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Regulation of Expression at Multiple Steps01:23

Regulation of Expression at Multiple Steps

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The gene expression in cells is regulated at different stages: (i) transcription, (ii) RNA processing, (iii) RNA localization, and (iv) translation. Transcriptional regulation is mediated by regulatory proteins such as transcription factors, activators, or repressors—these control gene expression by initiating or inhibiting the transcription of genes. Once a precursor or pre-mRNA is produced, it undergoes post-transcriptional modification, including 5' capping, splicing, and the...
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Translational regulation in prokaryotes ensures efficient protein synthesis by controlling ribosome access to mRNA. This regulation is mediated by secondary RNA structures, including translational riboswitches, RNA thermometers, and small RNAs (sRNAs), which respond to intracellular and environmental signals to modulate gene expression.Translational RiboswitchesRiboswitches in the leader region of mRNAs can regulate translation by altering the accessibility of the Shine-Dalgarno (SD) sequence,...
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Transcriptional Regulation: Riboswitches01:23

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Riboswitches are RNA elements that regulate gene expression by altering their secondary structures in response to specific effector molecules. These elements, located in the leader regions of certain mRNAs, act as transcriptional regulators by toggling between alternative conformations to control downstream gene expression. Riboswitch-mediated regulation is a precise mechanism for modulating biosynthetic pathways, as exemplified by the riboflavin biosynthesis pathway in Bacillus...
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Regulation of Expression Occurs at Multiple Steps02:24

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Gene expression can be regulated at almost every step from gene to protein. Transcription is the step that is most commonly regulated. This involves the binding of proteins to short regulatory sequences on the DNA. This association can either promote or inhibit the transcription of a gene associated with the respective sequence.
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Retargeting a Dual-Acting sRNA for Multiple mRNA Transcript Regulation.

Ashwin Lahiry1, Samuel D Stimple2, David W Wood2,1

  • 1Department of Microbiology, The Ohio State University , 484 W. 12th Avenue, Columbus, Ohio 43210, United States.

ACS Synthetic Biology
|January 10, 2017
PubMed
Summary

We engineered a system in E. coli to create dual-acting small regulatory RNAs (sRNAs) for precise control of gene expression. This platform enables multitargeting sRNAs for advanced metabolic engineering applications.

Keywords:
biofuelsgene regulationmetabolic engineeringnoncoding sRNAsynthetic biology

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Area of Science:

  • Synthetic biology
  • Molecular biology
  • Metabolic engineering

Background:

  • Small regulatory RNAs (sRNAs) naturally control bacterial gene expression.
  • Multitargeting sRNAs offer potential for precise metabolic engineering.
  • The native DsrA sRNA from E. coli has a well-characterized structure amenable to modification.

Purpose of the Study:

  • To develop a system for designing and assaying dual-acting sRNAs.
  • To create retargeted sRNA variants for controlling specific mRNA targets.
  • To enable precise, in vivo quantification of sRNA-mRNA interactions.

Main Methods:

  • Designed an Escherichia coli genetic system for creating and testing dual-acting sRNA variants.
  • Utilized a native DsrA sRNA scaffold with separable functional domains.
  • Employed independently controlled sRNA and reporter mRNA expression with small inducer molecules.
  • Performed microtiter plate assays for in vivo quantification.
  • Semirationally designed and screened DsrA variants targeting Clostridium acetobutylicum n-butanol pathway mRNAs.

Main Results:

  • Successfully created and assayed dual-acting retargeted-sRNA variants.
  • Demonstrated coordinate translational regulation of two distinct mRNA leaders.
  • Developed bifunctional sRNA prototypes by combining modified DsrA sequences.
  • Showcased the ability to target specific mRNA sequences from a metabolic pathway.

Conclusions:

  • The developed system provides a versatile platform for designing multitargeting sRNAs.
  • This approach facilitates the retargeting and characterization of sRNAs for metabolic engineering.
  • Enables precise control over multiple genes within a metabolic pathway.