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菌能够利用宿主细胞的肌动蛋白聚合机制推动其穿过细胞质，从而在细胞之间传播。尽管经过多年的广泛研究，人们对这种机制仍然知之甚少，并且几种肌动蛋白调节蛋白的作用尚未确定。其中一种蛋白质——Profilin——的功能仍然存在争议。我们认为，Profilin 在李斯特菌基于肌动蛋白的运动中发挥着关键作用，并且必须集中在细菌表面，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的快速速率。我们相信肌动蛋白-肌动蛋白复合物是向生长的肌动蛋白丝添加肌动蛋白单体的优选种类。为此，我们将利用两种实验方法研究该复合物的功能。在第一种方法中，我们将 profilin、肌动蛋白、profilin-actin 复合物或突变的 profilin-actin 复合物引入感染李斯特菌的 PtK2 细胞中。我们预计添加肌动蛋白-肌动蛋白复合物将比其他添加物更能增强细菌的运动性。为了补充这些实验，我们还将将相同的蛋白质引入到Profilin耗尽的非洲爪蟾卵提取物中，以支持李斯特菌。在无细胞模型中，我们还期望profilin-actin复合物成为恢复细菌运动性的最有效成分。最后，我们将利用荧光能量转移探索 profilin 与肌动蛋白聚合区中发现的其他蛋白质的相互作用。这些研究对于确定 profilin 在李斯特菌基于肌动蛋白的运动中的作用至关重要，这将进一步了解这种细菌如何引起疾病。