Transcript analysis reveals an extended regulon and the importance of protein–protein co-operativity for the <i>Escherichia coli</i> methionine repressor

Marincs, Ferenc; Manfield, Iain W.; Stead, Jonathan A.; McDowall, Kenneth J.; Stockley, Peter G.

doi:10.1042/bj20060021

Cited by 46 publications

(42 citation statements)

References 45 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The tolerance associated with mutations in metJ, rho, and rpsQ implicated translation and transcription as targets of ethanol. MetJ represses expression of genes involved in methionine biosynthesis (31). Rho terminates transcription uncoupled from translation, halting mRNA synthesis when translation terminates prematurely (29).…”

Section: Resultsmentioning

confidence: 99%

Correcting direct effects of ethanol on translation and transcription machinery confers ethanol tolerance in bacteria

Haft

Keating

Schwaegler

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

130

121

View full text Add to dashboard Cite

The molecular mechanisms of ethanol toxicity and tolerance in bacteria, although important for biotechnology and bioenergy applications, remain incompletely understood. Genetic studies have identified potential cellular targets for ethanol and have revealed multiple mechanisms of tolerance, but it remains difficult to separate the direct and indirect effects of ethanol. We used adaptive evolution to generate spontaneous ethanol-tolerant strains of Escherichia coli, and then characterized mechanisms of toxicity and resistance using genome-scale DNAseq, RNAseq, and ribosome profiling coupled with specific assays of ribosome and RNA polymerase function. Evolved alleles of metJ, rho, and rpsQ recapitulated most of the observed ethanol tolerance, implicating translation and transcription as key processes affected by ethanol. Ethanol induced miscoding errors during protein synthesis, from which the evolved rpsQ allele protected cells by increasing ribosome accuracy. Ribosome profiling and RNAseq analyses established that ethanol negatively affects transcriptional and translational processivity. Ethanol-stressed cells exhibited ribosomal stalling at internal AUG codons, which may be ameliorated by the adaptive inactivation of the MetJ repressor of methionine biosynthesis genes. Ethanol also caused aberrant intragenic transcription termination for mRNAs with low ribosome density, which was reduced in a strain with the adaptive rho mutation. Furthermore, ethanol inhibited transcript elongation by RNA polymerase in vitro. We propose that ethanol-induced inhibition and uncoupling of mRNA and protein synthesis through direct effects on ribosomes and RNA polymerase conformations are major contributors to ethanol toxicity in E. coli, and that adaptive mutations in metJ, rho, and rpsQ help protect these central dogma processes in the presence of ethanol.

show abstract

Section: Resultsmentioning

confidence: 99%

Correcting direct effects of ethanol on translation and transcription machinery confers ethanol tolerance in bacteria

Haft

Keating

Schwaegler

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

130

121

View full text Add to dashboard Cite

show abstract

“…190, 2008 PRE-Q 0 BIOSYNTHESIS 7881 on May 11, 2018 by guest http://jb.asm.org/ ulatory protein (24), but this is not observed in bacteria as they lack this protein, and the overexpression of folate or BH4 biosynthetic genes results in overproduction of the final products (52,57). In E. coli, the expression of the folE gene has been shown to be under the control of the negative regulator MetJ (30). The folE1 gene (mtrA) of B. subtilis is in an operon with the TRAP-encoding gene (mtrB).…”

