CifR is a 196-residue TetR-family transcriptional repressor encoded divergently from the morB-cfrX-cif operon. As a homodimer it binds two operators in the cifR-morB intergenic region, which overlap the start sites of both divergent transcripts, and so represses the cif operon, encoding the secreted epoxide hydrolase virulence factor Cif, as well as its own gene. CifR is an epoxide sensor: the xenobiotic epoxide epibromohydrin releases it from the operator in vitro and derepresses cif in vivo, and the conserved ligand-binding-domain cysteine Cys107, seen oxidised to sulfenic and sulfinic acid in the crystal structures, is required for epoxide-triggered DNA release. Crystal structures of free CifR and of the CifR-operator complex show one dimer bound per operator, with the N-terminal extension reading the DNA minor groove.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000976 transcription cis-regulatory region binding | IMP PMID:22843844 Epoxide-mediated CifR repression of cif gene expression util... | ACCEPT | Summary: CifR binds two mapped 27-bp operators in the cifR-morB intergenic region that overlap the -10 and +1 elements of both promoters; six-base changes abolish binding in vitro and raise transcription in vivo. Reason: Operator binding was mapped by EMSA with purified protein and confirmed on the chromosome, and the 2024 structures show a CifR dimer on the operator duplex. This is the specific DNA-binding claim for the gene. Supporting Evidence: PMID:22843844 We identified two sites of CifR binding in the intergenic space between cifR and morB, the first gene in the operon containing the cif gene. PMID:22843844 Mutagenesis of these binding sites resulted in a loss of DNA binding in vitro, and mutation of one of these sites in vivo resulted in an increase in transcription of both the cif and cifR genes. PMID:39413156 Despite the length of the operator sequence, the overall structure turned out to involve a single CifR dimer binding per operator site, with unusual protein:DNA contacts at the periphery. |
| GO:0001217 DNA-binding transcription repressor activity | IMP PMID:22843844 Epoxide-mediated CifR repression of cif gene expression util... | ACCEPT | Summary: Core molecular function: operator-bound CifR represses the divergent cif and cifR transcripts, and loss of the operator or of CifR raises their expression. Reason: Both the repressor activity and the DNA-binding mode are directly established; the activity is relieved by epoxide binding, which is the regulatory switch of the circuit. Supporting Evidence: PMID:18458065 We show that the predicted TetR family transcriptional repressor encoded by the PA2931 gene, which is immediately adjacent to and divergently transcribed from the cif-containing, three-gene operon, negatively regulates cif gene expression by binding to the promoter region immediately upstream of the cif-containing operon. PMID:22843844 Mutagenesis of the operator proximal to morB resulted in an increase in expression of both the cif and cifR transcripts, suggesting that both sites are required for complete repression of each of these genes. |
| GO:0003677 DNA binding | IEA GO_REF:0000120 | MODIFY | Summary: Correct but general; the DNA bound is a mapped cis-regulatory operator. Reason: The HTH tetR-type signature justifies DNA binding, and the specific descendant GO:0000976 transcription cis-regulatory region binding is already carried by this gene with experimental evidence. Proposed replacements: transcription cis-regulatory region binding Supporting Evidence: file:PSEAE/cifR/cifR-uniprot.txt DR InterPro; IPR001647; HTH_TetR. PMID:22843844 We identified two sites of CifR binding in the intergenic space between cifR and morB, the first gene in the operon containing the cif gene. |
| GO:0006355 regulation of DNA-templated transcription | IEA GO_REF:0000117 | MODIFY | Summary: True but unsigned; CifR is a repressor. Reason: GO:0045892 negative regulation of DNA-templated transcription is a descendant of GO:0006355 and is already on this gene with EXP, IMP and IEP evidence. Proposed replacements: negative regulation of DNA-templated transcription Supporting Evidence: PMID:18458065 Given its role as a repressor of cif gene expression, we have renamed PA2931 as CifR. |
| GO:0032993 protein-DNA complex | IMP PMID:22843844 Epoxide-mediated CifR repression of cif gene expression util... | ACCEPT | Summary: CifR forms a specific protein-DNA complex with its operators, shown by EMSA and by co-crystal structures of the CifR dimer bound to operator DNA. Reason: Operator occupancy is the state in which CifR acts, and the complex has been visualised crystallographically, so the component claim is directly supported (the 2012 evidence is gel-shift plus chromosomal mutation, for which CollecTF's IMP code is conservative). Supporting Evidence: PMID:39413156 We present here the first molecular structures of CifR alone and in complex with operator DNA, resolved in a single crystal lattice. PMID:22843844 CifR binding to the intergenic region was previously demonstrated by EMSA using a purified hexahistidine-tagged CifR protein ( 25 ). |
