In recent years, genomic searches performed in several laboratories have indicated the high prevalence of identified new small, untranslated RNAs in all organisms, from bacteria to animals and plants. In the enterobacterial model E. coli, many new RNAs were identified (50) or predicted (250). However, although their expression patterns suggest involvement in stress responses, few of them have at present a biological function assigned to it. Moreover, even for the few for which information about a biological role has been obtained, detailed mechanistic understanding has not been obtained. Different approaches can be used to address function. Since E. coli is a genetically tractable organism, small RNA knockout or overexpression strains can easily be constructed for subsequent phenotypic analyses. Functional genomics approaches such as phenotype arrays, 2D gels, and microarrays will help to identify the regulated target genes and/ or pathways affected by chosen small RNAs (monitoring changes in mRNAs, or proteins). In addition, a bioinformatics approach developed recently in the host group will be employed to search for target mRNA, on the assumption that most of the small RNAs may act as antisense RNAs. This will be followed up by measuring small RNA effects on target/ reporter gene fusion constructs. Furthermore, small RNA/ target control systems will be characterised in detail by mechanistic studies, primarily carried out in vitro (RNA structure determination, RNA-RNA interaction, effects of helper proteins etc.). In conclusion the proposed work is expected to highlight the variety of ways in which sRNAs can regulate gene expression and the physiological conditions under which they are believed to function, as well as their biochemical properties, and their mode of action. In addition, an elucidation of sRNA-mediated control in bacteria may be of great relevance for comparisons to small RNA (e.g. microRNA) dependent regulation.
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