An N-Terminal Retention Module Anchors the Giant Adhesin LapA of Pseudomonas fluorescens at the Cell Surface: a Novel Subfamily of Type I Secretion Systems

ABSTRACT LapA of Pseudomonas fluorescens Pf0-1 belongs to a diverse family of cell surface-associated bacterial adhesins that are secreted via the type I secretion system (T1SS). We previously reported that the periplasmic protease LapG cleaves the N terminus of LapA at a canonical dialanine motif to release the adhesin from the cell surface under conditions unfavorable to biofilm formation, thus decreasing biofilm formation. Here, we characterize LapA as the first type I secreted substrate that does not follow the “one-step” rule of T1SS. Rather, a novel N-terminal element, called the retention module (RM), localizes LapA at the cell surface as a secretion intermediate. Our genetic, biochemical, and molecular modeling analyses support a model wherein LapA is tethered to the cell surface through its T1SS outer membrane TolC-like pore, LapE, until LapG cleaves LapA in the periplasm. We further demonstrate that this unusual retention strategy is likely conserved among LapA-like proteins, and it reveals a new subclass of T1SS ABC transporters involved in transporting this group of surface-associated LapA-like adhesins. These studies demonstrate a novel cell surface retention strategy used throughout the Proteobacteria and highlight a previously unappreciated flexibility of function for T1SS. IMPORTANCE Bacteria have evolved multiple secretion strategies to interact with their environment. For many bacteria, the secretion of cell surface-associated adhesins is key for initiating contact with a preferred substratum to facilitate biofilm formation. Our work demonstrates that P. fluorescens uses a previously unrecognized secretion strategy to retain the giant adhesin LapA at its cell surface. Further, we identify likely LapA-like adhesins in various pathogenic and commensal proteobacteria and provide phylogenetic evidence that these adhesins are secreted by a new subclass of T1SS ABC transporters.