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Cells have evolved endocytic pathways to internalize different molecules, to regulate intra-cellular communication and their interaction with external environment. Pathogens have co-evolved with cells to exploit these processes for infections. In particular, the exotoxins of bacteria target precise cellular-pathways to get internalized and exert their activity in the cytosol. Therefore, studying these intoxication processes provide unique understanding not only of bacterial infections, but also of host cellular mechanisms. Previous work on anthrax toxins have shown that the initial steps of toxin entry, which occur at the plasma-membrane, are modulated by S-acylation. In this thesis, the quest for a deeper understanding of anthrax toxin entry led us to study the cellular trafficking of an acyl-thioesterase, APT2 which de-acylates anthrax toxin receptor, CMG2 at plasma-membrane. Here, we show that APT2 accesses cellular membranes using multi-step process including its S-acylation at the endoplasmic reticulum (ER) and Golgi. This S-acylation event is indispensable for its Golgi localization and trafficking to plasma-membrane, where it modifies local substrates such as CMG2. At plasma-membrane, APT2 is deacylated by resident acyl-thioesterase(s). Toxin endocytosis further relies on the existence of lipid-nanodomains which promote clustering of receptors and cleavage of toxin subunits by pro-protein convertases. A screen aimed at identifying novel genes involved in anthrax toxin uptake unraveled an unexpected mechanism underlying the formation of these nanostructures. Our data show that lipid exchange at the ER-Golgi membrane contact sites is required for nanodomain assembly at the plasma-membrane. At these contacts, OSBP and CERT complexes interact via the p24 family members, TMED2/10, making a supercomplex which drives cholesterol and ceramide transfer from ER to Golgi membranes. Lastly, we show that this ER-Golgi localized supercomplex is maintained by S-acylation. Altogether, these data highlight S-acylation and lipid exchange at the ER-Golgi membrane contact sites as major regulators of anthrax toxin entry in human cells.
Françoise Gisou van der Goot Grunberg, Laurence Gouzi Abrami, Valeria Cagno
Vassily Hatzimanikatis, Françoise Gisou van der Goot Grunberg, Graham Knott, Kathryn Hess Bellwald, Laurence Gouzi Abrami, Luciano Andres Abriata, Béatrice Kunz, Sylvia Ho, Patrick Alain Sandoz, Catherine Maclachlan, Robin Alexander Denhardt-Eriksson, Gard Spreemann