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    Udo Bläsi
    Renee Schroeder
    Andrea Barta
    Denise Barlow
    Kristina Djinovic-Carugo
    Michael Jantsch
    Robert Konrat
    Anton Wutz
    Silke Dorner
    Isabella Moll
    Christina Waldsich
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Group: Udo Bläsi

David Hasenöhrl | Postdoc
Birgit Märtens | Postdoc
Armin Resch | Postdoc
Branislav Vecerek | Postdoc
Theresa Sorger-Domenigg | PhD Student
Salim Manoharadas | PhD Student
Alessandra Romeo | PhD Student
Sarah Schlosser | PhD student
Hermann Hämmerle | Diploma Student
 


Post-transcriptional Regulation by small non-coding RNAs and Hfq in Bacteria
(SFB projects 1715 / 1720)

Bacteria are constantly challenged by changing environmental conditions. They employ a number of translational control mechanisms including trans-acting proteins, small non-coding RNAs (sRNAs) as well as features inherent to mRNA structure, which permit a fast adaptation to new environments or to different kinds of stress. E. coli host factor I (Hfq) has a hexameric ring-shaped structure and belongs to the large family of Sm and Sm-like proteins with RNA binding activity. The work on Hfq was initiated by our observation that translation initiation of the E. coli ompA gene is mediated by Hfq. This work directly demonstrated that a 30S ribosome bound to the ompA 5'-UTR protected the transcript from RNase E cleavage, and thereby contributed to a better understanding of the interrelation between translation (initiation) and mRNA turnover. Subsequent work revealed that Hfq induces structural changes in RNA and that it functions as a genuine RNA chaperone. In Gram-negative Bacteria, Hfq acts as a global regulator involved in post-transcriptional regulation and virulence. Hfq binds to a number of sRNAs, contributes to their stability, and is often required for translational silencing and activation mediated by these riboregulators. At present, it is poorly understood how the protein interacts with sRNAs and their mRNA substrates. Therefore, collaborative efforts with the Djinovic-Carugo and Konrat groups concern the elucidation of the complete 3D-structure of Hfq and that of Hfq-RNA complexes. Other sub-projects aim at defining (i) Hfq interacting proteins involved in post-transcriptional processes, (ii) the molecular events underlying sRNA-based mRNA silencing, and (iii) the molecular events leading to decay or stabilization of sRNA/mRNAs complexes. In the context of (ii), we were recently able to demonstrate that a sRNA does not necessarily regulate translation of a target mRNA in a direct manner, i.e. by binding to or in the vicinity of its ribosome binding site but can do so by regulation of an upstream open reading frame to which the downstream gene is translationally coupled. In addition, this work provided evidence for an iron-dependent decoding mechanism of mRNAs encoding iron-containing proteins.
Pseudomonas aeruginosa (PAO1) is a major cause of hospital acquired infections, exerts the highest case fatality rate among Gram-negative pathogens, and contributes to mortality in cystic fibrosis patients. As we have previously shown that a PAO1 hfq- mutant was significantly attenuated in virulence, it can be anticipated that sRNAs are involved in regulation of many virulence genes. The complexity of post-transcriptional regulation mediated by Hfq became apparent when we compared the transcriptome profiles of PAO1 with that of an isogenic hfq- mutant. The complex regulatory circuit affected by Hfq involved stabilization of a regulatory RNA, which indirectly affects a regulatory cascade, mediating virulence gene expression. The P. aeruginosa project is concentrating on the following topics: (i) detection of sRNAs under various stress conditions, including identification of Hfq-binding sRNAs expressed under mimicked infection conditions, and (ii) validation of their potential target genes using biochemical and genetic methods. It is anticipated that these approaches will reveal sRNAs involved in virulence gene expression, i.e. in post-transcriptional regulation of the PAO1 pathogenome. To this end we have obtained evidence that some of the identified and characterized sRNAs impact on multi-drug resistance in PAO1 and on synthesis of virulence factors .


Selected recent publications

Sonnleitner, E., Sorger-Domenigg, T., Madej, M.J., Findeiß, S., Hackermüller, J., Hüttenhofer, A., Stadler, P., Moll, I. and U. Bläsi (2008) Detection of small non-coding RNAs in Pseudomonas aeruginosa by RNomics and by structure-based bioinformatics tools. Microbiology 154: 3175-3187.

Resch, A., Afonyushkin, T., McDowall, K.J., Bläsi, U. and V.R. Kaberdin (2008) Translational activation by the non-coding RNA DsrA involves alternative RNase III processing in the rpoS 5’-leader. RNA 14:454-459.

Večerek, B., Rajkowitsch, L., Sonnleitner, E., Schroeder, R. and U. Bläsi (2008) The C-terminal domain of Escherichia coli Hfq is required for regulation. Nucleic Acids Research 36:133-143.

Večerek B., Moll, I. and Bläsi, U. (2007) Control of Fur synthesis by the non-coding RNA RyhB and iron-responsive decoding. EMBO J. 26:965-975.

Sonnleitner, E., Schuster, M., Sorger-Domenigg, T., Greenberg, E.P., and U. Bläsi (2006) Hfq-dependent alterations of the transcriptome profile and effects on quorum sensing in Pseudomonas aeruginosa. Mol. Microbiol. 59:1542-1558.