Architectural principles for Hfq/Crc-mediated regulation of gene expression.

Xue Yuan Pei,Ben F Luisi,Shaoxia Chen,Tom Dendooven,Udo Bläsi,Elisabeth Sonnleitner

doi:10.7554/elife.43158

Xue Yuan Pei, Ben F Luisi + Show 4 more

Open Access

https://doi.org/10.7554/elife.43158

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Abstract

In diverse bacterial species, the global regulator Hfq contributes to post-transcriptional networks that control expression of numerous genes. Hfq of the opportunistic pathogen Pseudomonas aeruginosa inhibits translation of target transcripts by forming a regulatory complex with the catabolite repression protein Crc. This repressive complex acts as part of an intricate mechanism of preferred nutrient utilisation. We describe high-resolution cryo-EM structures of the assembly of Hfq and Crc bound to the translation initiation site of a target mRNA. The core of the assembly is formed through interactions of two cognate RNAs, two Hfq hexamers and a Crc pair. Additional Crc protomers are recruited to the core to generate higher-order assemblies with demonstrated regulatory activity in vivo. This study reveals how Hfq cooperates with a partner protein to regulate translation, and provides a structural basis for an RNA code that guides global regulators to interact cooperatively and regulate different RNA targets.

Highlights

The RNA chaperone Hfq contributes to the control of mRNA translation through different modes of action
To gain insight into how P. aeruginosa Hfq cooperates with Crc in translational repression of amiE, we determined the structure of the complex they form on the Hfq binding motif of the carbon catabolite repression (CCR)-controlled amiE mRNA using cryo-electron microscopy
For cryo-EM structural studies of the Hfq/Crc/RNA complex, purified recombinant Hfq and Crc proteins were mixed with an 18 nucleotide Hfq binding motif from the translation initiation region of the CCR-controlled amiE mRNA

Summary

Introduction

The RNA chaperone Hfq contributes to the control of mRNA translation through different modes of action. Gram-negative pathogen Pseudomonas aeruginosa, Hfq acts as a pleiotropic regulator of metabolism (Sonnleitner and Blasi, 2014), virulence (Sonnleitner et al, 2003; Fernandez et al, 2016; Pusic et al, 2016), quorum sensing (Sonnleitner et al, 2006; Yang et al, 2015) and stress responses (Lu et al, 2016) Many of these roles are likely facilitated through partner molecules, and numerous putative protein interactors of P. aeruginosa Hfq have been identified with functions in transcription, translation and mRNA decay (Van den Bossche et al, 2014). To gain insight into how P. aeruginosa Hfq cooperates with Crc in translational repression of amiE, we determined the structure of the complex they form on the Hfq binding motif of the CCR-controlled amiE mRNA using cryo-electron microscopy (cryoEM). These findings expand the paradigm for in vivo action of Hfq through cooperation with the Crc helper protein and RNA to form effector assemblies

Results

Discussion

Materials and methods

Funding Funder Wellcome Trust