Burda Paul-Christian, Roelli Matthias A, Schaffner Marco, Khan Shahid M, Janse Chris J, Heussler Volker T
Institute of Cell Biology, University of Bern, Bern, Switzerland; Graduate School of Cellular Biology, University of Bern, Bern, Switzerland.
Institute of Cell Biology, University of Bern, Bern, Switzerland.
PLoS Pathog. 2015 Mar 18;11(3):e1004760. doi: 10.1371/journal.ppat.1004760. eCollection 2015 Mar.
The coordinated exit of intracellular pathogens from host cells is a process critical to the success and spread of an infection. While phospholipases have been shown to play important roles in bacteria host cell egress and virulence, their role in the release of intracellular eukaryotic parasites is largely unknown. We examined a malaria parasite protein with phospholipase activity and found it to be involved in hepatocyte egress. In hepatocytes, Plasmodium parasites are surrounded by a parasitophorous vacuole membrane (PVM), which must be disrupted before parasites are released into the blood. However, on a molecular basis, little is known about how the PVM is ruptured. We show that Plasmodium berghei phospholipase, PbPL, localizes to the PVM in infected hepatocytes. We provide evidence that parasites lacking PbPL undergo completely normal liver stage development until merozoites are produced but have a defect in egress from host hepatocytes. To investigate this further, we established a live-cell imaging-based assay, which enabled us to study the temporal dynamics of PVM rupture on a quantitative basis. Using this assay we could show that PbPL-deficient parasites exhibit impaired PVM rupture, resulting in delayed parasite egress. A wild-type phenotype could be re-established by gene complementation, demonstrating the specificity of the PbPL deletion phenotype. In conclusion, we have identified for the first time a Plasmodium phospholipase that is important for PVM rupture and in turn for parasite exit from the infected hepatocyte and therefore established a key role of a parasite phospholipase in egress.
细胞内病原体从宿主细胞中协调逸出是感染成功和传播的关键过程。虽然磷脂酶已被证明在细菌从宿主细胞逸出及毒力方面发挥重要作用，但其在细胞内真核寄生虫释放中的作用却 largely unknown。我们研究了一种具有磷脂酶活性的疟原虫蛋白，发现它参与肝细胞逸出过程。在肝细胞中，疟原虫被一层寄生泡膜（PVM）包围，在寄生虫释放到血液之前，这层膜必须被破坏。然而，在分子层面，关于PVM如何破裂知之甚少。我们发现伯氏疟原虫磷脂酶（PbPL）定位于受感染肝细胞的PVM上。我们提供的证据表明，缺乏PbPL的寄生虫在产生裂殖子之前肝脏阶段发育完全正常，但在从宿主肝细胞逸出方面存在缺陷。为进一步研究此问题，我们建立了一种基于活细胞成像的检测方法，这使我们能够在定量基础上研究PVM破裂的时间动态。利用该检测方法，我们发现缺乏PbPL的寄生虫PVM破裂受损，导致寄生虫逸出延迟。通过基因互补可恢复野生型表型，证明了PbPL缺失表型的特异性。总之，我们首次鉴定出一种对PVM破裂很重要的疟原虫磷脂酶，进而对寄生虫从受感染肝细胞逸出很重要，因此确立了寄生虫磷脂酶在逸出过程中的关键作用。
