The transcriptional repressor PheR regulates uptake and catabolism of phenanthrene in Sphingobium sp. SHPJ-2

Phenanthrene (PHE) is a typical polycyclic aromatic hydrocarbon (PAH) and a persistent pollutant. Aerobic catabolic metabolism of PHE involves the coordinated regulation between substrate uptake and the energy-intensive initial oxidation steps. Although Sphingobium sp. SHPJ-2 can efficiently degrade PHE, the mechanism linking PHE sensing to the activation of its catabolic pathway remains unclear. Here, we demonstrate that PheR, an IclR-family transcriptional repressor, links PHE availability to transcriptional activation of the phenanthrene-degradation operon in strain SHPJ-2. EMSA and DNase I footprinting analyses indicate that PheR specifically binds to P phnA1 , the promoter upstream of phnA1 that drives transcription of the phenanthrene-degradation operon. Through site-directed mutagenesis combined with biolayer interferometry identifies, we identify three residues (R63, R73, and R78) as critical for DNA binding and define 5′-GCAACG-3′ as the minimal recognition motif for PheR. Importantly, PHE acts as an effector molecule that diminishes PheR binding to the P phnA1 promoter DNA, supporting a model in which PHE availability triggers derepression of the catabolic operon. Consistently, deletion of pheR accelerates PHE degradation and markedly upregulates transcription of phenanthrene-degradation genes. Comparative transcriptomic analysis further indicates that PheR exerts broader downstream effects beyond the core catabolic cluster, including modulation of outer membrane–associated functions such as TonB/ExbBD-dependent energy transduction and envelope homeostasis. Collectively, this work links PHE availability to transcriptional control of phenanthrene catabolism in Sphingobium and identifies PheR as a potentially portable regulatory element for developing PAH-responsive whole-cell biosensors.