Curation Information

Publication
Mapping the regulon of Vibrio cholerae ferric uptake regulator expands its known network of gene regulation.;Davies BW, Bogard RW, Mekalanos JJ;Proceedings of the National Academy of Sciences of the United States of America 2011 Jul 26; 108(30):12467-72 [21750152]
TF
Fur [P0C6C8, view regulon]
Reported TF sp.
Vibrio cholerae El Tor biotype strain C6706 (948564)
Reported site sp.
Vibrio cholerae El Tor biotype strain C6706 (948564)
Created by
Erill Lab
Curation notes
ChIP-seq+MEME some sites verified by RT-PCR in independent curation

Experimental Process

ChIP seq, plus peak-calling. Peak fragments fed to MEME for detection of binding motif. Expression data curated from previous source [PMID: 16299312]. Details of ChIP-seq: "We compared the peak lists generated from all three samples and required that a peak be called in at least two of the three experiments to be considered a vcFur-binding site (Table S1). The average ChIP peak length was ∼500 bp."

ChIP assay conditions
VchFur: Vibrio cholerae El Tor biotype strain C6706 and a spontaneous lacZ− derivative of C6706 were used as parental (WT) strains. Fiftymilliliters ofexponentiallygrowing cultureinLB+ 40μM FeSO4 were induced with 0.1% arabinose for 30 min at 37° C.
ChIP notes
Formaldehyde was added to a final concentration of 1% and incubated at RT for 20 min with occasional swirling. Crosslinking was quenched by adding glycine to 0.5M. Cell pellets were washed in 1× TBS and resuspended in lysis buffer [10 mM Tris (pH 8.0), 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% deoxycholate (DOC), 0.5% N-lauroylsarcosine] plus protease inhibitor mixture (Sigma) and 1 mg/mL lysozyme and were incubated at 37° C for 30 min. The cells were sonicated 1×for 30 s with a needle sonicator, and unlysed debris was pelleted by centrifugation. The lysate was sonicated for 20 min with a 10-s on/10-s off cycle (Mixsonix). A sample was taken as a sequencing input control. Following clarification by centrifugation, 1/10 volume of 10% Triton X-100 in lysis buffer was added to each sample followed by 100 μL of Dynal-Protein G beads coated with anti-V5 monoclonal antibody (Sigma), and samples were incubated overnight with rotation. The beads were washed 5× with RIPA buffer [50 mM Hepes (pH7.5), 500 mM LiCl, 1 mM EDTA, 1% Nonidet P-40, 0.7% DOC] and then 1× in Tris-EDTA pH 8.0 plus 50 mM NaCl and were resuspended in 100μL elution buffer [50 mM Tris-HCl (pH 7.5), 10 mM EDTA, 1% SDS]. Samples were incubated at 65° C for 30 min, and the beads were pelleted by centrifugation. Supernatants were incubated at 65° C overnight to reverse crosslinks. Samples were incubated with 8 μL of 10-mg/mL RNase A for 2 h at 37° C and then with 4μL of 20 mg/mL proteinase K at 55° C for 2 h and were purified with Qiagen MinElute Reaction Cleanup Kit and quantitated with Pico green kit (Invitrogen). Experiments were performed in triplicate. One to three nanograms of ChIP or input DNA was processed for sequencing by the addition of a polyA tail as described by Helicos protocols (http://www.helicosbio.com/). Samples were sequenced using the HeliScope Single Molecule Sequencer at the Molecular Biology Core Facility in the Dana-Farber Cancer Institute, Boston, MA. The sequence reads were aligned to th

