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Quantitative characteristics of gene regulation by small RNA.

Levine E, Zhang Z, Kuhlman T, Hwa T - PLoS Biol. (2007)

Bottom Line: Our results demonstrate that sRNA provides a novel mode of gene regulation, with characteristics distinct from those of protein-mediated gene regulation.Knowledge of these special features of sRNA-mediated regulation may be crucial toward understanding the subtle functions that sRNAs can play in coordinating various stress-relief pathways.Our results may also help guide the design of synthetic genetic circuits that have properties difficult to attain with protein regulators alone.

View Article: PubMed Central - PubMed

Affiliation: Center for Theoretical Biological Physics, University of California San Diego, La Jolla, California, United States of America.

ABSTRACT
An increasing number of small RNAs (sRNAs) have been shown to regulate critical pathways in prokaryotes and eukaryotes. In bacteria, regulation by trans-encoded sRNAs is predominantly found in the coordination of intricate stress responses. The mechanisms by which sRNAs modulate expression of its targets are diverse. In common to most is the possibility that interference with the translation of mRNA targets may also alter the abundance of functional sRNAs. Aiming to understand the unique role played by sRNAs in gene regulation, we studied examples from two distinct classes of bacterial sRNAs in Escherichia coli using a quantitative approach combining experiment and theory. Our results demonstrate that sRNA provides a novel mode of gene regulation, with characteristics distinct from those of protein-mediated gene regulation. These include a threshold-linear response with a tunable threshold, a robust noise resistance characteristic, and a built-in capability for hierarchical cross-talk. Knowledge of these special features of sRNA-mediated regulation may be crucial toward understanding the subtle functions that sRNAs can play in coordinating various stress-relief pathways. Our results may also help guide the design of synthetic genetic circuits that have properties difficult to attain with protein regulators alone.

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Threshold-Linear Response of a Reporter Target of RNA-OUT(A) GFP expressions of strain ZZS35 (is10out+) are plotted on the vertical axis against the promoter activity, defined (as in Figure 2) as the GFP expression of strain ZZS31 (is10out−) grown in identical medium. The different symbols correspond to the different levels of RNA-OUT expressed by the PLtet-O1 promoter. The latter was controlled by varying amounts of aTc added to the growth medium (see legend). The solid lines are the steady-state solution (Equation 2) using the best-fit parameters listed in Table S2.(B) Ratio of GFP expression in is10out+ (ZZS35) and is10out− (ZZS31) strains measured through GFP fluorescence (solid bars) and RT-PCR (striped bars). Expression of RNA-OUT was induced by 10 ng/ml aTc (red), whereas target expression was induced with 0.3 mM IPTG [corresponding to the 4th diamond from the right in (A)]. Change in the mRNA level of cris10-gfp (striped bars) is insignificant compared with changes in GFP fluorescence (solid bars).
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pbio-0050229-g003: Threshold-Linear Response of a Reporter Target of RNA-OUT(A) GFP expressions of strain ZZS35 (is10out+) are plotted on the vertical axis against the promoter activity, defined (as in Figure 2) as the GFP expression of strain ZZS31 (is10out−) grown in identical medium. The different symbols correspond to the different levels of RNA-OUT expressed by the PLtet-O1 promoter. The latter was controlled by varying amounts of aTc added to the growth medium (see legend). The solid lines are the steady-state solution (Equation 2) using the best-fit parameters listed in Table S2.(B) Ratio of GFP expression in is10out+ (ZZS35) and is10out− (ZZS31) strains measured through GFP fluorescence (solid bars) and RT-PCR (striped bars). Expression of RNA-OUT was induced by 10 ng/ml aTc (red), whereas target expression was induced with 0.3 mM IPTG [corresponding to the 4th diamond from the right in (A)]. Change in the mRNA level of cris10-gfp (striped bars) is insignificant compared with changes in GFP fluorescence (solid bars).

Mentions: As a controlled source of RNA-OUT, we used the pZA31O plasmid, which harbors the is10out gene driven by the strong synthetic PLtet-O1 promoter [62]. We measured the response function at four different expression levels of RNA-OUT using different concentrations of the inducer aTc (0, 2, 6, and 10 ng/ml). The data obtained (symbols in Figure 3A) were fitted to the steady-state solution (Equation 2) as described above; best-fit parameters are given in Table S2. The fitted curves are presented as the solid lines in Figure 3A. In the absence of aTc, cris10-gfp expression coincides with that of the corresponding strain with no RNA-OUT source (dashed black line). At higher levels of aTc, the threshold-linear response is recovered, displaying a smooth transition as expected.


Quantitative characteristics of gene regulation by small RNA.

