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Sequencing technologies to measure translation in single cells

Nature Reviews Molecular Cell Biology volume 26, pages 337–346 (2025)Cite this article

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Abstract

Translation is one of the most energy-intensive processes in a cell and, accordingly, is tightly regulated. Genome-wide methods to measure translation and the translatome and to study the complex regulation of protein synthesis have enabled unprecedented characterization of this crucial step of gene expression. However, technological limitations have hampered our understanding of translation control in multicellular tissues, rare cell types and dynamic cellular processes. Recent optimizations, adaptations and new techniques have enabled these measurements to be made at single-cell resolution. In this Progress, we discuss single-cell sequencing technologies to measure translation, including ribosome profiling, ribosome affinity purification and spatial translatome methods.

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Fig. 1: Comparison of bulk and single-cell ribosome profiling methods.
Fig. 2: Single-cell technologies to measure the degree of ribosome association.

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Acknowledgements

This work was supported by a European Research Council Advanced grant (ERC-AdG 101053581-scTranslatomics).

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Authors and Affiliations

  1. Oncode Institute, Utrecht, the Netherlands

    Michael VanInsberghe & Alexander van Oudenaarden

  2. Hubrecht Institute-KNAW (Royal Netherlands Academy of Arts and Sciences), Utrecht, the Netherlands

    Michael VanInsberghe & Alexander van Oudenaarden

  3. University Medical Center Utrecht, Utrecht, the Netherlands

    Michael VanInsberghe & Alexander van Oudenaarden

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  1. Michael VanInsberghe
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  2. Alexander van Oudenaarden
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M.V. contributed to all aspects of the article. A.v.O contributed substantially to discussion of the content and reviewed and/or edited the manuscript before submission.

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Correspondence to Michael VanInsberghe.

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M.V. and A.v.O are inventors on a patent application related to measuring translation in single cells.

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Nature Reviews Molecular Cell Biology thanks Shu-Bing Qian, Marko Jovanovic, who co-reviewed with Ella Doron-Mandel, and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Glossary

Adapters

Sequences attached to a diverse set of RNA molecules, which are used to label, amplify and sequence those molecules.

Barcodes

Unique sequences that are added to all RNA fragments originating from the same source, thereby enabling pooling together of material from many samples for efficient sequencing, after which the barcodes are used to identify the original source of each read.

Isotachophoresis

(ITP). An electrophoretic technique for the selective separation and concentration of charged molecules.

Monosomes

Single ribosomes attached to an mRNA or mRNA fragment.

Polysome

Several ribosomes attached to an mRNA or mRNA fragment.

Random forest

A machine learning method that combines the output of multiple decision trees for classification and regression predictions.

Ribosome density

The number of ribosomes per mRNA.

Translation efficiency

The rate of polypeptide synthesis per mRNA per time.

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VanInsberghe, M., van Oudenaarden, A. Sequencing technologies to measure translation in single cells. Nat Rev Mol Cell Biol 26, 337–346 (2025). https://doi.org/10.1038/s41580-024-00822-z

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