Sigma E controls biogenesis of the antisense RNA MicA

Nucleic Acids Research, Feb 2007

Adaptation stress responses in the Gram-negative bacterium Escherichia coli and its relatives involve a growing list of small regulatory RNAs (sRNAs). Previous work by us and others showed that the antisense RNA MicA downregulates the synthesis of the outer membrane protein OmpA upon entry into stationary phase. This regulation is Hfq-dependent and occurs by MicA-dependent translational inhibition which facilitates mRNA decay. In this article, we investigate the transcriptional regulation of the micA gene. Induction of MicA is dependent on the alarmone ppGpp, suggestive of alternative σ factor involvement, yet MicA accumulates in the absence of the general stress/stationary phase σS. We identified stress conditions that induce high MicA levels even during exponential growth—a phase in which MicA levels are low (ethanol, hyperosmolarity and heat shock). Such treatments are sensed as envelope stress, upon which the extracytoplasmic sigma factor σE is activated. The strict dependence of micA transcription on σE is supported by three observations. Induced overexpression of σE increases micA transcription, an ΔrpoE mutant displays undetectable MicA levels and the micA promoter has the consensus σE signature. Thus, MicA is part of the σE regulon and downregulates its target gene, ompA, probably to alleviate membrane stress.

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Sigma E controls biogenesis of the antisense RNA MicA

Klas I. Udekwu 0 E. Gerhart H. Wagner 0 0 Department of Cell & Molecular Biology, Uppsala university, Biomedical Center , Box 596, S-75124 Uppsala, Sweden Adaptation stress responses in the Gram-negative bacterium Escherichia coli and its relatives involve a growing list of small regulatory RNAs (sRNAs). Previous work by us and others showed that the antisense RNA MicA downregulates the synthesis of the outer membrane protein OmpA upon entry into stationary phase. This regulation is Hfq-dependent and occurs by MicA-dependent translational inhibition which facilitates mRNA decay. In this article, we investigate the transcriptional regulation of the micA gene. Induction of MicA is dependent on the alarmone ppGpp, suggestive of alternative p factor involvement, yet MicA accumulates in the absence of the general stress/stationary phase pS. We identified stress conditions that induce high MicA levels even during exponential growtha phase in which MicA levels are low (ethanol, hyperosmolarity and heat shock). Such treatments are sensed as envelope stress, upon which the extracytoplasmic sigma factor pE is activated. The strict dependence of micA transcription on pE is supported by three observations. Induced overexpression of pE increases micA transcription, an "rpoE mutant displays undetectable MicA levels and the micA promoter has the consensus pE signature. Thus, MicA is part of the pE regulon and downregulates its target gene, ompA, probably to alleviate membrane stress. - Bacteria meet adverse environmental conditions by rapid adaptive changes in gene expression patterns, thereby mounting appropriate responses. This most often involves transcriptional regulation, by repressors or activators, often under the control of two-component regulatory systems, and/or by alternative sigma (s) factors. In addition, a second level of posttranscriptional control is frequently involved. In Escherichia coli, a growing number of small non-coding RNAs (sRNAs) have been implicated in the regulation of stress responses and virulence traits (13). Many of these are conserved in enteric relatives, and quite a few sRNAs have since been identified in other bacteria as well. Most of the sRNAs are antisense RNAs that inhibit (or, less frequently, activate) the translation of target mRNAs or promote their degradation. The ubiquitous RNA-binding protein Hfq (4) is often required for regulatory activity, though its mechanism of action is not yet fully understood. In contrast to the cis-encoded antisense RNAs in plasmids, most chromosomally encoded sRNAs are trans-encoded (5). Their genes do not overlap target genes, and thus complementarity to the target mRNA is limited and often non-contiguous. Therefore, the location of an sRNA gene does not automatically identify the target gene. Assigning functions for new sRNAs has been based on screening for downstream effects in strains lacking the sRNA, or overexpressing it. Microarray analyses, two-dimensional protein gels, phenotypic tests and bioinformatics-aided complementarity searches have resulted in the identification of targets (6). Conceptually, sRNAs are expected to be under appropriate transcriptional control, so that their induction matches requirements for their regulatory activity. This appears to be borne out by observations: for instance, RyhB is under control of the Fur repressor. When iron concentration is low, Fur repression of the ryhB promoter is abolished, and synthesized RyhB inhibits the synthesis of several iron-binding proteins (7). A similar logic guides the expression characteristics of several other sRNAs. Many sRNAs are upregulated immediately before or upon entry into stationary phase (8,9). This stress response in E. coli has been studied extensively and is characterized by major physiological changes arising from orchestrated alterations in gene expression (10), the majority of which are dependent on the stress/stationary phase sS. This transcription factor, in turn, is under intricate control involving several environmental signals that converge on the expression of its gene, the translation or stability of its mRNA and the activity/stability of the protein itself (11). Among these regulators are two antisense sRNAs, DsrA and RprA, that are induced by different signals and enhance translation of the rpoS mRNA (12). Transcriptome and proteome analyses have been used to chart the sS regulon, which includes transcriptional regulators important for the scavenging of nutrients, such as Crp and the two-component Ntr and Pho systems, and many genes whose products help in adaptation to stationary phase. Recently, microarray data have contributed to the understanding of the transcriptomes of other s factors, such as the heatshock sH (13) and the extracytoplasmic stress sE (14). Interestingly, sH responds not only to heat stress, but is itself transcriptionally upregulated in a sE-dependent manner (15), perhaps indicative of some regulatory hierarchy. The extra (...truncated)


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Klas I. Udekwu, E. Gerhart H. Wagner. Sigma E controls biogenesis of the antisense RNA MicA, Nucleic Acids Research, 2007, pp. 1279-1288, 35/4, DOI: 10.1093/nar/gkl1154