Sigma E controls biogenesis of the antisense RNA MicA
Klas I. Udekwu
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E. Gerhart H. Wagner
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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.
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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)