RNase G controls tpiA mRNA abundance in response to oxygen availability in Escherichia coli

Lee, Jaejin; Lee, Dong‐Ho; Jeon, Che Ok; Lee, Kangseok

doi:10.1007/s12275-019-9354-6

Cited by 10 publications

(8 citation statements)

References 40 publications

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“…Following confirmation of the quality and quantity of extracted total RNA samples with a Nanodrop 2000 (Thermo Scientific, Wilmington, DE, USA), cDNA was synthesized using an iScript cDNA Synthesis Kit (Bio‐Rad Laboratories, Hercules, CA, USA). Samples for RT–qPCR were prepared and analyzed as previously described (Lee et al , 2019). The same primers described above for the allele‐specific PCR analysis were used for RT–qPCR analysis of FOXL2 and GAPDH expression.…”

Section: Methodsmentioning

confidence: 99%

An alternative miRISC targets a cancer‐associated coding sequence mutation in FOXL2

et al. 2020

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Recent evidence suggests that animal microRNAs (miRNAs) can target coding sequences (CDSs); however, the pathophysiological importance of such targeting remains unknown. Here, we show that a somatic heterozygous missense mutation (c.402C>G; p.C134W) in FOXL2, a feature shared by virtually all adult-type granulosa cell tumors (AGCTs), introduces a target site for miR-1236, which causes haploinsufficiency of the tumor-suppressor FOXL2. This miR-1236-mediated selective degradation of the variant FOXL2 mRNA is preferentially conducted by a distinct miRNAloaded RNA-induced silencing complex (miRISC) directed by the Argonaute3 (AGO3) and DHX9 proteins. In both patients and a mouse model of AGCT, abundance of the inversely regulated variant FOXL2 with miR-1236 levels is highly correlated with malignant features of AGCT. Our study provides a molecular basis for understanding the conserved FOXL2 CDS mutation-mediated etiology of AGCT, revealing the existence of a previously unidentified mechanism of miRNA-targeting disease-associated mutations in the CDS by forming a non-canonical miRISC.

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Section: Methodsmentioning

confidence: 99%

An alternative miRISC targets a cancer‐associated coding sequence mutation in FOXL2

et al. 2020

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“…TM447 and W3110 PBAD eno strains were cultured in the same medium supplemented with 0.001% L-arabinose 24 . For anaerobic growth of E. coli cells, a 30 ml cylindrical bottle containing a sterilised stir bar was fully filled with LB medium with 0.2% glucose, sealed with sealing tape, and then cultured on a magnetic stirrer 55 . For aerobic–anaerobic–aerobic alternating experiments, the cells were grown aerobically to OD 600 ~0.15 (t0) as described above.…”

Section: Methodsmentioning

confidence: 99%

The coordinated action of RNase III and RNase G controls enolase expression in response to oxygen availability in Escherichia coli

Joo

Sim

et al. 2019

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Rapid modulation of RNA function by endoribonucleases during physiological responses to environmental changes is known to be an effective bacterial biochemical adaptation. We report a molecular mechanism underlying the regulation of enolase (eno) expression by two endoribonucleases, RNase G and RNase III, the expression levels of which are modulated by oxygen availability in Escherichia coli. Analyses of transcriptional eno-cat fusion constructs strongly suggested the existence of cis-acting elements in the eno 5′ untranslated region that respond to RNase III and RNase G cellular concentrations. Primer extension and S1 nuclease mapping analyses of eno mRNA in vivo identified three eno mRNA transcripts that are generated in a manner dependent on RNase III expression, one of which was found to accumulate in rng-deleted cells. Moreover, our data suggested that RNase III-mediated cleavage of primary eno mRNA transcripts enhanced Eno protein production, a process that involved putative cis-antisense RNA. We found that decreased RNase G protein abundance coincided with enhanced RNase III expression in E. coli grown anaerobically, leading to enhanced eno expression. Thereby, this posttranscriptional up-regulation of eno expression helps E. coli cells adjust their physiological reactions to oxygen-deficient metabolic modes. Our results revealed a molecular network of coordinated endoribonuclease activity that post-transcriptionally modulates the expression of Eno, a key enzyme in glycolysis.

