Cholesterol-dependent cytolysins: from water-soluble state to membrane pore

Abstract

The cholesterol-dependent cytolysins (CDCs) are a family of bacterial toxins that are important virulence factors for a number of pathogenic Gram-positive bacterial species. CDCs are secreted as soluble, stable monomeric proteins that bind specifically to cholesterol-rich cell membranes, where they assemble into well-defined ring-shaped complexes of around 40 monomers. The complex then undergoes a concerted structural change, driving a large pore through the membrane, potentially lysing the target cell. Understanding the details of this process as the protein transitions from a discrete monomer to a complex, membrane-spanning protein machine is an ongoing challenge. While many of the details have been revealed, there are still questions that remain unanswered. In this review, we present an overview of some of the key features of the structure and function of the CDCs, including the structure of the secreted monomers, the process of interaction with target membranes, and the transition from bound monomers to complete pores. Future directions in CDC research and the potential of CDCs as research tools will also be discussed.

Critical Role of Lipid Scramblase TMEM16F in Phosphatidylserine Exposure and Repair of Plasma Membrane after Pore Formation

“…Interestingly, when individual cells were analyzed, $50% of LLO-treated cells were positive for annexin V but remained negative for PI, indicating that lipid scrambling was taking place even in the absence of compromised membrane integrity ( Figure 1B). Analogous effects were seen with other pore-forming agents; namely, the bacterial toxin SLO (which is implicated in the pathogenicity of b-hemolytic Streptococcus strains), the detergent saponin (which creates pores by extracting membrane cholesterol and phospholipids), and the complement cascade (a component of the mammalian immune system capable of killing pathogens and, sometimes, host cells through pore formation) (Brito et al, 2019;Christie et al, 2018;Hamon et al, 2012;Jimenez et al, 2014;Seveau, 2014; Figure 1C; Figure S1B). Similar effects were noted in various lymphoid cell lines, both T and B cells (Figures S1C-S1E).…”

“…These pores are initially sufficient to initiate a flux of extracellular calcium but too small for appreciable entry of PI or DAPI and release of CFSE. In the case of toxins, the smaller pores may be due to insertion of toxin monomers or oligomers, rather than polymers, in the membrane (Christie et al, 2018;Hamon et al, 2012;Radoshevich and Cossart, 2018;Seveau, 2014). The binding of pore-forming agents to the membrane is independent of calcium and TMEM16F.…”

“…In addition to initiating intracellular responses to external cues, the plasma membrane serves as a physical barrier protecting cells from external attack (Andrews and Corrotte, 2018;Christie et al, 2018). Agents such as pore-forming proteins can kill cells directly by binding and damaging the plasma membrane (Christie et al, 2018;Hamon et al, 2012;Radoshevich and Cossart, 2018;Seveau, 2014). Pore-forming agents include a variety of bacterial toxins, such as listeriolysin O (LLO) and streptolysin O (SLO), which create pores in cellular membranes for pathogenicity.…”

All major cholesterol-dependent cytolysins use glycans as cellular receptors

“…The structural basis of CDC-glycan receptor interactions X-ray crystal structures are available for all CDCs investigated in this study [reviewed in (29)]. To identify the structural basis of glycan binding to the CDCs observed in our analyses, crystal soaks and cocrystallization studies were attempted with the highest affinity ligands in Table 1.…”

Listeriolysin O: A phagosome-specific cytolysin revisited

“…Secreted monomers bind to cell membranes and oligomerise into arc and ring prepore assemblies, which may contain up to 50 subunits. Following membrane binding, α‐helical regions in Domain 3 of each monomer refold into two β‐hairpins that insert into the membrane and form a β‐barrel pore (Christie, Johnstone, Tweten, Parker, & Morton, ; Leung et al, ; Mulvihill et al, ). Domain 4 contains the signature undecapeptide sequence (ECTGLAWEWWR) that is the most highly conserved region in the primary CDC sequence and is required for coupling of cholesterol binding to Domain 3 rearrangement (Dowd & Tweten, ).…”