Skip to main content

Ribosome Profiling in Trypanosomatids

  • Protocol
  • First Online:
Leishmania

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1971))

Abstract

Ribosomes are the machinery responsible for reading mRNAs and translating them into proteins. The ribosome profiling approach is based on high-throughput sequencing of ribosome-protected mRNAs. RNAs not harboring ribosomes are removed by nuclease digestion leaving the so-called ribosome “footprints.” The purified “footprint” RNA molecules are processed into DNA libraries and their individual abundance is determined by deep sequencing. Ribosome profiling reveals the portion of transcripts which are actually protein-coding and can be used for differential gene expression analysis addressing rates of protein synthesis, and translational control and efficiency.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 149.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Hardcover Book
USD 199.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Clayton CE (2002) Life without transcriptional control? From fly to man and back again. EMBO J 21(8):1881–1888

    Article  CAS  Google Scholar 

  2. Ingolia NT et al (2009) Genome-wide analysis in vivo of translation with nucleotide resolution using ribosome profiling. Science 324(5924):218–223

    Article  CAS  Google Scholar 

  3. Vasquez JJ et al (2014) Comparative ribosome profiling reveals extensive translational complexity in different Trypanosoma brucei life cycle stages. Nucleic Acids Res 42(6):3623–3637

    Article  CAS  Google Scholar 

  4. Smircich P et al (2015) Ribosome profiling reveals translation control as a key mechanism generating differential gene expression in Trypanosoma cruzi. BMC Genomics 16:443

    Article  Google Scholar 

  5. Ingolia NT et al (2012) The ribosome profiling strategy for monitoring translation in vivo by deep sequencing of ribosome-protected mRNA fragments. Nat Protoc 7(8):1534–1550

    Article  CAS  Google Scholar 

  6. Jensen BC et al (2005) Species specificity in ribosome biogenesis: a nonconserved phosphoprotein is required for formation of the large ribosomal subunit in Trypanosoma brucei. Eukaryot Cell 4(1):30–35

    Article  CAS  Google Scholar 

  7. Ingolia NT (2010) Genome-wide translational profiling by ribosome footprinting. Methods Enzymol 470:119–142

    Article  CAS  Google Scholar 

  8. Webb H et al (2005) Developmentally regulated instability of the GPI-PLC mRNA is dependent on a short-lived protein factor. Nucleic Acids Res 33(5):1503–1512

    Article  CAS  Google Scholar 

Download references

Acknowledgments

We thank T. Nicolai Siegel and Juan José Vasquez for establishing the ribosome profiling protocol in Trypanosoma brucei. This work was funded by a European Research Council Starting Grant (3D_Tryps 715466). R.O.C was supported by a Georg Forster Fellowship (Humboldt Foundation).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Raúl O. Cosentino .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2019 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Kraus, A.J., Cosentino, R.O. (2019). Ribosome Profiling in Trypanosomatids. In: Clos, J. (eds) Leishmania. Methods in Molecular Biology, vol 1971. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-9210-2_5

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-9210-2_5

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-9209-6

  • Online ISBN: 978-1-4939-9210-2

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics