YihE is a novel binding partner of Rho and regulates Rho-dependent transcription termination in the Cpx stress response

Wang, Biying; Pei, Hairun; Lu, Zhonghua; Xu, Yingying; Han, Shengnan; Jia, Zongchao; Zheng, Jimin

doi:10.1016/j.isci.2022.105483

Cited by 5 publications

(9 citation statements)

References 64 publications

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“…YihE is induced upon periplasmatic stress (43, 51), whereas stress conditions under which Hfq and Rof regulate ρ activity remain to be identified. YihE binds the ρ NTD, inhibiting ρ-RNA interactions (20); the mechanism of ρ regulation by Rof has been determined in this study, and molecular details of Hfq-mediated ρ inhibition remain to be solved (16, 19). By directly binding ρ, each of these anti-terminators may interfere with the formation of ρ-dependent termination complexes that assemble in an EC-dependent manner (center left) (31, 32) or RNA-dependent manner (center right) (47, 48).…”

Section: Resultsmentioning

confidence: 99%

“…Under stress conditions when dedicated anti-termination machineries and translating ribosomes are scarce, ρ action may need to be tuned down. To date, three cellular proteins that inhibit ρ function, Hfq, YihE and Rof, have been described (19)(20)(21)50). While a recent study revealed molecular details for YihE (20), the mechanisms of ρ inhibition by the Sm-like proteins Rof and Hfq remain incompletely understood.…”

Section: Discussionmentioning

confidence: 99%

“…Hfq can directly bind ρ, and inhibits ρ by an unknown, and possibly indirect, mechanism (16, 19). YihE forms a complex with ρ and inhibits ρ-RNA interactions (20). Rof proteins adopt an Sm-like fold, resembling Hfq (Fig.…”

Section: Introductionmentioning

confidence: 99%

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Sm-like protein Rof inhibits transcription termination factor ρ by binding site obstruction and conformational insulation

Said,

Finazzo,

Hilal

et al. 2023

Preprint

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Transcription termination factor Rho is a hexameric, RNA-dependent NTPase that can adopt active closed-ring and inactive open-ring conformations. The Sm-like protein Rof, a homolog of the RNA chaperone Hfq, inhibits Rho-dependent termination in vivo but recapitulation of this activity in vitro has proven difficult and the precise mode of Rof action is presently unknown. Our electron microscopic structures of Rho-Rof and Rho-RNA complexes show that Rof undergoes pronounced conformational changes to bind Rho at the protomer interfaces, undercutting Rho conformational dynamics associated with ring closure and occluding extended primary RNA-binding sites that are also part of interfaces between Rho and RNA polymerase. Consistently, Rof impedes Rho ring closure, Rho-RNA interactions, and Rho association with transcription elongation complexes. Structure-guided mutagenesis coupled with functional assays confirmed that the observed Rho-Rof interface is required for Rof-mediated inhibition of cell growth and Rho-termination in vitro. Bioinformatic analyses revealed that Rof is restricted to Pseudomonadota and that the Rho-Rof interface is conserved. Genomic contexts of rof differ between Enterobacteriaceae and Vibrionaceae, suggesting distinct modes of Rof regulation. We hypothesize that Rof and other cellular anti-terminators silence Rho under diverse, but yet to be identified, stress conditions when unrestrained transcription termination by Rho would be lethal.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Sm-like protein Rof inhibits transcription termination factor ρ by binding site obstruction and conformational insulation

Said,

Finazzo,

Hilal

et al. 2023

Preprint

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“…A recent study used structural, genetic, and biochemical methods to determine that Rof binding to Rho inhibits Rho association with both RNA and RNAP (Said et al , 2023). YihE is the third protein factor that has been reported to modulate Rho-dependent termination, and it also binds Rho and inhibits Rho-RNA interactions (Wang et al , 2022). What remains a mystery is when and why these regulators act on Rho.…”

Section: Discussionmentioning

confidence: 99%

Multiple Small RNAs Modulate Rho-Dependent Termination in the Cyclopropane Fatty Acid Synthase mRNA 5’ Untranslated Region

Farley,

Bianco,

Vanderpool

2024

Preprint

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Bacterial small RNAs (sRNAs) have been commonly characterized as regulators of post-transcriptional steps of gene expression including translation and stability of mRNA targets. Previous work revealed that the Escherichia coli cyclopropane fatty acid synthase (cfa) mRNA is regulated by at least five different sRNAs by a proposed mechanism involving regulated cfa mRNA turnover by the RNase E degradosome. However, recent work identified the long 5' untranslated region (UTR) of cfa mRNA as a potential target for Rho-dependent transcription termination, leading us to question whether sRNAs might regulate cfa gene expression at the level of transcription elongation. In this study we report evidence for premature Rho-dependent termination within the long 5' UTR of cfa, and demonstrate that a pyrimidine-only tract within the 5' UTR is required for efficient Rho-dependent regulation of cfa. Our data suggest that all of the sequence determinants required for efficient Rho-mediated termination are harbored within the cfa long mRNA 5' UTR. Finally, we discovered that both the activating sRNA RydC and repressing sRNA CpxQ regulate cfa primarily by modulating Rho-dependent termination of cfa transcription, with only a minor effect on RNase E degradosome-dependent turnover of cfa mRNA. These results illustrate the versatile mechanisms sRNAs use to regulate target gene expression at transcriptional and post-transcriptional levels and suggest that regulation by sRNAs in long UTRs can involve modulation of transcription elongation.

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“…However, during slow growth or stress, when translation is inefficient, indiscriminate termination by ρ may be lethal; consistently, partial loss-offunction mutations in the rho gene enable E. coli survival on ethanol, which inhibits translation 13,14 . ρ cellular levels are subject to autoregulation 15 , suggesting that ρ must be inhibited during stress, e.g., by accessory proteins that inhibit ρ-RNA interactions 16,17 .…”

Section: Mainmentioning

confidence: 99%

Transcription termination factor ρ polymerizes under stress

Wang,

Said,

Hilal

et al. 2023

Preprint

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Bacterial RNA helicase ρ is a genome sentinel that terminates synthesis of damaged and junk RNAs that are not translated by the ribosome. Co-transcriptional RNA surveillance by ρ is essential for quality control of the transcriptome during optimal growth. However, it is unclear how bacteria protect their RNAs from overzealous ρ during dormancy or stress, conditions common in natural habitats. Here we used cryogenic electron microscopy, biochemical, and genetic approaches to show that residue substitutions, ADP, or ppGpp promote hyper-oligomerization of Escherichia coli ρ. Our results demonstrate that nucleotides bound at subunit interfaces control ρ switching from active hexamers to inactive higher-order oligomers and extended filaments. Polymers formed upon exposure to antibiotics or ppGpp disassemble when stress is relieved, thereby directly linking termination activity to cellular physiology. Inactivation of ρ through hyper-oligomerization is a regulatory strategy shared by RNA polymerases, ribosomes, and metabolic enzymes across all life.

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YihE is a novel binding partner of Rho and regulates Rho-dependent transcription termination in the Cpx stress response

Cited by 5 publications

References 64 publications

Sm-like protein Rof inhibits transcription termination factor ρ by binding site obstruction and conformational insulation

Sm-like protein Rof inhibits transcription termination factor ρ by binding site obstruction and conformational insulation

Multiple Small RNAs Modulate Rho-Dependent Termination in the Cyclopropane Fatty Acid Synthase mRNA 5’ Untranslated Region

Transcription termination factor ρ polymerizes under stress

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