Polarized exocytosis is essential for plant development and defence. The exocyst, an octameric protein complex that tethers exocytotic vesicles to the plasma membrane, targets exocytosis. Upon pathogen attack, secreted materials form papillae to halt pathogen penetration. To determine if the exocyst is directly involved in targeting exocytosis to infection sites, information about its localization is instrumental. Here, we investigated exocyst subunit localization in the moss Physcomitrella patens upon pathogen attack and infection by Phytophthora capsici. Time-gated confocal microscopy was used to eliminate autofluorescence of deposited material around infection sites, allowing the visualization of the subcellular localization of exocyst subunits and of v-SNARE Vamp72A1-labelled exocytotic vesicles during infection. This showed that exocyst subunits Sec3a, Sec5b, Sec5d, and Sec6 accumulated at sites of attempted pathogen penetration. Upon pathogen invasion, the exocyst subunits accumulated on the membrane surrounding papilla-like structures and hyphal encasements. Vamp72A1-labelled vesicles were found to localize in the cytoplasm around infection sites. The re-localization of exocyst subunits to infection sites suggests that the exocyst is directly involved in facilitating polarized exocytosis during pathogenesis.

中文翻译：

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时间门控共聚焦显微镜显示在植物-病原体界面上囊外亚基的积累。

极化胞吐作用对于植物发育和防御至关重要。外囊是一种八聚体蛋白复合物，可将胞吐小泡束缚在质膜上，是针对胞吐作用的。病原体侵袭时，分泌的物质形成乳头以阻止病原体渗透。为了确定胞囊是否直接参与将胞吐作用靶向感染部位，有关其定位的信息是有用的。在这里，我们调查了病原体侵染和辣椒疫霉感染后苔藓小立碗藓中囊外亚基的定位。时间门控共聚焦显微镜用于消除感染部位周围沉积物质的自发荧光，从而使感染过程中囊外亚基和v-SNARE Vamp72A1标记的胞吞小泡的亚细胞定位可视化。这表明囊外亚基Sec3a，Sec5b，Sec5d和Sec6累积在病原体试图穿透的位点。病原体入侵后，囊外亚基积聚在乳头状结构和菌丝包膜周围的膜上。发现Vamp72A1标记的囊泡位于感染部位周围的细胞质中。外囊亚基到感染部位的重新定位表明，在发病过程中，外囊直接参与促进极化的胞吐作用。