BioProject

MicF is a textbook example of a small regulatory RNA (sRNA) that acts on a trans-encoded target mRNA through imperfect base paring. More...

MicF is a textbook example of a small regulatory RNA (sRNA) that acts on a trans-encoded target mRNA through imperfect base paring. The discovery of MicF as a post-transcriptional repressor of the major Escherichia coli porin OmpF established the paradigm for a meanwhile common mechanism of translational inhibition, through antisense sequestration of a ribosome binding site. However, whether MicF regulates additional genes has remained unknown for almost three decades. Here, we have harnessed the novel superfolder variant of GFP for reporter-gene fusions to validate newly predicted targets of MicF in Salmonella. We show that the conserved 5’ end of MicF acts by seed pairing to repress the mRNAs of global transcriptional regulator Lrp, periplasmic protein YahO, and lipid A-modifying enzyme LpxR. Whilst MicF binds lrp and yahO in the 5’ UTR, it targets lpxR at both the ribosome binding site and deep within the coding sequence. Repression in the coding sequence of lpxR may be achieved by decreasing mRNA stability through exacerbating the use of a native RNase E site proximal to the short MicF-lpxR duplex. Altogether, this study assigns the classic MicF sRNA to the growing class of Hfqassociated regulators that use diverse mechanisms to impact multiple loci. Overall design: To determine the targets of the small regulatory RNA MicF in S. Typhimurium, we looked at the effect of a short pulse of MicF over-expression on the Salmonella transcriptome. To achieve over-expression, the micF gene was cloned in the pBAD plasmid and induced with 0.2% L-arabinose for 10 min. We then extracted the total RNA for transcriptional profiling. A strain carrying the pBAD plasmid w/o insert was used as negative control (also induced by L-arabinose). 3 biological replicates were performed. This sRNA target identification strategy has been described in Papenfort et al; Molecular Microbiology (2006) 62(6), 1674?1688. Less...