Dual modes of Zn2+-dependent and independent virulence regulation by CzcSR in Pseudomonas syringae pv. actinidiae

CzcSR acts as a positive regulator of Psa M228 virulence in host plants and the hypersensitive response in non-host plants. 2. CzcR directly targets the hrpR / S operon promoter to activate T3SS expression in Psa M228. 3. CzcSR balances Zn 2+ stress adaptation and virulence in Psa M228 via distinct metal-dependent and independent regulatory pathways. Bacterial pathogens harbor numerous two-component systems (TCSs) in their genomes, which enable rapid sensing and response to environmental fluctuations, thereby facilitating dynamic adaptation to diverse ecological niches. Pseudomonas syringae pv. actinidiae (Psa) is the causal agent of kiwifruit bacterial canker (KBC), a devastating disease threatening global kiwifruit production. However, the biological function of the metal-responsive TCS CzcSR in Psa remains largely uncharacterized. In this study, we demonstrated that CzcSR plays a crucial role in regulating Psa pathogenicity in the host plant and the hypersensitive response (HR) in the non-host plant. Under zinc ion (Zn 2+) stress, Psa exhibited suppressed motility and enhanced oxidative stress tolerance; notably, this phenotype depends on the Zn 2+ -binding sites of CzcS and the phosphorylation status of CzcR. However, the key virulence factor type III secretion system (T3SS) of Psa is unaffected by Zn 2+ stress, and CzcSR-mediated regulation of the T3SS is independent of both the Zn 2+ -binding sites of CzcS and the phosphorylation status of CzcR. Instead, CzcR controls T3SS expression by binding to the promoter region of hrpR and modulates the c-di-GMP level via interacting with diguanylate cyclase (DGC) PSA₄781. Collectively, our findings expand CzcSR's functional repertoire, highlight TCS complexity, and deepen understanding of TCS versatility—CzcSR integrates Zn 2+ signals for canonical regulation of phenotypes (e. g. , motility, antioxidant defense) while using a signal-independent mechanism for T3SS control.