Research Article

Translation dynamics of single mRNAs in live cells and neurons

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Science  17 Jun 2016:
Vol. 352, Issue 6292, pp. 1430-1435
DOI: 10.1126/science.aaf1084

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The when, where, and how of translation

High-resolution single-molecule imaging shows the spatial and temporal dynamics of molecular events (see the Perspective by Iwasaki and Ingolia). Wu et al. and Morisaki et al. developed an approach to study the translation of single messenger RNAs (mRNAs) in live cells. Nascent polypeptides containing multimerized epitopes were imaged with fluorescent antibody fragments, while simultaneously detecting the single mRNAs using a different fluorescent tag. The approach enabled a direct readout of initiation and elongation, as well as revealing the spatial distribution of translation and allowing the correlation of polysome motility with translation dynamics. Membrane-targeted mRNAs could be distinguished from cytoplasmic mRNAs, as could single polysomes from higher-order polysomal complexes. Furthermore, the work reveals the stochasticity of translation, which can occur constitutively or in bursts, much like transcription, and the spatial regulation of translation in neuronal dendrites.

Science, this issue p. 1430, p. 1425; see also p. 1391

Abstract

Translation is the fundamental biological process converting mRNA information into proteins. Single-molecule imaging in live cells has illuminated the dynamics of RNA transcription; however, it is not yet applicable to translation. Here, we report single-molecule imaging of nascent peptides (SINAPS) to assess translation in live cells. The approach provides direct readout of initiation, elongation, and location of translation. We show that mRNAs coding for endoplasmic reticulum (ER) proteins are translated when they encounter the ER membrane. Single-molecule fluorescence recovery after photobleaching provides direct measurement of elongation speed (5 amino acids per second). In primary neurons, mRNAs are translated in proximal dendrites but repressed in distal dendrites and display “bursting” translation. This technology provides a tool with which to address the spatiotemporal translation mechanism of single mRNAs in living cells.

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