AmpR (PA4109) is the LysR-type transcriptional regulator (LTTR) of the chromosomal AmpC beta-lactamase of Pseudomonas aeruginosa PAO1, encoded divergently from ampC. It binds the ampC promoter through its N-terminal helix-turn-helix domain (Ser38 and Lys42 are needed for binding) and activates ampC transcription in response to beta-lactams; its effector-binding domain recognises cell-wall-derived muropeptides, linking peptidoglycan recycling to beta-lactamase induction. AmpR dimerises, may be membrane associated, and does not autoregulate. Beyond ampC it is a global regulator: loss of ampR alters expression of over 500 genes, including the PoxB beta-lactamase, the MexEF-OprN efflux pump, quorum-sensing regulators and QS-controlled virulence factors (pyocyanin, LasA and LasB proteases), type III and VI secretion genes and biofilm formation. Many of these effects are probably indirect, through other regulators.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003700 DNA-binding transcription factor activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA DNA-binding TF activity from the LysR node PTN000794521. Reason: AmpR binds the ampC promoter in vitro and in vivo and activates its transcription; the LysR clade seeds (GcvA, TtdR, DmlR) are characterised TFs. Supporting Evidence: PMID:25182487 Gel shifts using membrane fractions showed that AmpR binds to PampC in vitro whereas in vivo binding was demonstrated using chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR). |
| GO:0003700 DNA-binding transcription factor activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO DNA-binding TF activity (LysR HTH). Reason: Consistent with the direct evidence. Supporting Evidence: PMID:25182487 a potential membrane-associated AmpR dimer regulates ampC expression by direct binding |
| GO:0003700 DNA-binding transcription factor activity | IDA PMID:25182487 Structural and functional characterization of Pseudomonas ae... | ACCEPT | Summary: UniProt IDA DNA-binding TF activity; gel shift and ChIP-qPCR at PampC plus HTH mutagenesis. Reason: Direct binding plus loss of function of HTH mutants. Supporting Evidence: PMID:25182487 Gel shifts using membrane fractions showed that AmpR binds to PampC in vitro whereas in vivo binding was demonstrated using chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR). |
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular-location mapping (Cytoplasm, ECO:0000305). Reason: A transcription factor that binds chromosomal DNA acts in the bacterial cytoplasm (nucleoid); the membrane-association data do not exclude this. Supporting Evidence: PMID:25182487 whereas in vivo binding was demonstrated using chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) |
| GO:0006351 DNA-templated transcription | IDA PMID:25182487 Structural and functional characterization of Pseudomonas ae... | MODIFY | Summary: UniProt IDA DNA-templated transcription. Reason: AmpR regulates transcription of ampC by binding its promoter; it is not part of the transcription machinery. The process is regulation of DNA-templated transcription, which is not an is_a descendant of GO:0006351, so the row is modified. Proposed replacements: regulation of DNA-templated transcription Supporting Evidence: PMID:25182487 a potential membrane-associated AmpR dimer regulates ampC expression by direct binding |
| GO:0006355 regulation of DNA-templated transcription | IBA GO_REF:0000033 | ACCEPT | Summary: IBA regulation of transcription from PTN000794521, seeded by four E. coli LysR regulators and two P. aeruginosa ones (AmpR, TrpI). Reason: Sound; AmpR's own seed is a PseudoCAP row and appears in WITH/FROM as expected. AmpR both activates (ampC) and represses (poxB, mexEF-oprN) genes, so the undirected term is the right level. Supporting Evidence: PMID:25182487 a potential membrane-associated AmpR dimer regulates ampC expression by direct binding |
| GO:0006355 regulation of DNA-templated transcription | IEA GO_REF:0000120 | ACCEPT | Summary: Combined IEA regulation of transcription (LysR InterPro entries). Reason: Consistent with the direct evidence. Supporting Evidence: PMID:25182487 a potential membrane-associated AmpR dimer regulates ampC expression by direct binding |
| GO:0006355 regulation of DNA-templated transcription | IDA PMID:22479525 The regulatory repertoire of Pseudomonas aeruginosa AmpC Γ-l... | ACCEPT | Summary: PseudoCAP IDA regulation of transcription from the PAO1 versus deletion-mutant transcriptome; a seed of the PTN000794521 IBD. Reason: Full text read. The term is correct (more than 500 genes change expression in the ampR deletion, confirmed by qPCR), and AmpR is an established DNA-binding regulator (PMID:25182487). The evidence, however, is a mutant transcriptome and qPCR; the paper reports only bioinformatic prediction of AmpR binding sites and no direct assay, so the code should be IMP rather than IDA. Supporting Evidence: PMID:22479525 Using DNA microarrays, we compared the expression profiles of PAO1 and PAOΞampR, without (uninduced) and with (induced) sub-MIC Ξ²-lactam stress to identify genes that are regulated under the different conditions PMID:22479525 Studies aimed at identifying the direct targets of AmpR are needed to identify genes that are directly regulated by AmpR |
