Does Lipid Composition Regulate Anthrax Toxin Uptake

Biological roles of lipids are not limited toproviding structural stability to bilayer membranes. Lipids have numerous functions, e.g. acting in cellular signaling and protein regulation. Cells adjust bilayer biophysical parameters by controlling lipid content and gradient of the plasma and intracellular organelle membranes. Thus, membrane composition changes along the endocytic pathway, with early endosomal membranes resembling plasma membranes, and late endosomal membranes showing dramatic increase in anionic lysobisphosphatidic acid level.Here we investigate the role of the lipid environmentin the anthrax toxin uptake. The anthrax toxin, formed by three proteins (binding protective antigen, PA, and enzymatic lethal factor, LF and edema factor, EF), follows a multistep internalization process which involves PA binding to its cellular receptors, proteolytic cleavage to PA63, oligomerization to form prepores, and LF and EF binding followed by endocytosis. The acidic endosomal environment causes conformational changes in PA63 leading to its insertion into the early endosomal membrane and ion channel formation. The delivery of LF to cytoplasm was suggested to take place later in the endocytic pathway via the back-fusion of intraluminal vesicles of late endosomes. Given the variations in membrane lipid composition of the early endosome, the intraluminal vesicles and the late endosome, it becomes important to understand if these changes influence the PA63 insertion and structural dynamics, LF and EF binding and translocation.Using bilayer lipid membrane technique, we mimic the endosomal membrane composition probing its properties in-vitro. We show that at pH 5.5, PA63 insertion rate increases dramatically in PS compared with PC bilayers, whereas LF binds ten times less effectively to PA63 in PS compared to PC. When the solution pH is raised from 5 to 6.75, this difference is diminished.