5′ or 3′ UTR can determine the efficiency of translation or the half-life of the RNA. There are paths to modify such secondary structures, (1) by the action of a helicase that allows an alternative RNA structure to form, (2) by the formation of a duplex with another RNA, or (3) by cleavage of the RNA in a way that favors a different secondary structure. None of the three exclude the others, and in vivo it is common that two or all three work together to remodel an RNA to the desired form. However, while the first two solutions can be reversible, the cleavage of RNA is final, and there is no chance to go back. In this chapter, a method for tracking the 5′ end created by RNA processing on a transcriptome-wide scale is presented. The Exact Mapping Of Transcriptome Ends (EMOTE) allows the large-scale identification of mono-phosphorylated RNA 5′-ends and provides the exact processing sites.

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References

  1. Sambrook J, Russell DW (2001) Molecular cloning. Cold Spring Harbor Laboratory Press, New York, NY

    Google Scholar 

  2. Sharma CM, Hoffmann S, Darfeuille F et al (2010) The primary transcriptome of the major human pathogen Helicobacter pylori. Nature 464:250–255

    Article  CAS  PubMed  Google Scholar 

  3. Dugar G, Herbig A, Förstner KU et al (2013) High-resolution transcriptome maps reveal strain-specific regulatory features of multiple Campylobacter jejuni isolates. PLoS Genet 9:e1003495

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  4. Jorjani H, Zavolan M (2014) TSSer: an automated method to identify transcription start sites in prokaryotic genomes from differential RNA sequencing data. Bioinformatics 30:971–974

    Article  CAS  PubMed  Google Scholar 

  5. Deana A, Celesnik H, Belasco JG (2008) The bacterial enzyme RppH triggers messenger RNA degradation by 5′ pyrophosphate removal. Nature 451:355–358

    Article  CAS  PubMed  Google Scholar 

  6. Hsieh PK, Richards J, Liu Q et al (2013) Specificity of RppH-dependent RNA degradation in Bacillus subtilis. Proc Natl Acad Sci U S A 110:8864–8869

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  7. Piton J, Larue V, Thillier Y et al (2013) Bacillus subtilis RNA deprotection enzyme RppH recognizes guanosine in the second position of its substrates. Proc Natl Acad Sci U S A 110:8858–8863

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  8. Linder P, Lemeille S, Redder P (2014) Transcriptome-wide analyses of 5′-ends in RNase J mutants of a gram-positive pathogen reveal a role in RNA maturation, regulation and degradation. PLoS Genet 10:e1004207

    Article  PubMed Central  PubMed  Google Scholar 

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Acknowledgements

The critical reading of this manuscript by Vanessa Guimarães and Patrick Linder was highly appreciated. Work in the laboratory is funded by SwissLife Jubiläumsstiftung, Novartis Consumer Health Foundation, the Faculty of Medicine at University of Geneva, the Swiss National Science Foundation, and the Canton of Geneva.

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Correspondence to Peter Redder .

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Redder, P. (2015). Using EMOTE to Map the Exact 5′-Ends of Processed RNA on a Transcriptome-Wide Scale. In: Boudvillain, M. (eds) RNA Remodeling Proteins. Methods in Molecular Biology, vol 1259. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2214-7_5

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  • DOI: https://doi.org/10.1007/978-1-4939-2214-7_5

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-2213-0

  • Online ISBN: 978-1-4939-2214-7

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