Type 5 secretion as a novel mechanism to deliver protein effectors directly into mammalian host cells

All host-adapted Bordetella species encode large filamentous hemagglutinin-like adhesins termed FhaL. Global expression studies indicate that fhaL is a virulence-activated gene, and FhaL peptides are detected in the secretome of B. pertussis cells cultured under pathogenic growth conditions. Although the FhaL expression profile suggests a function during host colonization, the activity of this putative adhesin has not been examined experimentally. We find that FhaL has a remarkably similar domain architecture as antibacterial CdiA effector proteins, which deliver C-terminal toxin domains into neighboring bacteria to inhibit target-cell growth. We propose the FhaL uses a similar delivery mechanism to translocate putative effector domains into eukaryotic host cells. FhaL carries four cargo modules, each composed of C80 cysteine peptidase and putative effector domains. The peptidase domains are homologous to auto-proteolytic domains found in MARTX toxins, suggesting that FhaL releases it effector domains through auto-processing after translocation into the host cell cytosol. FhaL effector domains share between 25% and 45% pair-wise identity with each other, but are not homologous to any other protein of known function. AlphaFold2 modeling indicates that the effector domains adopt similar folds with distant structural homology to esterases and hydrolases. This project will determine whether FhaL acts as a contact-dependent effector delivery system and test whether the effector domains act as toxic hydrolases to disrupt host cell physiology. Project Number: 1R21AI191036-01 | Fiscal Year: 2025 | NIH Institute/Center: National Institute of Allergy and Infectious Diseases (NIAID) | Principal Investigator: Christopher Hayes | Institution: UNIVERSITY OF CALIFORNIA SANTA BARBARA, SANTA BARBARA, CA | Award Amount: $410,497 | Activity Code: R21 | Study Section: Prokaryotic Cell and Molecular Biology Study Section[PCMB] View on NIH RePORTER: https://reporter.nih.gov/project-details/1R21AI19103601