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Silencing of DNase Colicin E8 Gene Expression by a Complex Nucleoprotein Assembly Ensures Timely Colicin Induction.

Kamenšek S, Browning DF, Podlesek Z, Busby SJ, Žgur-Bertok D, Butala M - PLoS Genet. (2015)

Bottom Line: We show that L-asparagine is an environmental factor that has a marked impact on cea8 promoter regulation.Our results show that AsnC also modulates the expression of several other DNase and RNase colicin genes but does not substantially affect pore-forming colicin K gene expression.We propose that selection pressure has "chosen" highly conserved regulators to control colicin expression in E. coli strains, enabling similar colicin gene silencing among bacteria upon exchange of colicinogenic plasmids.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia.

ABSTRACT
Colicins are plasmid-encoded narrow spectrum antibiotics that are synthesized by strains of Escherichia coli and govern intraspecies competition. In a previous report, we demonstrated that the global transcriptional factor IscR, co dependently with the master regulator of the DNA damage response, LexA, delays induction of the pore forming colicin genes after SOS induction. Here we show that IscR is not involved in the regulation of nuclease colicins, but that the AsnC protein is. We report that AsnC, in concert with LexA, is the key controller of the temporal induction of the DNA degrading colicin E8 gene (cea8), after DNA damage. We demonstrate that a large AsnC nucleosome-like structure, in conjunction with two LexA molecules, prevent cea8 transcription initiation and that AsnC binding activity is directly modulated by L asparagine. We show that L-asparagine is an environmental factor that has a marked impact on cea8 promoter regulation. Our results show that AsnC also modulates the expression of several other DNase and RNase colicin genes but does not substantially affect pore-forming colicin K gene expression. We propose that selection pressure has "chosen" highly conserved regulators to control colicin expression in E. coli strains, enabling similar colicin gene silencing among bacteria upon exchange of colicinogenic plasmids.

No MeSH data available.


Related in: MedlinePlus

The cae8 promoter is sensitive to the amino acid L-asparagine.The figure shows measured β-galactosidase activities from wild-type BW25113 cells, carrying the cea8 promoter region subcloned into pRW50. Cells were grown in M9 minimal medium containing 0.5 mM NH4Cl until an OD600 of ~0.2, when the culture was split in to two and grown further in the presence of either 10 mM NH4Cl or 20 mM L-asparagine. The arrow indicates the time of addition of nalidixic acid and the dashed lines represent OD600. Each value is the average of duplicate experiments and the standard deviation is shown.
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pgen.1005354.g005: The cae8 promoter is sensitive to the amino acid L-asparagine.The figure shows measured β-galactosidase activities from wild-type BW25113 cells, carrying the cea8 promoter region subcloned into pRW50. Cells were grown in M9 minimal medium containing 0.5 mM NH4Cl until an OD600 of ~0.2, when the culture was split in to two and grown further in the presence of either 10 mM NH4Cl or 20 mM L-asparagine. The arrow indicates the time of addition of nalidixic acid and the dashed lines represent OD600. Each value is the average of duplicate experiments and the standard deviation is shown.

Mentions: Our data suggest that tight repression of DNase colicin E8 might be affected by the availability of amino acid L-asparagine and this signal is relayed via AsnC. To test this hypothesis we measured cea8 promoter activity in SOS-induced wild-type cells grown in the M9 minimal medium containing either 10 mM NH4Cl or 20 mM L-asparagine as the sole source of nitrogen. Fig 5 shows that in the L-asparagine containing medium, the expression from cea8 remains low, whilst it increases in medium containing higher levels of NH4Cl. This is in agreement with our in vitro data (Figs 3 and 4) and suggests that L-asparagine is needed to stabilize a specific AsnC assembly at the cea8 promoter region. Hence, we suggest that depletion of L-asparagine is the signal for AsnC de-repression at the cea8 promoter.


Silencing of DNase Colicin E8 Gene Expression by a Complex Nucleoprotein Assembly Ensures Timely Colicin Induction.

Kamenšek S, Browning DF, Podlesek Z, Busby SJ, Žgur-Bertok D, Butala M - PLoS Genet. (2015)

The cae8 promoter is sensitive to the amino acid L-asparagine.The figure shows measured β-galactosidase activities from wild-type BW25113 cells, carrying the cea8 promoter region subcloned into pRW50. Cells were grown in M9 minimal medium containing 0.5 mM NH4Cl until an OD600 of ~0.2, when the culture was split in to two and grown further in the presence of either 10 mM NH4Cl or 20 mM L-asparagine. The arrow indicates the time of addition of nalidixic acid and the dashed lines represent OD600. Each value is the average of duplicate experiments and the standard deviation is shown.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4482635&req=5

pgen.1005354.g005: The cae8 promoter is sensitive to the amino acid L-asparagine.The figure shows measured β-galactosidase activities from wild-type BW25113 cells, carrying the cea8 promoter region subcloned into pRW50. Cells were grown in M9 minimal medium containing 0.5 mM NH4Cl until an OD600 of ~0.2, when the culture was split in to two and grown further in the presence of either 10 mM NH4Cl or 20 mM L-asparagine. The arrow indicates the time of addition of nalidixic acid and the dashed lines represent OD600. Each value is the average of duplicate experiments and the standard deviation is shown.
Mentions: Our data suggest that tight repression of DNase colicin E8 might be affected by the availability of amino acid L-asparagine and this signal is relayed via AsnC. To test this hypothesis we measured cea8 promoter activity in SOS-induced wild-type cells grown in the M9 minimal medium containing either 10 mM NH4Cl or 20 mM L-asparagine as the sole source of nitrogen. Fig 5 shows that in the L-asparagine containing medium, the expression from cea8 remains low, whilst it increases in medium containing higher levels of NH4Cl. This is in agreement with our in vitro data (Figs 3 and 4) and suggests that L-asparagine is needed to stabilize a specific AsnC assembly at the cea8 promoter region. Hence, we suggest that depletion of L-asparagine is the signal for AsnC de-repression at the cea8 promoter.

Bottom Line: We show that L-asparagine is an environmental factor that has a marked impact on cea8 promoter regulation.Our results show that AsnC also modulates the expression of several other DNase and RNase colicin genes but does not substantially affect pore-forming colicin K gene expression.We propose that selection pressure has "chosen" highly conserved regulators to control colicin expression in E. coli strains, enabling similar colicin gene silencing among bacteria upon exchange of colicinogenic plasmids.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia.

ABSTRACT
Colicins are plasmid-encoded narrow spectrum antibiotics that are synthesized by strains of Escherichia coli and govern intraspecies competition. In a previous report, we demonstrated that the global transcriptional factor IscR, co dependently with the master regulator of the DNA damage response, LexA, delays induction of the pore forming colicin genes after SOS induction. Here we show that IscR is not involved in the regulation of nuclease colicins, but that the AsnC protein is. We report that AsnC, in concert with LexA, is the key controller of the temporal induction of the DNA degrading colicin E8 gene (cea8), after DNA damage. We demonstrate that a large AsnC nucleosome-like structure, in conjunction with two LexA molecules, prevent cea8 transcription initiation and that AsnC binding activity is directly modulated by L asparagine. We show that L-asparagine is an environmental factor that has a marked impact on cea8 promoter regulation. Our results show that AsnC also modulates the expression of several other DNase and RNase colicin genes but does not substantially affect pore-forming colicin K gene expression. We propose that selection pressure has "chosen" highly conserved regulators to control colicin expression in E. coli strains, enabling similar colicin gene silencing among bacteria upon exchange of colicinogenic plasmids.

No MeSH data available.


Related in: MedlinePlus