Department of Plant Pathology, University of Nebraska-Lincoln, Lincoln, Nebraska, United States of America.
Department of Agronomy and Horticulture, University of Nebraska-Lincoln, Lincoln, Nebraska, United States of America.
PLoS Genet. 2018 Nov 21;14(11):e1007814. doi: 10.1371/journal.pgen.1007814. eCollection 2018 Nov.
Like other intracellular eukaryotic phytopathogens, the devastating rice blast fungus Magnaporthe (Pyricularia) oryzae first infects living host cells by elaborating invasive hyphae (IH) surrounded by a plant-derived membrane. This forms an extended biotrophic interface enclosing an apoplastic compartment into which fungal effectors can be deployed to evade host detection. M. oryzae also forms a focal, plant membrane-rich structure, the biotrophic interfacial complex (BIC), that accumulates cytoplasmic effectors for translocation into host cells. Molecular decision-making processes integrating fungal growth and metabolism in host cells with interface function and dynamics are unknown. Here, we report unanticipated roles for the M. oryzae Target-of-Rapamycin (TOR) nutrient-signaling pathway in mediating plant-fungal biotrophic interface membrane integrity. Through a forward genetics screen for M. oryzae mutant strains resistant to the specific TOR kinase inhibitor rapamycin, we discovered IMP1 encoding a novel vacuolar protein required for membrane trafficking, V-ATPase assembly, organelle acidification and autophagy induction. During infection, Δimp1 deletants developed intracellular IH in the first infected rice cell following cuticle penetration. However, fluorescently labeled effector probes revealed that interface membrane integrity became compromised as biotrophy progressed, abolishing the BIC and releasing apoplastic effectors into host cytoplasm. Growth between rice cells was restricted. TOR-independent autophagy activation in Δimp1 deletants (following infection) remediated interface function and cell-to-cell growth. Autophagy inhibition in wild type (following infection) recapitulated Δimp1. In addition to vacuoles, Imp1GFP localized to IH membranes in an autophagy-dependent manner. Collectively, our results suggest TOR-Imp1-autophagy branch signaling mediates membrane homeostasis to prevent catastrophic erosion of the biotrophic interface, thus facilitating fungal growth in living rice cells. The significance of this work lays in elaborating a novel molecular mechanism of infection stressing the dominance of fungal metabolism and metabolic control in sustaining long-term plant-microbe interactions. This work also has implications for understanding the enigmatic biotrophy to necrotrophy transition.
与其他细胞内真核植物病原体一样，破坏性的稻瘟病菌 Magnaporthe (Pyricularia) oryzae 首先通过分泌被植物衍生膜包围的侵袭性菌丝（IH）来感染活宿主细胞。这形成了一个扩展的生物营养界面，将质外体隔室包围起来，真菌效应子可以部署到该隔室中以逃避宿主检测。M. oryzae 还形成了一个焦点、富含植物膜的结构，即生物营养界面复合物（BIC），该结构积累细胞质效应子以转位到宿主细胞中。将真菌在宿主细胞中的生长和代谢与界面功能和动力学整合在一起的分子决策过程尚不清楚。在这里，我们报告了出乎意料的 M. oryzae 雷帕霉素靶蛋白（TOR）营养信号通路在介导植物-真菌生物营养界面膜完整性中的作用。通过对耐特定 TOR 激酶抑制剂雷帕霉素的 M. oryzae 突变株进行正向遗传学筛选，我们发现 IMP1 编码一种新型液泡蛋白，该蛋白需要进行膜运输、V-ATPase 组装、细胞器酸化和自噬诱导。在感染过程中，在角质层穿透后第一个感染的水稻细胞中，Δimp1 缺失突变体形成了细胞内 IH。然而，荧光标记的效应子探针显示，随着生物营养的进展，界面膜完整性受到损害，BIC 被破坏，质外体效应子释放到宿主细胞质中。细胞间生长受到限制。在 Δimp1 缺失突变体中（感染后），TOR 非依赖性自噬激活修复了界面功能和细胞间生长。在野生型中（感染后）抑制自噬会再现 Δimp1 的情况。除了液泡，Imp1GFP 还以自噬依赖性方式定位于 IH 膜上。总的来说，我们的结果表明 TOR-Imp1-自噬分支信号传导介导了膜稳态，以防止生物营养界面的灾难性侵蚀，从而促进真菌在活水稻细胞中的生长。这项工作的意义在于阐述了一种新的感染分子机制，强调了真菌代谢及其在维持长期植物-微生物相互作用中的代谢控制的主导地位。这项工作对于理解神秘的生物营养到坏死营养的转变也具有重要意义。
