Melendez, A. V. | Search bioRxiv

bioRxiv2024

Borg3 / Cdc42EP5 regulates septin recruitment to primary cilia

Septin GTPases form linear hexa- or octameric rods that polymerize into higher order structures. They are incorporated into the cytoskeleton and involved in vital cellular functions. Among these, they play a role in the formation of primary cilia. Primary cilia are evolutionary conserved cellular signaling hubs. While it is accepted that septins localize to primary cilia and are involved in their formation and function, the regulation of septin assembly in the confined ciliary compartment remains elusive. Here we show, that Borg3, also known as Cdc42 effector protein 5 (Cdc42EP5), is an essential component of primary cilia. Specific Borg3 localization is facilitated by switching the Rho-GTPase Cdc42 between an inactive- and active state at the base of the primary cilium. The active form of Cdc42 has a defined localization restricted to the base of the primary cilium. Knockout of Borg3 as well as dysregulation of Cdc42 reduces septin dynamics at cilia and consequently, the formation of cilia. The study demonstrates that Borg3 is a novel and essential regulator of ciliogenesis through the spatiotemporal control of septin dynamics downstream of Cdc42.

cell biology↗

bioRxiv2020

The Gb3-enriched CD59/flotillin plasma membrane domain regulates host cell invasion by Pseudomonas aeruginosa

The opportunistic pathogen Pseudomonas aeruginosa is responsible for a high number of acute and chronic hospital-acquired infections. As it develops more and more resistances against existing antibiotics, P. aeruginosa has been placed highest on the global priority list of antibiotic-resistant bacteria for which alternative treatments are urgently needed. Former studies have highlighted the crucial role of the bacterial lectin LecA and the host cell glycosphingolipid globotriaosylceramide (Gb3) for the cellular uptake of P. aeruginosa into epithelial cells via the lipid zipper mechanism. To further characterize the host cell plasma membrane domain for LecA-driven attachment and invasion, we analyzed the protein and lipid composition of pulled-down membrane domains for novel interaction partners of LecA by mass spectrometry. We unraveled a predilection of LecA for binding to saturated Gb3 species in the extracellular membrane leaflet and an induction of dynamic phosphatidylinositol (3,4,5)-trisphosphate clusters at the intracellular leaflet co-localizing with sites of LecA binding. Moreover, we identified the GPI-anchored protein CD59 and flotillins, known as cargo and eponymous component of flotillin-assisted endocytosis, as LecA interaction partners. Depletion of each of these host cell proteins resulted in more than 50% of reduction in invasiveness of the P. aeruginosa strain PAO1 highlighting the importance of this LecA-induced plasma membrane domain. Our strategy to reduce the complexity of host-pathogen interactions by first identifying interaction partners of a single virulence factor and subsequently transferring these findings to the bacterium has been proven to be a successful approach in elucidating the molecular mechanisms of bacterial infections.

cell biology↗