Transcription Factor Binding Sites


AAGATGATAATGAATCTCAAA
TAAATGAGAACTATTATTATT
CAAATGATAGTAATTTTCATT
GTAATGATAACCTTTATCAAT
TAAATAATAACAATTATCATT
TTAATAAAAACTATTCTCATT
CAAATGATAATAATTTGCAAT
ATAATGAGAGCGTTTCTCAAT
AATGTGATAATAATTATCATT
TTGTCGATAATAATTCTCATT
TTAATAATAATCATTATCAAT
CAAATGAGAATGTATATCATT
CAAATGTAAAGCATTCTCATT
TTAATGATAATTAATATCATT
AATCTGTAAGCCTTTCTCAAT
TAAATGAGAACGATTTTCTTG
TAACTAAGAATTAAATTCTAG
AACTTGACCATCAATCTCTTT
TTCTCTAACATTGTTCTAAAT
CAATTGATAACCAATCTCAAC
TGTTCAAGAACTATTCTAAAC
CGTCAATGAGTTTTTCTCATT
CAACTGAAAATGTTTCTCAAG
TGGGTGAGCAGGGTTCTCAAT
GCGCTAATAACCTTTCTCATC
CGTCTGAGAATAATTCGTATT
AAACGGGTAAATTATCTCTTG
CGAGCGAAAGTCATTCTCATC
TAATTGATAATTATTATCATG
TAACTGATAACGATTCTCATT
ACCGCGAAAAAGATTCTCAAC
ATGTTGAGAGCTTATCCCATT
GCGTAGTTCATCATTATCATT
AAATTGTACCGTATTCTCATC
ATGACAGTCGGTATTATCATG
CAAATGATAACTGCTCTCAAT
AAGATGATAATGAATCTCAAA
TAAATGAGAACTATTATTATT
CAAATGATAGTAATTTTCATT
GTAATGATAACCTTTATCAAT
TAAATAATAACAATTATCATT
TTAATAAAAACTATTCTCATT
CAAATGATAATAATTTGCAAT
ATAATGAGAGCGTTTCTCAAT
AATGTGATAATAATTATCATT
TTGTCGATAATAATTCTCATT
TTAATAATAATCATTATCAAT
CAAATGAGAATGTATATCATT
CAAATGTAAAGCATTCTCATT
TTAATGATAATTAATATCATT
AATCTGTAAGCCTTTCTCAAT
TAAATGAGAACGATTTTCTTG
TAACTAAGAATTAAATTCTAG
AACTTGACCATCAATCTCTTT
TTCTCTAACATTGTTCTAAAT
CAATTGATAACCAATCTCAAC
TGTTCAAGAACTATTCTAAAC
CGTCAATGAGTTTTTCTCATT
CAACTGAAAATGTTTCTCAAG
TGGGTGAGCAGGGTTCTCAAT
GCGCTAATAACCTTTCTCATC
CGTCTGAGAATAATTCGTATT
AAACGGGTAAATTATCTCTTG
CGAGCGAAAGTCATTCTCATC
TAATTGATAATTATTATCATG
TAACTGATAACGATTCTCATT
ACCGCGAAAAAGATTCTCAAC
ATGTTGAGAGCTTATCCCATT
GCGTAGTTCATCATTATCATT
AAATTGTACCGTATTCTCATC
ATGACAGTCGGTATTATCATG
CAAATGATAACTGCTCTCAAT

Gene Regulation

Regulated genes for each binding site are displayed below. Gene regulation diagrams show binding sites, positively-regulated genes, negatively-regulated genes, both positively and negatively regulated genes, genes with unspecified type of regulation. For each indvidual site, experimental techniques used to determine the site are also given.

