Single-molecule imaging reveals the role of membrane-binding motif and C-terminal domain of RNase E in its localization and diffusion in Escherichia coli

Troyer, L.; Wang, Y.-H.; Shobhna, S.; Kim, S.; Woo, J.; Tajkhorshid, E.

doi:10.1101/2024.11.05.622141

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bioRxiv · 10.1101/2024.11.05.622141

Single-molecule imaging reveals the role of membrane-binding motif and C-terminal domain of RNase E in its localization and diffusion in Escherichia coli

Troyer, L.·Wang, Y.-H.·Shobhna, S.·Kim, S.·Woo, J.·Tajkhorshid, E.

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Abstract

In Escherichia coli, RNase E, a central enzyme in RNA processing and mRNA degradation, contains a catalytic N-terminal domain (NTD), a membrane-targeting sequence (MTS), and a C-terminal domain (CTD). We investigated how MTS and CTD influence RNase E localization, diffusion, and function. Super-resolution microscopy revealed that [~]93% of RNase E localizes to the inner membrane and exhibits slow diffusion similar to polysomes. Comparing the native amphipathic MTS with a transmembrane motif showed that the MTS confers slower diffusion and stronger membrane binding. The CTD further slows diffusion by increasing mass but unexpectedly weakens membrane association. RNase E mutants with partial cytoplasmic localization displayed enhanced co-transcriptional degradation of lacZ mRNA. These findings indicate that variations in the MTS and the presence of the CTD shape the spatiotemporal organization of RNA processing in bacterial cells, providing mechanistic insight into how RNase E domain architecture influences its cellular function.

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Troyer, L., Wang, Y.-H., Shobhna, S., Kim, S., Woo, J., Tajkhorshid, E.. 2024-11-05. Single-molecule imaging reveals the role of membrane-binding motif and C-terminal domain of RNase E in its localization and diffusion in Escherichia coli. https://doi.org/10.1101/2024.11.05.622141

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