Pneumolysin-damaged cells benefit from non-homogeneous toxin binding to cholesterol-rich membrane domains

Abstract

Nucleated cells eliminate lesions induced by bacterial pore-forming toxins, such as pneumolysin via shedding patches of damaged plasmalemma into the extracellular milieu. Recently, we have shown that the majority of shed pneumolysin is present in the form of inactive pre-pores. This finding is surprising considering that shedding is triggered by Ca-influx following membrane perforation and therefore is expected to positively discriminate for active pores versus inactive pre-pores. Here we provide evidence for the existence of plasmalemmal domains that are able to attract pneumolysin at high local concentrations. Within such a domain an immediate plasmalemmal perforation induced by a small number of pneumolysin pores would be capable of triggering the elimination of a large number of not yet active pre-pores/monomers and thus pre-empt more frequent and perilous perforation events. Our findings provide further insights into the functioning of the cellular repair machinery which benefits from an inhomogeneous plasmalemmal distribution of pneumolysin.

Structural insights into loss of function of a pore forming toxin and its role in pneumococcal adaptation to an intracellular lifestyle

“…Indeed Ply-H, but not Ply-NH, reduced the cellular internalization of cholera toxin coated latex beads, which enter cells specifically through a lipid raft mediated pathway. Recent studies have described increased affinity of Ply towards cholesterol rich domains(35) and calcium signalling triggered shedding of damaged membrane patches as a repair mechanism for Ply-created pores on plasma membranes(36). Integrating our findings with these reports, we propose that the attempt of SPN to enter host cells is thwarted by Ply-H in its vicinity that forms pores on lipid was not certified by peer review) is the author/funder.…”

Bacterial pore‐forming toxin pneumolysin: Cell membrane structure and microvesicle shedding capacity determines differential survival of immune cell types

“…27 PLY preferentially binds to lymphoid cells, that is, Jurkat T cells. 14,29 Increased numbers of such cholesterol-rich domains or a preferred membrane raft distribution may favor the rapid binding and pore-formation of PLY in Jurkat T cells. It is important to note here that cell surface area and cholesterol content of the microsomal membrane fraction did not significantly differ in the three cell types.…”

“…These microdomains are either permanently present, as a result of protein-driven lipid segregation, or their formation is triggered by PLY, which may stabilize cholesterol-rich platforms and promote self-association and fusion of highly dynamic lipid nanodomains. 14,29 Increased numbers of such cholesterol-rich domains or a preferred membrane raft distribution may favor the rapid binding and pore-formation of PLY in Jurkat T cells. Ca 2+ influx induced by PFTs is a key signal strictly required for plasma membrane repair.…”

“…14,15 For the viability experiments, 2 × 10 6 cells were washed in Ca 2+ Tyrode's buffer (140 mM NaCl, 5 mM KCl, 1 mM MgCl 2 , 10 mM glucose, 10 mM HEPES (pH 7.4), 2.5 mM Ca 2+ ) and incubated with PLY as indicated in Ca 2+ Tyrode's buffer for 30 minutes at room temperature. 14,15 For the viability experiments, 2 × 10 6 cells were washed in Ca 2+ Tyrode's buffer (140 mM NaCl, 5 mM KCl, 1 mM MgCl 2 , 10 mM glucose, 10 mM HEPES (pH 7.4), 2.5 mM Ca 2+ ) and incubated with PLY as indicated in Ca 2+ Tyrode's buffer for 30 minutes at room temperature.…”

The iron chelator Deferasirox causes severe mitochondrial swelling without depolarization due to a specific effect on inner membrane permeability

“…Giant vesicles for measurements of proton permeability were prepared in PBS pH 7.4 containing 0.8 mg/ml pHrodo by gentle swelling as previously reported 66 , trapped in a microfluidic chamber and incubated with DFX or DFA and pHrodo at 28 °C. Control experiments containing DMSO did not alter the results (data not shown).…”