Section: Discussionmentioning

confidence: 99%

Biosynthesis of 7-Deazaguanosine-Modified tRNA Nucleosides: a New Role for GTP Cyclohydrolase I

et al. 2008

View full text Add to dashboard Cite

Queuosine (Q) and archaeosine (G ؉ ) are hypermodified ribonucleosides found in tRNA. Q is present in the anticodon region of tRNA GUN in Eukarya and Bacteria, while G ؉ is found at position 15 in the D-loop of archaeal tRNA. Prokaryotes produce these 7-deazaguanosine derivatives de novo from GTP through the 7-cyano-7-deazaguanine (pre-Q 0 ) intermediate, but mammals import the free base, queuine, obtained from the diet or the intestinal flora. By combining the results of comparative genomic analysis with those of genetic studies, we show that the first enzyme of the folate pathway, GTP cyclohydrolase I (GCYH-I), encoded in Escherichia coli by folE, is also the first enzyme of pre-Q 0 biosynthesis in both prokaryotic kingdoms. Indeed, tRNA extracted from an E. coli ⌬folE strain is devoid of Q and the deficiency is complemented by expressing GCYH-I-encoding genes from different bacterial or archaeal origins. In a similar fashion, tRNA extracted from a Haloferax volcanii strain carrying a deletion of the GCYH-I-encoding gene contains only traces of G ؉ . These results link the production of a tRNA-modified base to primary metabolism and further clarify the biosynthetic pathway for these complex modified nucleosides.It is well established that GTP is not only a molecule central to energy conservation and a precursor to the synthesis of RNA and DNA but also the precursor to a number of essential metabolites. These include riboflavin and the pterin-related coenzymes tetrahydropterin (BH4), tetrahydrofolate (THF), methanopterin, and molybdopterin. It is less well known that GTP is also the precursor of the 7-deazapurine derivatives found in tRNA (queuosine [Q] and archaeosine

show abstract

“…5). Derepression of methionine and SAM biosynthesis in a metJ knockout strain (JW3909; derivative of BW25113) (30,31) resulted in higher levels of repression, further indicating that the magnitude of repression correlates with intracellular SAM (Fig. S4B).…”

Section: Sam Binding Stabilizes the Pk-2 Subdomain Required For Regulmentioning

confidence: 98%

Structural basis for diversity in the SAM clan of riboswitches

Trausch¹,

Xu²,

Edwards³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

In bacteria, sulfur metabolism is regulated in part by seven known families of riboswitches that bind S-adenosyl-L-methionine (SAM). Direct binding of SAM to these mRNA regulatory elements governs a downstream secondary structural switch that communicates with the transcriptional and/or translational expression machinery. The most widely distributed SAM-binding riboswitches belong to the SAM clan, comprising three families that share a common SAMbinding core but differ radically in their peripheral architecture. Although the structure of the SAM-I member of this clan has been extensively studied, how the alternative peripheral architecture of the other families supports the common SAM-binding core remains unknown. We have therefore solved the X-ray structure of a member of the SAM-I/IV family containing the alternative "PK-2" subdomain shared with the SAM-IV family. This structure reveals that this subdomain forms extensive interactions with the helix housing the SAM-binding pocket, including a highly unusual mode of helix packing in which two helices pack in a perpendicular fashion. Biochemical and genetic analysis of this RNA reveals that SAM binding induces many of these interactions, including stabilization of a pseudoknot that is part of the regulatory switch. Despite strong structural similarity between the cores of SAM-I and SAM-I/IV members, a phylogenetic analysis of sequences does not indicate that they derive from a common ancestor.RNA structure | X-ray crystallography | chemical probing | isothermal titration calorimetry | gene regulation R iboswitches are noncoding RNA elements generally found in the leader of bacterial mRNAs that regulate expression via direct binding of a specific cellular metabolite (reviewed in refs.

show abstract

Transcript analysis reveals an extended regulon and the importance of protein–protein co-operativity for the Escherichia coli methionine repressor

Cited by 46 publications

References 45 publications

Correcting direct effects of ethanol on translation and transcription machinery confers ethanol tolerance in bacteria

Correcting direct effects of ethanol on translation and transcription machinery confers ethanol tolerance in bacteria

Biosynthesis of 7-Deazaguanosine-Modified tRNA Nucleosides: a New Role for GTP Cyclohydrolase I

Structural basis for diversity in the SAM clan of riboswitches

Contact Info

Product

Resources

About