Characterization of MicA interactions suggests a potential novel means of gene regulation by small non-coding RNAs.
MicA is a small non-coding RNA that regulates ompA mRNA translation in Escherichia coli. MicA has an inhibitory function, base pairing to the translation initiation region of target mRNAs through short sequences of complementarity, blocking their ribosome-binding sites. The MicA structure contains two stem loops, which impede its interaction with target mRNAs, and it is thought that the RNA chaperone protein Hfq, known to be involved in MicA regulation of ompA, may structurally remodel MicA to reveal the ompA-binding site for cognate pairing. To further characterize these interactions, we undertook biochemical and biophysical studies using native MicA and a 'stabilized' version, modified to mimic the conformational state of MicA where the ompA-binding site is exposed. Our data corroborate two proposed roles for Hfq: first, to bring both MicA and ompA into close proximity, and second, to restructure MicA to allow exposure of the ompA-binding site for pairing, thereby demonstrating the RNA chaperone function of Hfq. Additionally, at accumulated MicA levels, we identified a Mg(2+)-dependent self-association that occludes the ompA-recognition region. We discuss the potential contribution of an Mg(2+)-mediated conformational switch of MicA for the regulation of MicA function.
Biophysics Laboratories, School of Biological Sciences, Institute of Biomedical and Biomolecular Sciences, University of Portsmouth, Portsmouth PO1 2DT, UK.
Charlotte A
CA
Henderson
Helen A
HA
Vincent
Carlanne M
CM
Stone
Jack O
JO
Phillips
Peter D
PD
Cary
Darren M
DM
Gowers
Anastasia J
AJ
Callaghan
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000737:chemistry; D015964:Gene Expression Regulation, Bacterial; D035001:Host Factor 1 Protein / Q000737:chemistry; D055029:Inverted Repeat Sequences; D008274:Magnesium / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D058727:RNA, Small Untranslated / Q000737:chemistry / Q000235:genetics*
D001425:Bacterial Outer Membrane Proteins; C405431:DsrA RNA, E coli; D029968:Escherichia coli Proteins; C087373:Hfq protein, E coli; D035001:Host Factor 1 Protein; D058727:RNA, Small Untranslated; C078167:OMPA outer membrane proteins; D008274:Magnesium