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1.
FIG. 5.

FIG. 5. From: MicC, a Second Small-RNA Regulator of Omp Protein Expression in Escherichia coli.

Effects of compensatory mutations. Luciferase activities (luminescence counts per microgram of protein) for the micC deletion strain carrying the wild-type ompC-luc fusion (SC218) or the mutant ompC-luc fusion (SC216) and transformed with pAlter-Ex2, pAE-micC, or pAE-micCmutant are given. The experiment was repeated three times, and averages and standard deviations are presented.

Shuo Chen, et al. J Bacteriol. 2004 October;186(20):6689-6697.
2.
FIG. 6.

FIG. 6. From: MicC, a Second Small-RNA Regulator of Omp Protein Expression in Escherichia coli.

Toeprinting analysis of 30S ribosomal subunit binding to ompC mRNA. The arrow indicates the toeprint signal at the C residue, and the three small dots indicate the AUG start codon. The DNA sequencing reactions were carried out with the same end-labeled oligonucleotide used in the toeprinting assay.

Shuo Chen, et al. J Bacteriol. 2004 October;186(20):6689-6697.
3.
FIG. 3.

FIG. 3. From: MicC, a Second Small-RNA Regulator of Omp Protein Expression in Escherichia coli.

Proposed formation of MicC-ompC and MicF-ompF duplexes. (A) Base pairing between wild-type ompC mRNA leader and wild-type MicC RNA (pAE-micC) identified by a BLASTN search (http://www.ncbi.nlm.nih.gov/BLAST/) of the E. coli genome. (B) Base pairing between a mutant ompC mRNA leader and a mutant MicC RNA (pAE-micCmutant). The sequence of the PacI restriction site is UUAAUUAA. (C) Base pairing between wild-type ompC mRNA leader and wild-type and mutant oligonucleotides. (D) Base pairing between wild-type ompF mRNA leader and wild-type MicF RNA (24). Ribosome-binding sites (RBS) and start codons for ompC and ompF are underlined, and the mutant sequences are indicated with lowercase letters.

Shuo Chen, et al. J Bacteriol. 2004 October;186(20):6689-6697.
4.
FIG. 7.

FIG. 7. From: MicC, a Second Small-RNA Regulator of Omp Protein Expression in Escherichia coli.

Requirement of Hfq for MicC repression of ompC. (A) Hfq binding to MicC RNA. Cell extracts were prepared from wild-type (BW25113) or hfq-1 mutant (GSO107) cells grown to an OD600 of 0.6 in LB medium at 24°C. Immunoprecipitations were carried out with the wild-type extracts and an Hfq antiserum or a preimmune serum and were compared to total RNAs from 1/10 extract equivalents of the wild-type and hfq-1 mutant RNAs. The levels of MicC and the DsrA positive control were determined by Northern hybridization. (B) Levels of OmpC, OmpF, and OmpA proteins in wild-type and hfq-1 mutant strains without and with pAlter-Ex2 or pAE-micC grown to an OD600 of 0.6 in LB medium at 24°C.

Shuo Chen, et al. J Bacteriol. 2004 October;186(20):6689-6697.
5.
FIG. 2.

FIG. 2. From: MicC, a Second Small-RNA Regulator of Omp Protein Expression in Escherichia coli.

Sequence of MicC RNA. (A) Primer extension analysis of MicC RNA. Reverse transcriptase reactions were carried out as described in Materials and Methods, using total RNA isolated from JM109 cells grown overnight in LB medium at room temperature. The transcription initiation site corresponding to a G is indicated with an arrow. (B) Sequence of micC gene in E. coli K-12. The −10 and −35 promoter sequences are underlined, and bold letters denote the micC coding sequence. The stem-loop of the predicted terminator is indicated by arrows. (C) Chromosomal position of micC. micC is transcribed clockwise on the chromosome on the strand opposite the adjacent ompN and ydbK genes. (D) Alignment of micC homologs by the CLUSTALW program (http://molbio.info.nih.gov/molbio/gcglite/clustalw18.html).

Shuo Chen, et al. J Bacteriol. 2004 October;186(20):6689-6697.
6.
FIG. 4.

FIG. 4. From: MicC, a Second Small-RNA Regulator of Omp Protein Expression in Escherichia coli.

Effects of increased and decreased expression of MicC on ompC expression. (A) Luciferase activities (luminescence counts per microgram of protein) for the wild-type strain (SC201), the wild-type strain carrying the control pAlter-Ex2 vector, the wild-type strain carrying pAE-micC, and the corresponding micC deletion strain (SC204), with each grown for 6 h in LB medium at 37°C. The experiment was repeated three times, and averages and standard deviations are presented. (B) Levels of OmpC, OmpF, and OmpA proteins in the wild-type strain (BW25113), the wild-type strain carrying the control pAlter-Ex2 vector, the wild-type strain carrying pAE-micC, and the corresponding micC deletion strain (SC200), with each grown to an OD600 of 0.6 in LB medium at 24°C or an OD600 of 0.4 in M9-glycerol medium at 37°C and then treated with 0.3 mM NaCl for 20 min. The strains assayed in panel B were the same as the strains assayed in panel A, except they did not carry the ompC-luc fusion.

Shuo Chen, et al. J Bacteriol. 2004 October;186(20):6689-6697.
7.
FIG. 1.

FIG. 1. From: MicC, a Second Small-RNA Regulator of Omp Protein Expression in Escherichia coli.

MicC and MicF RNA levels under various growth conditions. (A) Levels of MicC and MicF RNAs in JM109 cells grown under different growth conditions as follows: stationary phase (overnight; lane 1) or exponential phase (OD600 = 0.4; lane 2) in LB medium at 24°C; stationary phase (lane 3) or exponential phase (lane 4) in LB medium at 37°C; growth in LB medium at 37°C to exponential phase and then a switch to 42°C for 20 min (lane 5), a treatment with 0.5 mM paraquat for 20 min (lane 6), a treatment with 0.2 mM H2O2 for 5 min (lane 7), or a switch to pH 4.5 for 20 min (lane 8); stationary phase (lane 9) or exponential phase (lane 10) in M9-glycerol medium at 37°C; growth in M9-glycerol medium at 37°C to exponential phase and then a treatment with 0.3 mM NaCl (lane 11) or 10% ethanol (lane 12) for 20 min. The bands corresponding to the MicC and MicF RNAs are denoted by arrows. (B) Levels of MicC and MicF RNAs in wild-type and ompR mutant strains grown to exponential phase (OD600 = 0.4) in LB medium or M9-glycerol medium at 37°C. For both panels, 10-μg samples of total RNA were fractionated in 8% polyacrylamide-urea gels and analyzed by Northern hybridization with a labeled oligonucleotide complementary to MicC or a labeled RNA complementary to MicF.

Shuo Chen, et al. J Bacteriol. 2004 October;186(20):6689-6697.

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