Bursting translation on single mRNAs in live cells

Nathan M Livingston; Jiwoong Kwon; Oliver Valera; James A Saba; Niladri K Sinha; Pranav Reddy; Blake Nelson; Clara Wolfe; Taekjip Ha; Rachel Green; Jian Liu; Bin Wu

doi:10.1016/j.molcel.2023.05.019

Bursting translation on single mRNAs in live cells

Mol Cell. 2023 Jul 6;83(13):2276-2289.e11. doi: 10.1016/j.molcel.2023.05.019. Epub 2023 Jun 16.

Authors

Nathan M Livingston¹, Jiwoong Kwon¹, Oliver Valera², James A Saba³, Niladri K Sinha³, Pranav Reddy¹, Blake Nelson¹, Clara Wolfe⁴, Taekjip Ha⁵, Rachel Green⁶, Jian Liu⁷, Bin Wu⁸

Affiliations

¹ Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Center for Cell Dynamics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
² Department of Cell Biology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Center for Cell Dynamics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
³ Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
⁴ Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
⁵ Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Center for Cell Dynamics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA.
⁶ Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA.
⁷ Department of Cell Biology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Center for Cell Dynamics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. Electronic address: jliu187@jhmi.edu.
⁸ Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Center for Cell Dynamics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Solomon H Snyder Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. Electronic address: bwu20@jhmi.edu.

Abstract

Stochasticity has emerged as a mechanism of gene regulation. Much of this so-called "noise" has been attributed to bursting transcription. Although bursting transcription has been studied extensively, the role of stochasticity in translation has not been fully investigated due to the lack of enabling imaging technology. In this study, we developed techniques to track single mRNAs and their translation in live cells for hours, allowing the measurement of previously uncharacterized translation dynamics. We applied genetic and pharmacological perturbations to control translation kinetics and found that, like transcription, translation is not a constitutive process but instead cycles between inactive and active states, or "bursts." However, unlike transcription, which is largely frequency-modulated, complex structures in the 5'-untranslated region alter burst amplitudes. Bursting frequency can be controlled through cap-proximal sequences and trans-acting factors such as eIF4F. We coupled single-molecule imaging with stochastic modeling to quantitatively determine the kinetic parameters of translational bursting.

Keywords: RNA; bursting; single molecule; stochastic gene expression; translation.

Bursting translation on single mRNAs in live cells

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