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. 1995 Mar 28;92(7):2979–2983. doi: 10.1073/pnas.92.7.2979

Membrane permeabilization by Listeria monocytogenes phosphatidylinositol-specific phospholipase C is independent of phospholipid hydrolysis and cooperative with listeriolysin O.

H Goldfine 1, C Knob 1, D Alford 1, J Bentz 1
PMCID: PMC42342  PMID: 7708759

Abstract

We have examined potential cooperative interactions of Listeria monocytogenes phosphatidylinositol-specific phospholipase C (PI-PLC) and listeriolysin O (LLO), a pore-forming hemolysin, in a liposome lysis assay. Large unilamellar vesicles, approximately 0.1 micron in diameter, encapsulating the fluorescent probe calcein, were treated with PI-PLC or LLO at pH 6.0, and each was capable of causing dye release. With phosphatidylcholine/phosphatidylinositol/cholesterol liposomes at 0.1 microM lipid, minimal release of dye was observed on addition of 80 pM LLO or 7 nM PI-PLC. Addition of the two proteins together produced rapid dye release. Unexpectedly, essentially identical results were obtained with phosphatidylcholine/cholesterol liposomes. Thus, the effect of PI-PLC did not depend on lipid hydrolysis. Both proteins also released inulin (M(r) 5200) from liposomes. Membrane permeabilization was not accompanied by membrane fusion. Very little dye release from phosphatidylcholine/phosphatidylinositol/cholesterol liposomes was seen with PI-PLC from Bacillus thuringiensis, and addition of this enzyme to LLO produced no additional dye release; however PI-PLC from L. monocytogenes cooperated with perfringolysin O from Clostridium perfringens. PI-PLC from L. monocytogenes and LLO bind to phosphatidylcholine/cholesterol liposomes, and the rate of binding of each protein was not influenced by the presence of the other. These data support a postulated accessory role for PI-PLC with LLO in lysing the primary phagosome of a macrophage.

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Selected References

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