Department of Agriculture and Ecology, Faculty of Science, University of Copenhagen, DK-1871 Frederiksberg C, Copenhagen, Denmark.
Proc Natl Acad Sci U S A. 2012 Jul 10;109(28):11443-8. doi: 10.1073/pnas.1117596109. Epub 2012 Jun 25.
Penetration resistance to powdery mildew fungi, conferred by localized cell wall appositions (papillae), is one of the best-studied processes in plant innate immunity. The syntaxin PENETRATION (PEN)1 is required for timely appearance of papillae, which contain callose and extracellular membrane material, as well as PEN1 itself. Appearance of membrane material in papillae suggests secretion of exosomes. These are potentially derived from multivesicular bodies (MVBs), supported by our observation that ARA6-labeled organelles assemble at the fungal attack site. However, the trafficking components that mediate delivery of extracellular membrane material are unknown. Here, we show that the delivery is independent of PEN1 function. Instead, we find that application of brefeldin (BF)A blocks the papillary accumulation of GFP-PEN1-labeled extracellular membrane and callose, while impeding penetration resistance. We subsequently provide evidence indicating that the responsible BFA-sensitive ADP ribosylation factor-GTP exchange factor (ARF-GEF) is GNOM. Firstly, analysis of the transheterozygote gnom(B4049/emb30-1) (gnom(B)(/E)) mutant revealed a delay in papilla formation and reduced penetration resistance. Furthermore, a BFA-resistant version of GNOM restored the BFA-sensitive papillary accumulation of GFP-PEN1 and callose. Our data, therefore, provide a link between GNOM and disease resistance. We suggest that papilla formation requires rapid reorganization of material from the plasma membrane mediated by GNOM. The papilla material is subsequently presumed to be sorted into MVBs and directed to the site of fungal attack, rendering the epidermal plant cell inaccessible for the invading powdery mildew fungus.
对粉孢菌（powdery mildew fungi）穿透抗性的研究，是植物先天免疫中研究得最好的过程之一，这种抗性是通过局部细胞壁附加物（papillae）实现的。结构域交换蛋白 PENETRATION（PEN）1 对于维持细胞壁结构和发挥功能是必需的，这种蛋白定位在表皮细胞表面的 papillae 中，papillae 中包含 callose 和细胞外膜物质，同时 PEN1 自身也存在于 papillae 中。膜物质在 papillae 中的出现表明存在外泌体的分泌。外泌体很可能来源于多泡体（MVBs），这一推测得到了以下观察结果的支持：在真菌攻击部位观察到 ARA6 标记的细胞器聚集。然而，介导细胞外膜物质运输的运输成分仍不清楚。在这里，我们证明这种运输不依赖于 PEN1 功能。相反，我们发现布雷菲德菌素 A（BFA）的应用会阻止 GFP-PEN1 标记的细胞外膜和 callose 在 papillae 中的积累，同时阻碍穿透抗性。随后，我们提供了证据表明，负责 BFA 敏感的 ADP 核糖基化因子-GTP 交换因子（ARF-GEF）是 GNOM。首先，对 transheterozygote gnom（B4049/emb30-1）（gnom（B）（/E））突变体的分析表明，papillae 的形成延迟，穿透抗性降低。此外，BFA 抗性版本的 GNOM 恢复了 GFP-PEN1 和 callose 在 papillae 中的 BFA 敏感积累。因此，我们的数据为 GNOM 与疾病抗性之间建立了联系。我们认为 papillae 的形成需要 GNOM 介导的质膜的快速重组。随后，papillae 中的物质被分拣到 MVBs 中，并定向到真菌攻击部位，使表皮植物细胞对入侵的粉孢菌不可接近。
