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首页> 外文期刊>Infection and immunity >Compartmentalization of the Broad-Range Phospholipase C Activity to the Spreading Vacuole Is Critical for Listeria monocytogenes Virulence
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Compartmentalization of the Broad-Range Phospholipase C Activity to the Spreading Vacuole Is Critical for Listeria monocytogenes Virulence

机译：范围广的磷脂酶C活性对扩散液泡的室间隔对于单核细胞增生性李斯特菌毒力至关重要

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Listeria monocytogenes is a bacterial pathogen that multiplies in the cytosol of host cells and spreads directly from cell to cell by using an actin-based mechanism of motility. The broad-range phospholipase C (PC-PLC) of L. monocytogenes contributes to bacterial escape from vacuoles formed upon cell-to-cell spread. PC-PLC is made as an inactive proenzyme whose activation requires cleavage of an N-terminal propeptide. During infection, PC-PLC is activated specifically in acidified vacuoles. To assess the importance of compartmentalizing PC-PLC activity during infection, we created a mutant that makes constitutively active PC-PLC (the plcBΔpro mutant). Results from intracellular growth and cell-to-cell spread assays showed that the plcBΔpro mutant was sensitive to gentamicin, suggesting that unregulated PC-PLC activity causes damage to host cell membranes. This was confirmed by the observation of a twofold increase in staining of live infected cells by a non-membrane-permeant DNA fluorescent dye. However, membrane damage was not sufficient to cause cell lysis and was dependent on bacterial cell-to-cell spread, suggesting that damage was localized to bacterium-containing filopodia. Using an in vivo competitive infection assay, we observed that the plcBΔpro mutant was outcompeted up to 200-fold by the wild-type strain in BALB/c mice. Virulence attenuation was greater when mice were infected orally than when they were infected intravenously, presumably because the plcBΔpro mutant was initially outcompeted in the intestines, reducing the number of mutant bacteria reaching the liver and spleen. Together, these results emphasize the importance for L. monocytogenes virulence of compartmentalizing the activity of PC-PLC during infection.
机译：单核细胞增生李斯特菌是一种细菌病原体，它在宿主细胞的细胞质中繁殖，并通过基于肌动蛋白的运动机制直接在细胞之间传播。 L的宽范围磷脂酶C（PC-PLC）。单核细胞增生因子有助于细菌从细胞间扩散形成的液泡中逸出。 PC-PLC被制成无活性的酶，其激活需要裂解N端前肽。在感染期间，PC-PLC在酸化的液泡中被专门激活。为了评估在感染过程中区分PC-PLC活性的重要性，我们创建了一个突变型突变体，该突变体构成了具有活性的PC-PLC（ plcB Δpro突变体）。细胞内生长和细胞间扩散试验的结果表明， plcB Δpro突变体对庆大霉素敏感，表明未调节的PC-PLC活性会破坏宿主细胞膜。通过观察非膜渗透性DNA荧光染料使活感染细胞染色增加了两倍，可以证实这一点。然而，膜损伤不足以引起细胞裂解，并且依赖于细菌细胞间的扩散，这表明损伤局限于含有细菌的丝状伪足。使用体内竞争感染试验，我们观察到野生型菌株在野生型BALB / c小鼠中， plcB Δpro突变体的竞争能力高达200倍。小鼠经口感染的毒力衰减比经静脉内感染的毒力衰减更大，这可能是因为 plcB Δpro突变体最初在肠道中的竞争能力较弱，从而减少了到达肝脏和脾脏的突变细菌的数量。总之，这些结果强调了 L的重要性。单核细胞增生性在感染过程中分隔PC-PLC活性的毒力。

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