| GO:0016020 membrane | IEA GO_REF:0000044 | KEEP AS NON CORE | Summary: UniProt subcellular-location mapping (Membrane, ECO:0000303 from PMID:25182487). Reason: The source is the authors' suggestion from alkaline phosphatase fusion and shaving experiments. AmpR has no predicted transmembrane segment; peripheral membrane association is possible but not established, and is not where its DNA-binding function is shown. Supporting Evidence: PMID:25182487 Alkaline phosphatase fusion and shaving experiments suggest that AmpR is likely to be membrane associated. |
| GO:0016020 membrane | NAS PMID:25182487 Structural and functional characterization of Pseudomonas ae... | KEEP AS NON CORE | Summary: UniProt NAS membrane from the AmpR characterisation paper. Reason: As for the IEA row; the paper itself frames this as a suggestion. Supporting Evidence: PMID:25182487 Alkaline phosphatase fusion and shaving experiments suggest that AmpR is likely to be membrane associated. |
| GO:0045862 positive regulation of proteolysis | IMP PMID:22479525 The regulatory repertoire of Pseudomonas aeruginosa AmpC Γ-l... | MARK AS OVER ANNOTATED | Summary: PseudoCAP IMP positive regulation of proteolysis; lower LasA and LasB activity in the ampR mutant. Reason: Full text read. The phenotype is lower production of secreted QS-controlled proteases, a downstream effect of AmpR on gene expression (probably via the QS regulators); AmpR does not regulate a proteolytic process as such. The direction is also inconsistent across papers: PMID:16251297 reported higher LasA and lower LasB in an ampR mutant. Supporting Evidence: PMID:22479525 The LasB elastase production was also severely affected due to the loss of ampR |
| GO:0046677 response to antibiotic | IMP PMID:16251297 Pseudomonas aeruginosa AmpR is a global transcriptional fact... | KEEP AS NON CORE | Summary: UniProt IMP response to antibiotic; AmpR controls beta-lactam induction of AmpC. Reason: True; AmpR is the transducer of beta-lactam induction of ampC. The core function is the transcriptional regulation itself. Supporting Evidence: PMID:16251297 The ampR mutation greatly altered AmpC production. |
| GO:0050714 positive regulation of protein secretion | IMP PMID:22479525 The regulatory repertoire of Pseudomonas aeruginosa AmpC Γ-l... | MARK AS OVER ANNOTATED | Summary: PseudoCAP IMP positive regulation of protein secretion. Reason: Full text read. The data are lower transcript levels of type III secretion genes in the ampR mutant (microarray and qPCR) and lower amounts of secreted factors; secretion itself was not measured. The effect is on gene expression, probably indirect, so a protein-secretion regulation term overstates it. Supporting Evidence: PMID:22479525 Fold changes (FC) as determined by microarray experiments in the absence of Ξ²-lactam stress of the Type VI secretion (β΄, HSI-I) and Type III secretion (βͺ) genes in the ampR mutant strain |
| GO:1900232 negative regulation of single-species biofilm formation on inanimate substrate | IMP PMID:22479525 The regulatory repertoire of Pseudomonas aeruginosa AmpC Γ-l... | KEEP AS NON CORE | Summary: PseudoCAP IMP; the ampR deletion forms more biofilm over 72 h. Reason: Full text read. Direct mutant phenotype in a biofilm assay; a pleiotropic, probably indirect output of the AmpR regulon. Supporting Evidence: PMID:22479525 over a period of 72 hours indicating that AmpR is a negative regulator of biofilm formation |
| GO:1900378 positive regulation of secondary metabolite biosynthetic process | IMP PMID:22479525 The regulatory repertoire of Pseudomonas aeruginosa AmpC Γ-l... | KEEP AS NON CORE | Summary: PseudoCAP IMP; the ampR deletion makes less pyocyanin. Reason: Full text read. Measured pyocyanin phenotype; peripheral, probably via QS regulators. Note that PMID:16251297 reported more pyocyanin in an ampR mutant, so the direction may depend on strain or conditions. Supporting Evidence: PMID:22479525 reduction in pyocyanin production by the ampR mutant, compared to PAO1 |
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Download this section (compressed HTML)Q: Which genes of the >500-gene AmpR regulon are direct targets? ChIP-seq of AmpR with and without beta-lactam would separate direct from indirect (QS- or sigma-factor-mediated) effects.
Experiment: ChIP-seq of epitope-tagged AmpR in PAO1 with and without sub-MIC beta-lactam, combined with EMSA of purified AmpR at selected promoters (mexR, lasR, rhlR, poxB).
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