Clostridium septicum Alpha Toxin Uses Glycosylphosphatidylinositol-anchored Protein Receptors

Abstract

The alpha toxin produced by Clostridium septicum is a channel-forming protein that is an important contributor to the virulence of the organism. Chinese hamster ovary (CHO) cells are sensitive to low concentrations of the toxin, indicating that they contain toxin receptors. Using retroviral mutagenesis, a mutant CHO line (BAG15) was generated that is resistant to alpha toxin. FACS analysis showed that the mutant cells have lost the ability to bind the toxin, indicating that they lack an alpha toxin receptor. The mutant cells are also resistant to aerolysin, a channel-forming protein secreted by Aeromonas spp., which is structurally and functionally related to alpha toxin and which is known to bind to glycosylphosphatidylinositol (GPI)-anchored proteins, such as Thy-1. We obtained evidence that the BAG15 cells lack N-acetylglucosaminyl-phosphatidylinositol deacetylase-L, needed for the second step in GPI anchor biosynthesis. Several lymphocyte cell lines lacking GPIanchored proteins were also shown to be less sensitive to alpha toxin. On the other hand, the sensitivity of CHO cells to alpha toxin was increased when the cells were transfected with the GPI-anchored folate receptor. We conclude that alpha toxin, like aerolysin, binds to GPIanchored protein receptors. Evidence is also presented that the two toxins bind to different subsets of GPIanchored proteins.

Bacterial protein toxins and lipids: pore formation or toxin entry into cells

“…The hydrophobic amino acid side chains of each β-strand are probably located on the outside of the β-barrel and interact with the lipid bilayer, whereas the alternating hydrophilic residues are directed toward the fluid-filled channel. The α-toxin pores are estimated to be 1-2 nm in diameter, a size similar to that of pores from other related toxins (Ballard et al, 1993;Gordon et al, 1999;Tweten et al, 2001;Melton et al, 2004).…”

Identification of a target protein of Hydra actinoporin-like toxin-1 (HALT-1) using GST affinity purification and SILAC-based quantitative proteomics

“…Therefore, the folate receptor alpha is localized on the outer surface of the cell membrane and could serve as the membrane attachment site for extracellular proteins, possibly including HALT-1. This hypothesis is supported by the fact that the alpha-toxin of Clostridium septicum binds to the GPI-anchored form of folate receptor alpha (Gordon et al 1999) before it is cleaved approximately 4 kDa from the C-terminus to become an active cytolysin (Melton-Witt et al 2006). Similarly, we suggest that folate receptor alpha could serve as a membrane receptor for HALT-1 instead of the lipid sphingomyelin, which has been previously thought to be the solely attachment site for HALT-1 on the cell membrane (Liew et al 2015).…”

Association of Helicobacter pylori Vacuolating Toxin (VacA) with Lipid Rafts

“…Two pore-forming toxins, aerolysin and Clostridium septicum alpha toxin, each bind to GPI-anchored proteins, which are enriched in lipid raft microdomains of the plasma membrane (57,66). Other pore-forming toxins, such as perfringolysin, bind to cholesterol components of lipid rafts (43).…”