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The mechanical stability of proteins regulates their translocation rate into the cell nucleus

Abstract

A cell’s ability to react to mechanical stimuli is known to be affected by the transport of transcription factors, the proteins responsible for regulating transcription of DNA into RNA, across the membrane enveloping its nucleus. Yet the molecular mechanisms by which mechanical cues control this process remain unclear. Here we show that one such protein, myocardin-related transcription factor A (MRTFA), is imported into the nucleus at a rate that is inversely correlated with its nanomechanical stability, but independent of its thermodynamic stability. Attaching mechanically stable proteins to MRTFA results in reduced gene expression and the subsequent slowing down of cell migration. We conclude that the mechanical unfolding of proteins regulates their nuclear translocation rate, and highlight the role of the nuclear pore complex as a selective mechanosensor that is capable of detecting forces as low as 10 pN. The modulation of the mechanical stability of transcription factors may represent a general strategy for the control of gene expression.

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Fig. 1: Protein unfolding modulates the kinetics of nuclear import.
Fig. 2: The kinetics of MRTFA nuclear import is regulated by its mechanical properties.
Fig. 3: The NPC is highly mechanoselective.
Fig. 4: The mechanical selectivity of the NPC probed with optogenetic constructs.
Fig. 5: Mechanically stable MRTFA constructs downregulate gene expression and cellular motility.

Data availability

Data supporting this research can be obtained from the corresponding author on reasonable request.

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Acknowledgements

We thank M. Vartiainen (University of Helsinki) for sharing the MRTFA–GFP plasmid, M. Parsons (King’s College London) for sharing the pEBFP2–N1 and pEYFP–N1 vectors and the MDA-MB-231 cell line, and U. Eggert and M. Pfuhl (King’s College London) for providing HeLa and Top10 competent cells, respectively. We thank the Nikon Imaging Centre at King’s College London for assistance in setting up the cell-imaging experiments. We wish to thank G. Yang for help in qPCR analysis, and C. Nichols and S. Conte (King’s College London) for help with differential scanning fluorimetry experiments. A.E.M.B. is the recipient of a Sir Henry Wellcome fellowship (210887/Z/18/Z). V.S.R. was funded by the BHF Centre for Research Excellence at King’s College London. This work was supported by BHF grant PG/13/50/30426, the European Commission (Mechanocontrol, grant agreement SEP-210342844), EPSRC Fellowship K00641X/1, EPSRC Strategic Equipment Grant EP/M022536/1, Leverhulme Trust Project Grant RPG-2015-225, Leverhulme Trust Research Leadership Award RL-2016-015, Wellcome Trust Investigator Award 212218/Z/18/Z and Royal Society Wolfson Fellowship RSWF/R3/183006, all to S.G.-M.

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S.G.-M. conceived the research. E.I. designed and performed cell biology, live-cell imaging experiments, motility assays and qPCR experiments. A.S. designed and performed live-cell imaging analysis and kinetic modelling. P.R.-L., S.J.B., E.R. and A.L. performed molecular biology work. A.E.M.B. and Y.J.W. conducted single-molecule nanomechanical experiments and A.E.M.B. analysed data. E.R. with A.S. conducted and analysed differential scanning fluorimetry experiments. S.G.B., F.P. and V.S.R. acquired preliminary data. C.S. and P.R.-C. participated in data discussion. S.G.-M., E.I. and A.S. wrote the paper.

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Correspondence to Sergi Garcia-Manyes.

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Infante, E., Stannard, A., Board, S.J. et al. The mechanical stability of proteins regulates their translocation rate into the cell nucleus. Nat. Phys. 15, 973–981 (2019). https://doi.org/10.1038/s41567-019-0551-3

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