Phosphorylation of a novel STK substrate SstP1 links SUF Fe-S cluster biogenesis to oxidative stress resistance and virulence in Streptococcus suis.

Iron-sulfur (Fe-S) clusters are ancient prosthetic groups that are incorporated into apoproteins to generate fully functional Fe-S proteins, representing a conserved regulatory pathway in both Eukarya and Prokaryotes. In bacteria, the sulfur utilization factor (SUF) system mediates Fe-S cluster biosynthesis under stress conditions, thereby supporting essential metabolic processes to resist oxidative stress. In this study, we identify a novel substrate of serine/threonine kinase (STK), the protein with a DUF1831 domain (named as SstP1) in Streptococcus suis. GST pull-down and enzymatic activity assays demonstrated that STK-dependent phosphorylation of SstP1 at residue threonine 35 is required to sustain SUF-dependent Fe-S cluster biosynthesis under oxidative stress through direct interaction with the scaffold protein SufB. Functional analyses revealed that both deletion of sstP1 (ΔsstP1) or introduction of a phospho-ablative T35A mutant impaired bacterial growth and markedly reduced survival under oxidative stress and iron limitation conditions. In a mouse infection model, both the ΔsstP1 and T35A mutants exhibited significantly reduced bacterial burdens in the blood, lung, spleen, and brain, accompanied by attenuated virulence compared to the WT, phospho-mimetic T35E, and sstP1 complementary (CΔsstP1) mutants. Collectively, these findings reveal that STK-mediated phosphorylation of SstP1 at Thr35 is essential for oxidative stress resistance and virulence in S. suis, uncovering a novel mechanism that links phosphorylation signaling to SUF-dependent Fe-S cluster biosynthesis in bacterial pathogenesis.