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“…RNase G is a bacterial endoribonuclease known to negatively regulate gene expression via mRNA decay [9,[15][16][17]; however, SDS-PAGE and mass spectrometry revealed that some SPI-1 T3SS effectors (SlrP, SopA, SipA, and SipC) were up-regulated in the WT strain compared to that in the Δrng strain, indicating that changes in their mRNA levels were not a direct consequence of mRNA decay by RNase G. To assess this further, we analyzed upstream pathways to identify a SPI-1 T3SS repressor that can be negatively regulated by RNase G. When S. Typhimurium invades host cells or responds to elevated sucrose or salt concentrations, SPI-1 T3SS effector gene expression is activated by the major transcriptional activator HilA, which is generally repressed by H-NS [31,35,52]. To explore the relationships between sipA, sipC, hilA, and hns mRNA expression levels, real-time reverse transcription polymerase chain reaction (qRT-PCR) was performed using total RNA from WT cells cultured under aerobic and anaerobic conditions, since SPI-1 T3SS expression is known to be up-regulated during Salmonella growth under low oxygen conditions and during host cell infection [27,53].…”

Section: Effect Of Rnase G Levels On T3ss Activitymentioning

confidence: 99%

“…Since RNase E/G family proteins are known to influence mRNA half-life by cleaving the 5 0 untranslated region (UTR) of most biochemically characterised mRNA substrates [15][16][17]54], we performed primer extension analysis to identify the exact RNase G cleavage sites in the 5 0 UTR of hns mRNA. Total RNA extracted from the WT, Δrng, and Δrng comp strains cultivated under anaerobic conditions was used to synthesise cDNA, which formed three bands To demonstrate biochemically enzymatic cleavage of hns mRNA by RNase G, we carried out an in vitro cleavage assay using purified S. Typhimurium RNase G protein and synthetic RNA containing the full-length hns sequence.…”

Section: Effect Of Rnase G Levels On Hns Mrna Stabilitymentioning

confidence: 99%

“…Both enzymes are single-stranded RNA-specific endoribonucleases with a preference for cleaving AU-rich sequences [11,13,14]. RNase G participates in rRNA maturation and mRNA degradation [9,11,[15][16][17][18][19]. E. coli lacking rne can be made viable by overexpressing RNase G; however, rngcomplementation affects the abundance of a small portion of mRNAs in rne-deficient cells [9], indicating that RNase G and E play distinct physiological roles.…”

Section: Introductionmentioning

confidence: 99%

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Endoribonuclease-mediated control of hns mRNA stability constitutes a key regulatory pathway for Salmonella Typhimurium pathogenicity island 1 expression

et al. 2021

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Bacteria utilize endoribonuclease-mediated RNA processing and decay to rapidly adapt to environmental changes. Here, we report that the modulation of hns mRNA stability by the endoribonuclease RNase G plays a key role in Salmonella Typhimurium pathogenicity. We found that RNase G determines the half-life of hns mRNA by cleaving its 5′ untranslated region and that altering its cleavage sites by genome editing stabilizes hns mRNA, thus decreasing S. Typhimurium virulence in mice. Under anaerobic conditions, the FNR-mediated transcriptional repression of rnc encoding RNase III, which degrades rng mRNA, and simultaneous induction of rng transcription resulted in rapid hns mRNA degradation, leading to the derepression of genes involved in the Salmonella pathogenicity island 1 (SPI-1) type III secretion system (T3SS). Together, our findings show that RNase III and RNase G levels-mediated control of hns mRNA abundance acts as a regulatory pathway upstream of a complex feed-forward loop for SPI-1 expression.

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RNase G controls tpiA mRNA abundance in response to oxygen availability in Escherichia coli

Cited by 10 publications

References 40 publications

An alternative miRISC targets a cancer‐associated coding sequence mutation in FOXL2

An alternative miRISC targets a cancer‐associated coding sequence mutation in FOXL2

The coordinated action of RNase III and RNase G controls enolase expression in response to oxygen availability in Escherichia coli

Endoribonuclease-mediated control of hns mRNA stability constitutes a key regulatory pathway for Salmonella Typhimurium pathogenicity island 1 expression

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