Maximization of transcription of the serC (pdxF)-aroA multifunctional operon by antagonistic effects of the cyclic AMP (cAMP) receptor protein-cAMP complex and Lrp global regulators of Escherichia coli K-12

Man, Tsz-Kwong; Pease, A J; Winkler, Malcolm E.

doi:10.1128/jb.179.11.3458-3469.1997

Cited by 20 publications

(22 citation statements)

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“…␤-Galactosidase specific activities in exponentially growing (50K) or stationary-phase (24-h) cultures were measured with a plate reader as described before (43). Differential rates of ␤-galactosidase synthesis were determined from plots of total ␤-galactosidase activity per milliliter of culture per reaction time graphed against cell mass (A 600 ) as described previously (43).…”

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confidence: 99%

Negative regulation of mutS and mutH repair gene expression by the Hfq and RpoS global regulators of Escherichia coli K-12

1997

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The MutS, MutL, and MutH proteins play major roles in several DNA repair pathways. We previously reported that the cellular amounts of MutS and MutH decreased by as much as 10-fold in stationary-phase cultures. Consequently, we tested whether the amounts of MutS, MutL, and MutH were regulated by two global regulators, RpoS ( 38 ) and Hfq (HF-I [putative RNA chaperone]), which are involved in stationary-phase transition. We report here that mutations in hfq and rpoS reversed the stationary-phase down-regulation of the amounts of MutS and MutH. hfq regulation of the amount of MutS in stationary-phase cultures was mediated by RpoS-dependent and -independent mechanisms, whereas hfq regulation of the amount of MutH was mediated only through RpoS. Consistent with this interpretation, the amount of MutS but not MutH was regulated by Hfq, but not RpoS, in exponentially growing cells. The amount of MutL remained unchanged in rpoS, hfq-1, and rpoS ؉ , hfq ؉ strains in exponentially growing and stationary-phase cultures and served as a control. The ␤-galactosidase activities of single-copy mutS-lacZ operon and gene fusions suggested that hfq regulates mutS posttranscriptionally in exponentially growing cultures. RNase T 2 protection assays revealed increased amounts of mutS transcript that are attributed to increased mutS transcript stability in hfq-1 mutants. Lack of Hfq also increased the amounts and stabilities of transcripts initiated from P miaA and P1 hfq HS, two of the promoters for hfq, suggesting autoregulation, but did not change the half-life of bulk mRNA. These results suggest that the amounts of MutS and MutH may be adjusted in cells subjected to different stress conditions by an RpoS-dependent mechanism. In addition, Hfq directly or indirectly regulates several genes, including mutS, hfq, and miaA, by an RpoS-independent mechanism that destabilizes transcripts.The MutS, MutL, and MutH proteins of Escherichia coli play important roles in several DNA repair pathways and the maintenance of chromosomal stability (51, 53). In the methyl-directed-mismatch (MDM) repair pathway, MutS, MutL, and MutH function as a DNA mismatch binding protein (75, 76), a putative molecular matchmaker (66), and a d(GATC)-specific endonuclease (83), respectively. MDM repair is a major pathway that sets spontaneous mutation rates by correcting mismatched base pairs and small single-strand loops that arise as replication errors (9,60,75). In addition, MutL and MutS play roles in very-short-patch repair (38, 40), transcription-coupled nucleotide excision repair (48), and the prevention of homeologous recombination (61, 62). Homologs of E. coli MutL and MutS have been found in most other bacteria and in eukaryotes (26, 51, 53), in which they play crucial roles in DNA repair, in maintaining chromosome stability, in meiotic recombination, and in preventing colon and sporadic cancers (31,52,53).Direct measurements showed that MutS and MutL proteins are present at about 186 and 113 dimers, respectively, per exponentially growing cell (15), wher...

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Negative regulation of mutS and mutH repair gene expression by the Hfq and RpoS global regulators of Escherichia coli K-12

1997

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“…To fully understand the control and integration of PLP biosynthesis, we have been determining whether the individual PLP biosynthetic genes are subject to forms of metabolic regulation. Previously, we reported that the serC (pdxF) gene is positively regulated by the Lrp protein and negatively regulated by the cyclic AMP receptor protein (43). Here we report that transcription of the pdxA and pdxB genes is positively growth rate regulated by two different mechanisms.…”

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confidence: 61%

“…Moreover, other PLP biosynthetic genes do not follow the pattern of positive growth rate regulation reported here for pdxA and pdxB. For example, expression of the serC (pdxF)-encoded transaminase, which acts between PdxB and PdxA in the de novo PLP pathway (40,43), is minimal in rich LB medium and maximal in minimal salts-glucose medium due to opposing positive and negative controls exerted by the Lrp and cyclic AMP receptor proteins, respectively (T.-K. Man and M. E. Winkler, unpublished results) (43). However, the regulation of the SerC (PdxF) transaminase may have been strongly set by its central role in serine biosynthesis (63).…”

