Dataset Information

Grc3 programs the essential endoribonuclease Las1 for specific RNA cleavage.

ABSTRACT: Las1 is a recently discovered endoribonuclease that collaborates with Grc3-Rat1-Rai1 to process precursor ribosomal RNA (rRNA), yet its mechanism of action remains unknown. Disruption of the mammalian Las1 gene has been linked to congenital lethal motor neuron disease and X-linked intellectual disability disorders, thus highlighting the necessity to understand Las1 regulation and function. Here, we report that the essential Las1 endoribonuclease requires its binding partner, the polynucleotide kinase Grc3, for specific C2 cleavage. Our results establish that Grc3 drives Las1 endoribonuclease cleavage to its targeted C2 site both in vitro and in Saccharomyces cerevisiae. Moreover, we observed Las1-dependent activation of the Grc3 kinase activity exclusively toward single-stranded RNA. Together, Las1 and Grc3 assemble into a tetrameric complex that is required for competent rRNA processing. The tetrameric Grc3/Las1 cross talk draws unexpected parallels to endoribonucleases RNaseL and Ire1, and establishes Grc3/Las1 as a unique member of the RNaseL/Ire1 RNA splicing family. Together, our work provides mechanistic insight for the regulation of the Las1 endoribonuclease and identifies the tetrameric Grc3/Las1 complex as a unique example of a protein-guided programmable endoribonuclease.

SUBMITTER: Pillon MC

PROVIDER: S-EPMC5514736 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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Grc3 programs the essential endoribonuclease Las1 for specific RNA cleavage.

Pillon Monica C MC Sobhany Mack M Borgnia Mario J MJ Williams Jason G JG Stanley Robin E RE

Proceedings of the National Academy of Sciences of the United States of America 20170626 28

Las1 is a recently discovered endoribonuclease that collaborates with Grc3-Rat1-Rai1 to process precursor ribosomal RNA (rRNA), yet its mechanism of action remains unknown. Disruption of the mammalian Las1 gene has been linked to congenital lethal motor neuron disease and X-linked intellectual disability disorders, thus highlighting the necessity to understand Las1 regulation and function. Here, we report that the essential Las1 endoribonuclease requires its binding partner, the polynucleotide k ...[more]

PMID: 28652339

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Project description:Posttranscriptional regulation plays an essential role in the quick adaptation of pathogenic bacteria to host environments, and RNases play key roles in this process by modifying small RNAs and mRNAs. We find that the Pseudomonas aeruginosa endonuclease YbeY is required for rRNA processing and the bacterial virulence in a murine acute pneumonia model. Transcriptomic analyses reveal that knocking out the ybeY gene results in downregulation of oxidative stress response genes, including the catalase genes katA and katB Consistently, the ybeY mutant is more susceptible to H2O2 and neutrophil-mediated killing. Overexpression of katA restores the bacterial tolerance to H2O2 and neutrophil killing as well as virulence. We further find that the downregulation of the oxidative stress response genes is due to defective expression of the stationary-phase sigma factor RpoS. We demonstrate an autoregulatory mechanism of RpoS and find that ybeY mutation increases the level of a small RNA, ReaL, which directly represses the translation of rpoS through the 5' UTR of its mRNA and subsequently reduces the expression of the oxidative stress response genes. In vitro assays demonstrate direct degradation of ReaL by YbeY. Deletion of reaL or overexpression of rpoS in the ybeY mutant restores the bacterial tolerance to oxidative stress and the virulence. We also demonstrate that YbeZ binds to YbeY and is involved in the 16S rRNA processing and regulation of reaL and rpoS as well as the bacterial virulence. Overall, our results reveal pleiotropic roles of YbeY and the YbeY-mediated regulation of rpoS through ReaL.IMPORTANCE The increasing bacterial antibiotic resistance imposes a severe threat to human health. For the development of effective treatment and prevention strategies, it is critical to understand the mechanisms employed by bacteria to grow in the human body. Posttranscriptional regulation plays an important role in bacterial adaptation to environmental changes. RNases and small RNAs are key players in this regulation. In this study, we demonstrate critical roles of the RNase YbeY in the virulence of the pathogenic bacterium Pseudomonas aeruginosa We further identify the small RNA ReaL as the direct target of YbeY and elucidate the YbeY-regulated pathway on the expression of bacterial virulence factors. Our results shed light on the complex regulatory network of P. aeruginosa and indicate that inference with the YbeY-mediated regulatory pathway might be a valid strategy for the development of a novel treatment strategy.

Dataset Information

Grc3 programs the essential endoribonuclease Las1 for specific RNA cleavage.

Publications

Grc3 programs the essential endoribonuclease Las1 for specific RNA cleavage.

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