The Role of Profilin in Listeria Actin-Based Motility

Profilin 在李斯特菌肌动蛋白运动中的作用

• 批准号: 6622385
• 负责人: LAURA L LARSON
• 金额: $ 1.82万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-02-28 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6622385
• 关键词: Listeria; Listeria infections; Xenopus oocyte; actin binding protein; actins; bacteria infection mechanism; bacterial proteins; cell free system; cell motility; fluorescence resonance energy transfer; postdoctoral investigator; tissue /cell culture

Listeria monocytogenes is able to spread from cell-to-cell by utilizing the host-cell's mechanism of actin polymerization to propel it through the cytoplasm. Despite years of extensive investigation, this mechanism is still poorly understood, and the roles of several actin regulatory proteins have yet to be established. The function of one of these proteins, profilin, remains controversial. We propose that profilin plays a critical role in actin-based motility in Listeria and must be concentrated on the surface of the bacterium to support the rapid rates of actin assembly associated with the migration of Listeria through the host-cell cytoplasm. We believe that the profilin-actin complex is the preferred species for addition of actin monomers to the growing actin filament. To that end, we will study the function of this complex utilizing two experimental approaches. In the first approach, we will introduce profilin, actin, profilin-actin complex, or mutated profilin-actin complex into PtK2 cells infected with Listeria. We expect than the addition of the profilin-actin complex will enhance bacterial motility more than the other additions. To complement these experiments, we will also introduce the same proteins into profilin-depleted Xenopus egg extracts supporting Listeria. In cell-free model, we also expect the profilin-actin complex to be the most effective component restoring bacterial motility. Finally, we will explore the interaction of profilin with other proteins found in the actin polymerization zone using fluorescent energy transfer. These investigations are central in determining the role of profilin in actin- based motility of Listeria, which will provide further insight into how this bacteria causes disease.

单核细胞增生李斯特氏菌能够通过利用宿主细胞的肌动蛋白聚合机制从细胞到细胞扩散，从而通过细胞质推动它。尽管进行了多年的广泛研究，但这种机制仍然很少了解，并且尚未确定几种肌动蛋白调节蛋白的作用。这些蛋白质之一的功能，profilin，仍然存在争议。我们提出，profilin在基于肌动蛋白的李斯特菌运动中起着至关重要的作用，必须集中在细菌的表面上，以支持与李斯特菌通过宿主细胞细胞质迁移相关的肌动蛋白组装速率。我们认为，在不断增长的肌动蛋白丝中添加肌动蛋白单体是酸性蛋白 - 肌动蛋白复合物。为此，我们将利用两种实验方法研究这种复合物的功能。在第一种方法中，我们将引入profilin，肌动蛋白，酸性肌动蛋白复合物或突变的profilin-actin复合物中，以感染李斯特菌的PTK2细胞。我们期望的是，添加酸蛋白酶 - 肌动蛋白复合物将比其他添加剂增强细菌运动能力。为了补充这些实验，我们还将将相同的蛋白质引入支撑李斯特菌的缺乏的Xenopus卵提取物。在无细胞模型中，我们还期望profilin-actin复合物是恢复细菌运动性的最有效成分。最后，我们将使用荧光能传递探索profilin与在肌动蛋白聚合区中发现的其他蛋白质的相互作用。这些研究在确定profilin在李斯特菌基于肌动蛋白的运动中的作用方面是核心，这将进一步了解该细菌如何引起疾病。