| GO:0045892 negative regulation of DNA-templated transcription | EXP PMID:22843844 Epoxide-mediated CifR repression of cif gene expression util... | ACCEPT | Summary: Core process: repression of the cif operon and of cifR itself from two intergenic operators. Reason: Directly shown by operator mapping, loss-of-binding mutants and in vivo transcript measurements. Supporting Evidence: PMID:22843844 Mutagenesis of these binding sites resulted in a loss of DNA binding in vitro, and mutation of one of these sites in vivo resulted in an increase in transcription of both the cif and cifR genes. |
| GO:0045892 negative regulation of DNA-templated transcription | IEP PMID:22843844 Epoxide-mediated CifR repression of cif gene expression util... | ACCEPT | Summary: Same claim from the expression-pattern side: cif and cifR transcript levels respond to operator mutation and to epoxide exposure. Reason: Reinforces the EXP and IMP rows for the same term; no inconsistency. Supporting Evidence: PMID:22843844 Interestingly, the cifR deletion mutant also exhibited a small, but significant 2-fold increase in cif gene expression in the presence of EBH, indicating additional, non-CifR-dependent, epoxide-mediated regulation of cif gene transcription. |
| GO:0045892 negative regulation of DNA-templated transcription | IMP PMID:18458065 Cif is negatively regulated by the TetR family repressor Cif... | ACCEPT | Summary: The PseudoCAP row, and the sole seed of the IBD at PTN002934616. It records the gene's own activity: PA2931 binds the cif promoter region and represses cif, and the complex is released by epibromohydrin. Reason: Not a downstream phenotype. The 2008 paper combines promoter binding by purified protein with cif expression in a cifR background, in P. aeruginosa, and names the gene cifR for that role. The seed is therefore sound and the IBD at PTN002934616 is justified. Supporting Evidence: PMID:18458065 Furthermore, this protein-DNA interaction is disrupted by the epoxide EBH in vitro, suggesting that the binding of EBH by the PA2931 protein product drives the disassociation from its DNA-binding site. PMID:18458065 Given its role as a repressor of cif gene expression, we have renamed PA2931 as CifR. |
| GO:0045892 negative regulation of DNA-templated transcription | IBA GO_REF:0000033 | ACCEPT | Summary: The IBD at PTN002934616 (PTHR47506) is seeded only by cifR's own experimental annotations, which is legitimate: the node term is a broad, sign-correct claim for a clade of TetR/MarR-type HTH repressors. Reason: cifR appearing in its own With/From is expected, not circular: its experimental rows (EXP, IEP, IMP) are the descendant evidence the PAINT curator used, and the IBA adds the judgment that repression is the inherited mode in this clade. A single seed is not weak support when the seed is a well-characterised repressor with structures of the protein-operator complex. The gene-specific claims (the cif operators, epoxide sensing by Cys107) were not propagated, which is correct. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN002934616 · PTN002934616 (PTHR47506) SUPPORTS TRANSFER QuickGO lists 25 GO:0045892 leaf IBAs in 9 taxa on 2026-10-28 (E. coli yjdC, Shewanella, Bradyrhizobium, Streptomyces, Vibrio, Xanthomonas, and P. aeruginosa cifR and PA2484), all TetR/MarR-type HTH regulators, for which the unsigned-to-negative generalisation is appropriate. UniProtKB:Q9HZR6 · cifR (self; sole IBD seed) SUPPORTS TRANSFER The target's own experimental annotations (CollecTF EXP/IEP PMID:22843844, PseudoCAP IMP PMID:18458065) are the descendant evidence for the node; this is the expected pattern, not circularity. Supporting Evidence: PMID:22843844 This virulence factor, named CFTR inhibitory factor (Cif), is regulated by a TetR-family, epoxide-responsive repressor known as CifR via direct binding and repression. |
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Download this section (compressed HTML)Q: What is the physiological epoxide that CifR senses in the airway, and is Cys107 modified covalently by it or only by oxidants such as N-ethylmaleimide in vitro?
Q: Should GO have a term for epoxide-responsive (ligand-released) repressor activity, or is the sensing better captured as an epoxide-binding molecular function on CifR once the physiological ligand is identified?
Experiment: Determine whether CifR binds epoxides directly and whether Cys107 is covalently modified in vivo: mass spectrometry of CifR from epoxide-exposed P. aeruginosa, with Cys107 substitution mutants assayed for operator occupancy (ChIP) and for cif derepression.
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