BMC Seminar Thursday 6th October 2016, at 12:10 room 201 Læknagarður

Speaker: Peter Šebo, Laboratory of Molecular Biology of Bacterial Pathogens, Institute of Microbiology, Czech Academy of Sciences, Prague, Czech Republic,

Title: The Yin and the Yang of a bacterial toxin: Fooling host phagocytes and delivering immunotherapeutic T cell vaccines

Abstract: The whooping cough agent, Bordetella pertussis, secretes an adenylate cyclase toxin (ACT) that penetrates host phagocytes bearing the α_Mβ₂ integrin receptor Mac-1 (also known as CR3 or CD11b^/CD18). It targets in particular the sentinel functions of neutrophils, macrophages and the professional antigen presenting dendritic cells (DC, CD11b^high). ACT recognizes a positively charged loop of the CD11b subunit of CR3 near the hinge region outside of the I/domain of CD11b and inserts directly across phagocyte membrane. ACT-mediated Ca²⁺ influx then induces calpain-mediated cleavage of talin, enabling ACT to hijack the receptor and mobilize it into membrane lipid rafts. There, translocation of the AC domain across cell membrane is completed across a tightly sealed protein-lipid interface. The AC binds cytosolic calmodulin and catalyzes conversion of ATP to cAMP, generating supraphysiologic cAMP levels that subvert phagocyte functions, causing phagocyte impotence due to inactivation of the Syk kinase and block of signaling of leukocyte receptors. Activation of PKA through cAMP next provokes transient inactivation of the small GTP-ase RhoA, causing rapid and unproductive cell ruffling. In parallel, transient activation of the tyrosine phosphatase SHP-1 occurs by an as yet unknown PKA-dependent mechanism and causes inhibition of oxidative burst and block of expression of iNOS, preventing bactericidal NO production in phagocytes. Simultaneously, activated SHP-1 causes stabilization of BimEL and activation of Bax, provoking induction of apoptosis.

A subpopulation of ACT molecules oligomerizes into small cation-selective pores that permeabilize cells for potassium efflux. This contributes to induction of maturation of dendritic cells that is, however, hijacked by cAMP signaling, which compromises the capacity of DCs to stimulate antigen-specific T cell immune responses. Migration of the incompletely mature DCs into lymph nodes then likely contributes to suppression of adaptive host immune responses to the pathogen and supports bacterial colonization of the host in early stages of infection. Later in infection, ACT action provokes NALP3 inflammasome activation in dendritic cells, which likely contributes to late inflammatory response and eventual development of Th1/Th17 polarized immune responses that support eventual clearance of the bacterial infection.

The amazing capacity of ACT to accommodate foreign antigenic polypeptides and the capacity of ACT to promote maturation of dendritic cells allowed development of genetically detoxified ACT (dACT) into a novel carrier for delivery of antigens for processing into both the MHC class I and II-restricted presentation pathways. This enables efficient induction of prophylactic as well as therapeutic antigen-specific CD4⁺ and CD8⁺ T cell immune responses. I will conclude by reviewing recent applications of ACT technology for immunotherapy of certain tumors.