Levine E, Zhang Z, Kuhlman T, Hwa T - PLoS Biol. (2007)

Threshold-Linear Response of a Reporter Target of RNA-OUT(A) GFP expressions of strain ZZS35 (is10out+) are plotted on the vertical axis against the promoter activity, defined (as in Figure 2) as the GFP expression of strain ZZS31 (is10out−) grown in identical medium. The different symbols correspond to the different levels of RNA-OUT expressed by the PLtet-O1 promoter. The latter was controlled by varying amounts of aTc added to the growth medium (see legend). The solid lines are the steady-state solution (Equation 2) using the best-fit parameters listed in Table S2.(B) Ratio of GFP expression in is10out+ (ZZS35) and is10out− (ZZS31) strains measured through GFP fluorescence (solid bars) and RT-PCR (striped bars). Expression of RNA-OUT was induced by 10 ng/ml aTc (red), whereas target expression was induced with 0.3 mM IPTG [corresponding to the 4th diamond from the right in (A)]. Change in the mRNA level of cris10-gfp (striped bars) is insignificant compared with changes in GFP fluorescence (solid bars).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC1994261&req=5

pbio-0050229-g003: Threshold-Linear Response of a Reporter Target of RNA-OUT(A) GFP expressions of strain ZZS35 (is10out+) are plotted on the vertical axis against the promoter activity, defined (as in Figure 2) as the GFP expression of strain ZZS31 (is10out−) grown in identical medium. The different symbols correspond to the different levels of RNA-OUT expressed by the PLtet-O1 promoter. The latter was controlled by varying amounts of aTc added to the growth medium (see legend). The solid lines are the steady-state solution (Equation 2) using the best-fit parameters listed in Table S2.(B) Ratio of GFP expression in is10out+ (ZZS35) and is10out− (ZZS31) strains measured through GFP fluorescence (solid bars) and RT-PCR (striped bars). Expression of RNA-OUT was induced by 10 ng/ml aTc (red), whereas target expression was induced with 0.3 mM IPTG [corresponding to the 4th diamond from the right in (A)]. Change in the mRNA level of cris10-gfp (striped bars) is insignificant compared with changes in GFP fluorescence (solid bars).
Mentions: As a controlled source of RNA-OUT, we used the pZA31O plasmid, which harbors the is10out gene driven by the strong synthetic PLtet-O1 promoter [62]. We measured the response function at four different expression levels of RNA-OUT using different concentrations of the inducer aTc (0, 2, 6, and 10 ng/ml). The data obtained (symbols in Figure 3A) were fitted to the steady-state solution (Equation 2) as described above; best-fit parameters are given in Table S2. The fitted curves are presented as the solid lines in Figure 3A. In the absence of aTc, cris10-gfp expression coincides with that of the corresponding strain with no RNA-OUT source (dashed black line). At higher levels of aTc, the threshold-linear response is recovered, displaying a smooth transition as expected.

Bottom Line: Our results demonstrate that sRNA provides a novel mode of gene regulation, with characteristics distinct from those of protein-mediated gene regulation.Knowledge of these special features of sRNA-mediated regulation may be crucial toward understanding the subtle functions that sRNAs can play in coordinating various stress-relief pathways.Our results may also help guide the design of synthetic genetic circuits that have properties difficult to attain with protein regulators alone.

View Article: PubMed Central - PubMed

Affiliation: Center for Theoretical Biological Physics, University of California San Diego, La Jolla, California, United States of America.

ABSTRACT
An increasing number of small RNAs (sRNAs) have been shown to regulate critical pathways in prokaryotes and eukaryotes. In bacteria, regulation by trans-encoded sRNAs is predominantly found in the coordination of intricate stress responses. The mechanisms by which sRNAs modulate expression of its targets are diverse. In common to most is the possibility that interference with the translation of mRNA targets may also alter the abundance of functional sRNAs. Aiming to understand the unique role played by sRNAs in gene regulation, we studied examples from two distinct classes of bacterial sRNAs in Escherichia coli using a quantitative approach combining experiment and theory. Our results demonstrate that sRNA provides a novel mode of gene regulation, with characteristics distinct from those of protein-mediated gene regulation. These include a threshold-linear response with a tunable threshold, a robust noise resistance characteristic, and a built-in capability for hierarchical cross-talk. Knowledge of these special features of sRNA-mediated regulation may be crucial toward understanding the subtle functions that sRNAs can play in coordinating various stress-relief pathways. Our results may also help guide the design of synthetic genetic circuits that have properties difficult to attain with protein regulators alone.

Show MeSH
Related in: MedlinePlus