| Aspect (definition/mechanism/phenotype/application/statistic) | Key points | Evidence (what experiment or statement) | Primary source (first author year) | URL | Pub date |
|---|---|---|---|---|---|
| Definition | **rpoN (PP_0952)** in *Pseudomonas putida* KT2440 encodes **RpoN/σ54/σN**, an alternative RNA polymerase sigma factor, matching UniProt P0A171. | KT2440 transcriptome study lists **rpoN (PP_0952)** as “Sigma factor RpoN”; broader σ54 literature defines RpoN as σ54/σN. (pqac-00000022, pqac-00000019) | Mozejko-Ciesielska 2017 | https://doi.org/10.1186/s13568-017-0396-z | 2017-05 |
| Mechanism | RpoN recognizes **−24/−12 promoters** with a conserved **GGN10GC** motif rather than σ70-type −35/−10 promoters. | Review statement describing direct σ54 promoter recognition at −24/−12 and motif architecture. (pqac-00000019) | Yu 2021 | https://doi.org/10.3390/ijms222312692 | 2021-11 |
| Mechanism | RpoN-dependent transcription requires **enhancer-binding proteins (EBPs)** with a central **AAA+ ATPase** domain. | Review and EcoSal article state σ54 forms a closed complex and needs ATP-driven remodeling by AAA+ EBPs to initiate transcription. (pqac-00000019, pqac-00000016, pqac-00000020) | Busby 2024 | https://doi.org/10.1128/ecosalplus.esp-0039-2020 | 2024-12 |
| Mechanism | Domain architecture aligns with UniProt family assignment: **N-terminal activator-interacting domain** and **C-terminal DNA-binding domain**. | Review explicitly describes two conserved functional domains in σ54/RpoN. (pqac-00000004, pqac-00000019) | Yu 2021 | https://doi.org/10.3390/ijms222312692 | 2021-11 |
| Localization/function | RpoN acts in the **cytoplasm/nucleoid-associated transcription machinery** as part of the **RNAP holoenzyme** rather than as a secreted or membrane protein. | Mechanistic reviews describe σ54 as binding core RNAP and promoter DNA to direct transcription initiation. (pqac-00000019, pqac-00000016) | Busby 2024 | https://doi.org/10.1128/ecosalplus.esp-0039-2020 | 2024-12 |
| Phenotype/pathway | Under nitrogen limitation, the **rpoN mutant transcriptome clusters closely with wild type** for PHA-related expression; mcl-PHA accumulation can still occur without RpoN under these conditions. | RNA-seq and RT-qPCR comparison of WT, relA/spoT, and rpoN mutant; authors state analyzed pha transcripts were at comparable levels in WT and rpoN mutant. (pqac-00000011, pqac-00000009) | Dabrowska 2020 | https://doi.org/10.3390/ijms22010152 | 2020-12 |
| Statistic | In nitrogen-limited cultures, **ammonium at 48 h** was **2.1 g/L (WT), 1.7 g/L (relA/spoT), 1.63 g/L (rpoN)**, indicating similar N consumption by the rpoN mutant. | Batch-culture physiology in PHA study. (pqac-00000011) | Dabrowska 2020 | https://doi.org/10.3390/ijms22010152 | 2020-12 |
| Statistic | In the same study, **phaI/phaF operon expression was ~40-fold higher** than **phaC1ZC2D**; however, WT and rpoN mutant showed comparable PHA-gene transcript levels. | RT-qPCR across pha loci under N limitation. (pqac-00000011) | Dabrowska 2020 | https://doi.org/10.3390/ijms22010152 | 2020-12 |
| Pathway/regulon | Nitrogen limitation in the **rpoN mutant** still induced multiple **nitrogen acquisition and transport genes**, including **urtA–urtD** urea transporter genes. | RNA-seq table lists strong upregulation of **urtA–urtD** in WT and rpoN backgrounds. Example: **urtA 12.7-fold (WT), 11.12-fold (rpoN)**. (pqac-00000009) | Dabrowska 2020 | https://doi.org/10.3390/ijms22010152 | 2020-12 |
| Statistic | Nitrogen limitation induced transporter genes associated with a **sigma-54-dependent regulator** region (**PP_2259–PP_2263**), but induction was reduced in the rpoN mutant. | RNA-seq table: **PP_2260** glycerol-phosphate ABC transporter **17.36-fold (WT) vs 6.74-fold (rpoN)**; **PP_2261** sugar ABC transporter **39.87-fold (WT) vs 8.01-fold (rpoN)**. (pqac-00000009) | Dabrowska 2020 | https://doi.org/10.3390/ijms22010152 | 2020-12 |
