Limits...
Intermediate Levels of Bacillus subtilis CodY Activity Are Required for Derepression of the Branched-Chain Amino Acid Permease, BraB.

Belitsky BR, Brinsmade SR, Sonenshein AL - PLoS Genet. (2015)

Bottom Line: However, under conditions of reduced CodY activity, CodY-mediated repression was relieved to a greater extent than ScoC-mediated repression was increased, leading to elevated braB expression.We conclude that restricting increased expression of braB to conditions of moderate nutrient limitation is the raison d'être of the feed-forward regulatory loop formed by CodY and ScoC at the braB promoter.The increase in BraB expression only at intermediate activities of CodY may facilitate the uptake of BCAA when they are not in excess but prevent unneeded BraB synthesis when other BCAA transporters are active.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts, United States of America.

ABSTRACT
The global transcriptional regulator, CodY, binds strongly to the regulatory region of the braB gene, which encodes a Bacillus subtilis branched-chain amino acid (BCAA) permease. However, under conditions that maximize CodY activity, braB expression was similar in wild-type and codY mutant cells. Nonetheless, expression from the braB promoter was significantly elevated in cells containing partially active mutant versions of CodY or in wild-type cells under growth conditions leading to intermediate levels of CodY activity. This novel pattern of regulation was shown to be due to two opposing mechanisms, negative and positive, by which CodY affects braB expression. A strong CodY-binding site located downstream of the transcription start point conferred negative regulation by direct interaction with CodY. Additionally, sequences upstream and downstream of the promoter were required for repression by a second pleiotropic B. subtilis regulator, ScoC, whose own expression is repressed by CodY. ScoC-mediated repression of braB in codY mutants cells was as efficient as direct, CodY-mediated repression in wild-type cells under conditions of high CodY activity. However, under conditions of reduced CodY activity, CodY-mediated repression was relieved to a greater extent than ScoC-mediated repression was increased, leading to elevated braB expression. We conclude that restricting increased expression of braB to conditions of moderate nutrient limitation is the raison d'ĂȘtre of the feed-forward regulatory loop formed by CodY and ScoC at the braB promoter. The increase in BraB expression only at intermediate activities of CodY may facilitate the uptake of BCAA when they are not in excess but prevent unneeded BraB synthesis when other BCAA transporters are active.

No MeSH data available.


Related in: MedlinePlus

Expression of the braB242-gfp fusion in individual cells.Cells were grown in TSS + 16 aa medium. GFP fluorescence was detected using Zeiss Axio Observer.Z1 microscope. A. Strain BB4082 (wild type). B. Strain BB4083 (codY::spc). C. Strain BB4084 [codY(R61K)].
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4608796&req=5

pgen.1005600.g006: Expression of the braB242-gfp fusion in individual cells.Cells were grown in TSS + 16 aa medium. GFP fluorescence was detected using Zeiss Axio Observer.Z1 microscope. A. Strain BB4082 (wild type). B. Strain BB4083 (codY::spc). C. Strain BB4084 [codY(R61K)].

Mentions: A braB242-gfp translational fusion was introduced into the wild-type strain, the codY mutant, and a codY point mutant (R61K) strain with intermediate residual activity. In all cases, the level of braB expression was rather similar across the cell population (Fig 6), eliminating the possibility that a bistable expression pattern could explain our results. As expected, the codY (R61K) mutant strain had elevated expression compared to the wild-type and codY mutant strains.


Intermediate Levels of Bacillus subtilis CodY Activity Are Required for Derepression of the Branched-Chain Amino Acid Permease, BraB.

Belitsky BR, Brinsmade SR, Sonenshein AL - PLoS Genet. (2015)

Expression of the braB242-gfp fusion in individual cells.Cells were grown in TSS + 16 aa medium. GFP fluorescence was detected using Zeiss Axio Observer.Z1 microscope. A. Strain BB4082 (wild type). B. Strain BB4083 (codY::spc). C. Strain BB4084 [codY(R61K)].
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4608796&req=5

pgen.1005600.g006: Expression of the braB242-gfp fusion in individual cells.Cells were grown in TSS + 16 aa medium. GFP fluorescence was detected using Zeiss Axio Observer.Z1 microscope. A. Strain BB4082 (wild type). B. Strain BB4083 (codY::spc). C. Strain BB4084 [codY(R61K)].
Mentions: A braB242-gfp translational fusion was introduced into the wild-type strain, the codY mutant, and a codY point mutant (R61K) strain with intermediate residual activity. In all cases, the level of braB expression was rather similar across the cell population (Fig 6), eliminating the possibility that a bistable expression pattern could explain our results. As expected, the codY (R61K) mutant strain had elevated expression compared to the wild-type and codY mutant strains.

Bottom Line: However, under conditions of reduced CodY activity, CodY-mediated repression was relieved to a greater extent than ScoC-mediated repression was increased, leading to elevated braB expression.We conclude that restricting increased expression of braB to conditions of moderate nutrient limitation is the raison d'être of the feed-forward regulatory loop formed by CodY and ScoC at the braB promoter.The increase in BraB expression only at intermediate activities of CodY may facilitate the uptake of BCAA when they are not in excess but prevent unneeded BraB synthesis when other BCAA transporters are active.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts, United States of America.

ABSTRACT
The global transcriptional regulator, CodY, binds strongly to the regulatory region of the braB gene, which encodes a Bacillus subtilis branched-chain amino acid (BCAA) permease. However, under conditions that maximize CodY activity, braB expression was similar in wild-type and codY mutant cells. Nonetheless, expression from the braB promoter was significantly elevated in cells containing partially active mutant versions of CodY or in wild-type cells under growth conditions leading to intermediate levels of CodY activity. This novel pattern of regulation was shown to be due to two opposing mechanisms, negative and positive, by which CodY affects braB expression. A strong CodY-binding site located downstream of the transcription start point conferred negative regulation by direct interaction with CodY. Additionally, sequences upstream and downstream of the promoter were required for repression by a second pleiotropic B. subtilis regulator, ScoC, whose own expression is repressed by CodY. ScoC-mediated repression of braB in codY mutants cells was as efficient as direct, CodY-mediated repression in wild-type cells under conditions of high CodY activity. However, under conditions of reduced CodY activity, CodY-mediated repression was relieved to a greater extent than ScoC-mediated repression was increased, leading to elevated braB expression. We conclude that restricting increased expression of braB to conditions of moderate nutrient limitation is the raison d'ĂȘtre of the feed-forward regulatory loop formed by CodY and ScoC at the braB promoter. The increase in BraB expression only at intermediate activities of CodY may facilitate the uptake of BCAA when they are not in excess but prevent unneeded BraB synthesis when other BCAA transporters are active.

No MeSH data available.


Related in: MedlinePlus