Site sequence Regulated genes Gene diagram Experimental techniques TF function TF type
AAGATGATAATGAATCTCAAA VC0091,
... ... VC0091 VC0090 VC0092
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TAAATGAGAACTATTATTATT
... ... VC0106 VC0107
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CAAATGATAGTAATTTTCATT VC0284,
... ... VC0284 VC0283 VC0285 VC0286
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
GTAATGATAACCTTTATCAAT VC0608,
... ... VC0608 VC0607 VC0606 VC0609 VC0610
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TAAATAATAACAATTATCATT VC0771,
... ... VC0771 VC0770
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TTAATAAAAACTATTCTCATT VC0773, VC0774,
... ... VC0773 VC0774 VC0772 VC0775
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CAAATGATAATAATTTGCAAT VC1264,
... ... VC1264 ribA VC1265 VC1266 VC1267
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
ATAATGAGAGCGTTTCTCAAT VC1548,
... ... VC1548 VC1547 VC1546 VC1545 VC1544 VC1543 VC1549
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
AATGTGATAATAATTATCATT VC1572,
... ... VC1572 VC1571 VC1570 VC1569 VC1568 VC1567 VC1566 VC1565 VC1564 VC1563 fumC
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TTGTCGATAATAATTCTCATT VC1688,
... ... VC1688 VC1687 VC1686 VC1685 VC1684 VC1683 VC1682 VC1681 VC1680 VC1689
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TTAATAATAATCATTATCAAT vibF,
... ... vibF VC2210
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CAAATGAGAATGTATATCATT VC2210,
... ... VC2210 VC2211
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CAAATGTAAAGCATTCTCATT VC2211,
... ... VC2211 VC2212
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TTAATGATAATTAATATCATT VC2694,
... ... VC2694 cpxA VC2692
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
AATCTGTAAGCCTTTCTCAAT
... ... VC0913 VC0912 tRNA-Tyr-5 VC0914
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TAAATGAGAACGATTTTCTTG
... ... VC1235 VC1236
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TAACTAAGAATTAAATTCTAG
... ... VC2011 VC2012
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
AACTTGACCATCAATCTCTTT
... ... VC2333 VC2332 VC2331 VC2330 VC2334 VC2335 VC2336
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TTCTCTAACATTGTTCTAAAT
... ... VC2080 VC2079 VC2081
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CAATTGATAACCAATCTCAAC
... ... VC0142 VC0143
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TGTTCAAGAACTATTCTAAAC
... ... avtA glyS glyQ
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CGTCAATGAGTTTTTCTCATT
... ... VC0034 VC0033 VC0035 VC0036
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CAACTGAAAATGTTTCTCAAG
... ... VC0239 VC0238 VC0237 VC0236 VC0235 VC0234 VC0233 VC0232 VC0231 VC0230 VC0229 VC0228 VC0227 VC0226 VC0225 VC0224 rfaD
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TGGGTGAGCAGGGTTCTCAAT
... ... pyrG eno VC2449 mazG
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
GCGCTAATAACCTTTCTCATC
... ... rnpA VC0005 VC0004 rpmH
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CGTCTGAGAATAATTCGTATT
... ... VC0073 VC0072 VC0074
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
AAACGGGTAAATTATCTCTTG
... ... VC0882 VC0881 VC0883 VC0884 VC0885 VC0886
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CGAGCGAAAGTCATTCTCATC
... ... VC0931 VC0930 VC0932 VC0933
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TAATTGATAATTATTATCATG
... ... VC1009 VC1008 VC1010 VC1011 VC1012 VC1013 rnfD VC1015 VC1016 VC1017
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
TAACTGATAACGATTCTCATT
... ... rrmA VC2002 VC2004
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
ACCGCGAAAAAGATTCTCAAC
... ... VC2232 fadE VC2233
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
ATGTTGAGAGCTTATCCCATT
... ... VC2489 VC2488 VC2487 VC2490
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
GCGTAGTTCATCATTATCATT
... ... VC2498 tRNA-Leu-10 VC2499 VC2500
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
AAATTGTACCGTATTCTCATC
... ... VC2738 VC2737 VC2739 VC2740
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
ATGACAGTCGGTATTATCATG
... ... VC0052 VC0051 VC0053 VC0054 VC0055
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer
CAAATGATAACTGCTCTCAAT
... ... VC0872 VC0871 VC0873 VC0874 VC0875
Experimental technique details ChIP-Seq (ECO:0006009) - Experimental technique details Motif-discovery (ECO:0005558) - repressor dimer