| Statistic | Several respiratory oxidase genes changed strongly in the rpoN mutant under N limitation. | RNA-seq table reports high fold changes in rpoN column, including **ccoO-I (PP_4251) 252.3**, **ccoQ-I (PP_4252) 209.35**, **ccoP-I (PP_4253) 118.18**, **cyoD (PP_0815) 37.9**, **cyoC (PP_0814) 15.99**. (pqac-00000008) | Dabrowska 2020 | https://doi.org/10.3390/ijms22010152 | 2020-12 |
| Pathway/regulon | The rpoN-linked nitrogen-response dataset also includes **glnL/glnG (NtrB/NtrC system)** and chemotaxis/flagellar genes, consistent with σ54 involvement in broader nutrient-response networks. | RNA-seq tables annotate **glnL**, **glnG**, chemotaxis genes, and **FlgE** among N-limitation-responsive loci. (pqac-00000008, pqac-00000009) | Dabrowska 2020 | https://doi.org/10.3390/ijms22010152 | 2020-12 |
| Phenotype/pathway | In KT2440, **K1-T6SS expression is repressed by RpoN and FleQ**, even though the T6SS promoters themselves are mainly **σ70-dependent**. | Promoter-lacZ assays in regulator mutants; authors define four σ70-like promoters and show increased expression in **rpoN** and **fleQ** mutants. (pqac-00000013, pqac-00000014, pqac-00000015) | Bernal 2023 | https://doi.org/10.1099/mic.0.001295 | 2023-01 |
| Statistic | **Promoter derepression in Bernal 2023:** deletion of **rpoN** caused a **clear qualitative increase** (substantial de-repression; approximately twofold for one tested construct, and stronger for some PS/PV promoter assays) in K1-T6SS promoter activity, especially in exponential phase; exact values are figure-based and not fully readable here. | β-galactosidase assays in WT vs ΔrpoN and ΔfleQ; figure interpretation indicates substantial derepression and one explicitly noted **twofold** effect for the **M1** construct in ΔrpoN. (pqac-00000012, pqac-00000015, pqac-00000023) | Bernal 2023 | https://doi.org/10.1099/mic.0.001295 | 2023-01 |
| Mechanism | For K1-T6SS, RpoN repression appears **indirect**, not due to direct binding of a canonical σ54 site in the PS2 promoter. | Site-directed mutagenesis of the putative RpoN box showed results inconsistent with direct promoter binding; authors conclude RpoN likely represses via another regulator or noncanonical mechanism. (pqac-00000012, pqac-00000014) | Bernal 2023 | https://doi.org/10.1099/mic.0.001295 | 2023-01 |
| Application | Understanding RpoN/FleQ control of **K1-T6SS**, a system used by *P. putida* to outcompete phytopathogens, is relevant to **biological control** and engineering of plant-protective pseudomonads. | Bernal et al. frame K1-T6SS as a relevant antimicrobial/biocontrol system whose expression could be manipulated through this regulatory network. (pqac-00000013) | Bernal 2023 | https://doi.org/10.1099/mic.0.001295 | 2023-01 |
| Application | In the industrial chassis KT2440, RpoN is part of the nitrogen-responsive regulatory context around **mcl-PHA bioplastic production**, but available data suggest **RpoN is not the main limiting regulator** for PHA accumulation on gluconate under N limitation. | Comparative transcriptomics and physiology show mcl-PHA production persists in the rpoN mutant, with WT and rpoN transcriptomes grouping together. (pqac-00000011, pqac-00000009) | Dabrowska 2020 | https://doi.org/10.3390/ijms22010152 | 2020-12 |
| Expert analysis | Current expert consensus is that σ54 systems are unusually specialized: unlike σ70, they often integrate environmental signals through multiple EBPs, making RpoN a **global transcriptional hub** rather than an enzyme with a substrate. | Review-level synthesis emphasizing enhancer dependence, ATP-coupled activation, and broad regulatory integration. (pqac-00000019, pqac-00000016, pqac-00000021) | Busby 2024 | https://doi.org/10.1128/ecosalplus.esp-0039-2020 | 2024-12 |


*Table: This table summarizes verified mechanistic, physiological, and application-relevant findings for *Pseudomonas putida* KT2440 rpoN (RpoN/σ54), with direct evidence and recent authoritative sources. It is useful as a compact evidence map linking sigma-54 mechanism to KT2440-specific phenotypes and quantitative transcriptomic observations.*