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bioRxiv · 10.1101/2022.06.02.494606

Actin nucleotide state modulates the F-actin structural landscape evoked by bending forces

Reynolds, M. J.·Hachicho, C.·Carl, A. G.·Gong, R.·Alushin, G. M.
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Abstract

ATP hydrolysis-coupled actin polymerization is a fundamental mechanism of cellular force generation. Force and actin filament (F-actin) nucleotide state in turn modulate the engagement of actin binding proteins (ABPs) to regulate actin dynamics through unknown mechanisms. Here, we show that bending forces evoke structural transitions in F-actin which are modulated by actin nucleotide state. Cryo-electron microscopy (cryo-EM) structures of ADP- and ADP-Pi-F-actin with sufficient resolution to visualize bound solvent reveal inter-subunit interactions primarily bridged by waters which could contribute to lattice flexibility. Despite substantial ordered solvent differences in the nucleotide binding cleft, these structures feature essentially indistinguishable protein backbone conformations which are unlikely to be discriminable by ABPs. We next introduce a machine-learning enabled pipeline for reconstructing bent filaments, enabling us to visualize both continuous structural variability and side-chain level detail. Bent F-actin structures reveal major rearrangements at inter-subunit interfaces characterized by striking alterations of helical twist and deformations of individual protomers which are distinct in ADP- and ADP-Pi-F-actin. This suggests phosphate rigidifies actin subunits to alter F-actins bending structural landscape. We therefore propose actin nucleotide state serves as a co-regulator of F-actin mechanical regulation, as bending forces evoke nucleotide-state dependent conformational transitions that could be readily detected by ABPs.

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Reynolds, M. J., Hachicho, C., Carl, A. G., Gong, R., Alushin, G. M.. 2022-06-02. Actin nucleotide state modulates the F-actin structural landscape evoked by bending forces. https://doi.org/10.1101/2022.06.02.494606

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