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confidence: 99%

Positive Growth Rate-Dependent Regulation of the pdxA , ksgA , and pdxB Genes of Escherichia coli K-12

et al. 2002

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We found that transcription of the pdxA and pdxB genes, which mediate steps in the biosynthesis of the essential coenzyme pyridoxal 5-phosphate, and the ksgA gene, which encodes an rRNA modification enzyme and is partly cotranscribed with pdxA, is subject to positive growth rate regulation in Escherichia coli K-12. The amounts of the pdxA-ksgA cotranscript and pdxB-and ksgA-specific transcripts and expression from pdxA-and pdxB-lacZ fusions increased as the growth rate increased. The half-lives of ksgA-and pdxB-specific transcripts were not affected by the growth rate, whereas the half-life of the pdxA-ksgA cotranscript was too short to be measured accurately. A method of normalization was applied to determine the amount of mRNA synthesized per gene and the rate of protein accumulation per gene. Normalization removed an apparent anomaly at fast growth rates and demonstrated that positive regulation of pdxB occurs at the level of transcription initiation over the whole range of growth rates tested. RNA polymerase limitation and autoregulation could not account for the positive growth rate regulation of pdxA, pdxB, and ksgA transcription. On the other hand, growth rate regulation of the amount of the pdxA-ksgA cotranscript was abolished by a fis mutation, suggesting a role for the Fis protein. In contrast, the fis mutation had no effect on pdxB-or ksgA-specific transcript amounts. The amounts of the pdxA-ksgA cotranscript and ksgA-specific transcript were repressed in the presence of high intracellular concentrations of guanosine tetraphosphate; however, this effect was independent of relA function for the pdxA-ksgA cotranscript. Amounts of the pdxB-specific transcript remained unchanged during amino acid starvation in wild-type and relA mutant strains.

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“…The general idea predicts that all isoprenoid biosynthetic genes are in multigene operons with essential distal genes. This prediction has not yet been tested, but such mixedfunction operons are known (17,19,34).…”

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confidence: 99%

Lethal Mutations in the Isoprenoid Pathway of Salmonella enterica

2006

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Essential isoprenoid compounds are synthesized using the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway in many gram-negative bacteria, some gram-positive bacteria, some apicomplexan parasites, and plant chloroplasts. The alternative mevalonate pathway is found in archaea and eukaryotes, including cytosolic biosynthesis in plants. The existence of orthogonal essential pathways in eukaryotes and bacteria makes the MEP pathway an attractive target for the development of antimicrobial agents. A system is described for identifying mutations in the MEP pathway of Salmonella enterica serovar Typhimurium. Using this system, point mutations induced by diethyl sulfate were found in the all genes of the essential MEP pathway and also in genes involved in uptake of methylerythritol. Curiously, none of the MEP pathway genes could be identified in the same parent strain by transposon mutagenesis, despite extensive searches. The results complement the biochemical and bioinformatic approaches to the elucidation of the genes involved in the MEP pathway and also identify key residues for activity in the enzymes of the pathway.It was recently discovered that bacteria synthesize isoprenoids by a pathway that differs from that found in eukaryotes. In bacteria, pyruvate and glyceraldehyde 3-phosphate are converted, through the 2-C-methyl-D-erythritol phosphate (MEP) pathway, to isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). This has focused recent research on the elucidation of the biosynthetic steps and genes encoding the catalytic enzymes in bacteria. To date, all synthetic steps of the MEP pathway and the genes encoding the biosynthetic enzymes have been identified. Isoprenoid biosynthesis begins with the condensation of pyruvate and glyceraldehyde 3-phosphate by 1-deoxy-D-xylulose 5-phosphate (DXP) synthase, encoded by dxs (26). Next, in the first committed step of the pathway, DXP reductoisomerase, encoded by dxr, converts DXP to MEP (16,32). In the following three steps, a cytidyl monophosphate moiety is attached to the phosphate of MEP; the resulting diphosphodiester is phosphorylated at the C-2 hydroxyl in the methylerythritol moiety and then cyclized to form an eight-membered cyclic diphosphate diester, and these steps are catalyzed by the proteins encoded by ispD (25), ispE (18), and ispF (11), respectively. The enzyme encoded by ispG, 1-hydroxy-2-methyl-2-(E)-butenol 4-diphosphate (HDMAPP) synthase, catalyzes the ring opening and reduction of 2-Cmethyl-D-erythritol-2,4-cyclodiphosphate to yield HDMAPP (10). Finally, HDMAPP is converted to a mixture of IPP and DMAPP by HDMAPP reductase encoded by ispH (1).Mutants blocked in the MEP pathway of Salmonella spp. or Escherichia coli are expected to be lethal, since these organisms are unable to utilize exogenously supplied IPP, DMAPP, or their corresponding alcohols. To allow viability of such mutants, genes of the alternative mevalonate pathway were introduced into bacteria, either on plasmids or in the chromosome (2,5,9,15,24). Strains containing the ...

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Maximization of transcription of the serC (pdxF)-aroA multifunctional operon by antagonistic effects of the cyclic AMP (cAMP) receptor protein-cAMP complex and Lrp global regulators of Escherichia coli K-12

Cited by 20 publications

References 57 publications

Negative regulation of mutS and mutH repair gene expression by the Hfq and RpoS global regulators of Escherichia coli K-12

Negative regulation of mutS and mutH repair gene expression by the Hfq and RpoS global regulators of Escherichia coli K-12

Positive Growth Rate-Dependent Regulation of the pdxA , ksgA , and pdxB Genes of Escherichia coli K-12

Lethal Mutations in the Isoprenoid Pathway of Salmonella enterica

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