Secretion of the Francisella pathogenicity island effector protein PdpE requires components of both the Sec and Type IV pilus secretion pathways

Francisella pathogenicity determinant protein E (PdpE) is a protein of unknown function encoded within the Type VI secretion system (T6SS) gene cluster of the Francisella pathogenicity island, a region essential for bacterial intracellular replication and virulence. We previously showed that PdpE is secreted by the bacteria via a T6SS-independent pathway, but the specific pathway involved was unclear. In this study, we investigated PdpE secretion in Francisella novicida and demonstrated that it requires components of both the Sec general secretory pathway and the Type IV pilus (T4P) secretion system. F. novicida possesses two SecB paralogs, and either of the two paralogs is sufficient for bacterial viability and for T4P-mediated secretion of chitinase and PepO enzymatic activities. In contrast, we found that both paralogs are required for T4P-mediated secretion of PdpE. Bacterial two-hybrid and pulldown assays confirmed interactions between these two SecB chaperones. Genomic analyses indicate that the presence of two secB paralogs is a conserved feature across the family Francisellaceae, including species that lack pdpE, suggesting that-while they have overlapping and partially redundant functions-the two paralogs have evolved to fulfill additional specialized chaperone and/or regulatory functions.IMPORTANCEOur research shows that pathogenicity determinant protein E (PdpE) secretion in Francisella requires components of both the Sec general secretory pathway and the Type IV pilus (T4P) secretion system, making it the first gene product encoded within a Type VI secretion system gene cluster known to use an alternative pathway. Although genetically linked to PdpC, PdpE's reliance on an alternative secretion pathway may offer insight into its physiological role. Furthermore, we show that dual SecB chaperone paralogs are required for T4P-mediated PdpE secretion by Francisella, and bioassays indicate a direct interaction between the two distinct chaperones. Genomic analysis reveals that possession of two secB paralogs is a conserved trait within the Francisellaceae family.