RocR (PA3947) is a soluble response regulator made of an N-terminal receiver (REC) domain and a C-terminal EAL domain. The EAL domain is a cyclic-di-GMP-specific phosphodiesterase that hydrolyses c-di-GMP to the linear dinucleotide pGpG, using Mg2+ (or Mn2+) and a general-base mechanism; it does not hydrolyse cAMP or cGMP. RocR forms tetramers, and the phosphorylation state of the receiver domain (Asp56) is thought to tune the active site through loop 6 of the EAL domain. RocR belongs to the RocS1-RocR-RocA1 (SadARS) signalling system, where it receives phosphoryl groups from the sensor kinase RocS1 and antagonises the DNA-binding response regulator RocA1. Through this system and through c-di-GMP turnover it controls cupB/cupC fimbrial gene expression, biofilm maturation, type III secretion and virulence.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005886 plasma membrane | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: PM IBD at PTN002105365, seeded only by membrane-anchored E. coli EAL proteins. RocR is a REC-EAL response regulator with no transmembrane segment and is a tetramer in solution. Reason: The node term does not suit the whole clade, which includes soluble EAL proteins such as RocR and PA2133. There is no evidence that RocR is active at the membrane; a peripheral association with RocS1 cannot be excluded, so the row is marked over-annotated rather than removed. Node-level defect recorded below. Propagation Review Root cause: PROPAGATION BAD Failure modes: COMPARTMENT OR COMPLEX MISMATCH Sources checked: PANTHER:PTN002105365 · PTN002105365 SUPPORTS SOURCE BUT NOT TARGET The membrane seeds (PdeA, PdeB, PdeC, PdeF, PdeG, PdeK, PdeN, CsrD) are TM proteins; RocR is a soluble response regulator. Supporting Evidence: PMID:22753070 a finding consistent with the observation that RocR forms tetramers in solution |
| GO:0071111 cyclic-guanylate-specific phosphodiesterase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from the PTN002105365 IBD (18 seeds). RocR appears among its own sources, which reflects its purified-enzyme evidence and is expected. RocR keeps the catalytic EAL residues and is an active PDE. Reason: Correct for RocR, with direct biochemical support. The IBD stands on many independent seeds. The node-level problem (degenerate EAL leaves such as CsrD, YdiV and BluF receive this IBA without NOT) does not affect RocR and is recorded as a GO_CENTRAL defect. Supporting Evidence: PMID:18344366 catalyzes the hydrolysis of cyclic di-GMP by using a general base-catalyzed mechanism with the assistance of Mg(2+) ion |
| GO:0000160 phosphorelay signal transduction system | IEA GO_REF:0000002 | ACCEPT | Summary: Receiver-domain mapping. RocR is the receiver of the RocS1 phosphorelay, and the D56 phosphorylation site modulates its PDE activity. Reason: RocR is a genuine response regulator of a two-component phosphorelay, and its receiver domain regulates the PDE output. Supporting Evidence: PMID:19376848 the catalytic activity of RocR was affected by mutating the putative phosphorylation site (D56N) in the phosphoreceiver domain |
| GO:0071111 cyclic-guanylate-specific phosphodiesterase activity | IEA GO_REF:0000003 | ACCEPT | Summary: EC 3.1.4.52 mapping; the EC number is experimentally supported for RocR. Reason: Correct core activity. Supporting Evidence: PMID:18344366 catalyzes the hydrolysis of cyclic di-GMP by using a general base-catalyzed mechanism with the assistance of Mg(2+) ion |
| GO:0071111 cyclic-guanylate-specific phosphodiesterase activity | EXP PMID:18344366 Catalytic mechanism of cyclic di-GMP-specific phosphodiester... | ACCEPT | Summary: Purified RocR kinetics and mechanism of c-di-GMP hydrolysis. Reason: Direct purified-enzyme evidence for the core activity. Supporting Evidence: PMID:18344366 catalyzes the hydrolysis of cyclic di-GMP by using a general base-catalyzed mechanism with the assistance of Mg(2+) ion |
| GO:0071111 cyclic-guanylate-specific phosphodiesterase activity | EXP PMID:19376848 The functional role of a conserved loop in EAL domain-based ... | ACCEPT | Summary: Mutational analysis of loop 6 and D56 with purified RocR PDE assays. Reason: Direct purified-enzyme evidence for the core activity. Supporting Evidence: PMID:19376848 the catalytic activity of RocR was affected by mutating the putative phosphorylation site (D56N) in the phosphoreceiver domain |
| GO:0045892 negative regulation of DNA-templated transcription | IMP PMID:15659157 A novel two-component system controls the expression of Pseu... | KEEP AS NON CORE | Summary: PseudoCAP IMP. RocR represses cupB/cupC expression by antagonising RocA1, the DNA-binding activator; RocR does not bind DNA itself. Reason: True as a regulatory outcome, but indirect: RocR acts upstream of the transcription factor. Full text not read, so the row is kept as non-core rather than challenged. Not a PAINT seed. Supporting Evidence: PMID:15659157 RocA1 is a DNA binding response regulator that activates cup genes, and RocR is an antagonist of RocA1 activity |
| GO:0071111 cyclic-guanylate-specific phosphodiesterase activity | IDA PMID:16477007 Analysis of Pseudomonas aeruginosa diguanylate cyclases and ... | ACCEPT | Summary: PseudoCAP IDA and one of the three RocR supports for the PTN002105365 seeding. Lysates of a RocR-overexpressing strain degraded 100% of added synthetic c-di-GMP (Table 1, full text read on PMC). Reason: Seed verdict SOUND. A lysate assay on an overexpressing strain is direct if crude evidence, and purified-enzyme work confirms it. Supporting Evidence: file:PSEAE/rocR/rocR-notes.md PA3825 EVL +(27) PA3947 EVL +(100) PMID:18344366 catalyzes the hydrolysis of cyclic di-GMP by using a general base-catalyzed mechanism with the assistance of Mg(2+) ion |
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Download this section (compressed HTML)Q: Does RocS1-mediated phosphorylation of RocR Asp56 activate or inhibit its PDE activity in vivo, and is the antagonism of RocA1 a sequestration of phosphoryl groups or a c-di-GMP-dependent effect?
Experiment: Compare cup gene expression in a rocR mutant complemented with catalytically dead (EAL-motif alanine substitution) RocR versus a D56N variant, to separate the PDE output from phosphorelay competition.
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