Small RNA-mediated Regulation of Gene Expression in Escherichia coli

• Main Author: Unoson, Cecilia
• Format: Doctoral Thesis
• Language: English
• Published: Uppsala universitet, Institutionen för cell- och molekylärbiologi 2010
• Subjects: Small RNA; non-coding RNA; antisense; translational control; ribosome standby; toxin-antitoxin; IstR; TisB; tisAB; MicF; Lrp; Microbiology; Mikrobiologi; Cell and molecular biology; Cell- och molekylärbiologi
• Online Access: http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-130885; http://nbn-resolving.de/urn:isbn:978-91-554-7891-9

Non-coding RNAs are highly abundant regulators of gene expression in all kingdoms of life that often play important roles in vital cellular functions. In bacteria, small regulatory RNAs (sRNAs) usually act post-transcriptionally by regulating mRNAs through base pairing within ribosome binding sites (RBS), thereby inhibiting translation initiation. tisB encodes a toxin, TisB, whose synthesis is controlled by the sRNA IstR-1. Intriguingly, IstR-1 base pairs far upstream of the RBS but nevertheless inhibits translation initiation. The tisB mRNA is unusual in that ribosomes cannot access the RBS directly, but instead need an unstructured upstream region. This is precisely where IstR-1 exerts its inhibitory effect. We propose this region to serve as a ribosome loading site (standby site) which permits ribosomes to overcome the obstacle of inhibitory RBS-containing structures. Sequence-independent ribosome binding to the standby site allows for efficient relocation to the RBS structure when it is transiently open. Thus, standby sites are translation enhancer elements. I also characterized TisB-mediated toxicity. The hydrophobic protein TisB is targeted to the inner membrane and causes damage. This decreases the intracellular ATP concentration and entails decreased  replication, transcription and translation rates. It is likely that this toxin is involved in multidrug tolerance under certain conditions. We identified the sRNA MicF as a negative regulator of lrp expression. Lrp is a global transcription factor that controls genes involved in amino acid metabolism and transport of small molecules. Interestingly, Lrp also downregulates MicF. Thus, this study established that the mutual downregulation of MicF/Lrp creates a positive feedback loop which gives a switch-like behavior important for fast adaptations.