Baranowski Łukasz, Różańska Elżbieta, Sańko-Sawczenko Izabela, Matuszkiewicz Mateusz, Znojek Ewa, Filipecki Marcin, Grundler Florian M W, Sobczak Mirosław
Department of Botany, Warsaw University of Life Sciences-SGGW, Nowoursynowska 159, 02-766, Warsaw, Poland.
Department of Plant Genetics, Breeding and Biotechnology, Warsaw University of Life Sciences-SGGW, Nowoursynowska 159, 02-766, Warsaw, Poland.
Protoplasma. 2019 Mar;256(2):419-429. doi: 10.1007/s00709-018-1303-4. Epub 2018 Sep 5.
Plant parasitic cyst nematodes induce specific hypermetabolic syncytial nurse cell structures in host roots. A characteristic feature of syncytia is the lack of the central vacuole and the formation of numerous small and larger vesicles. We show that these structures are formed de novo via widening of ER cisternae during the entire development of syncytium, whereas in advanced stages of syncytium development, larger vacuoles are also formed via fusion of vesicles/tubules surrounding organelle-free pre-vacuole regions. Immunogold transmission electron microscopy of syncytia localised the vacuolar markers E subunit of vacuolar H-adenosinetriphosphatase (V-ATPase) complex and tonoplast intrinsic protein (γ-TIP1;1) mostly in membranes surrounding syncytial vesicles, thus indicating that these structures are vacuoles and that some of them have a lytic character. To study the function of syncytial vacuoles, changes in expression of AtVHA-B1, AtVHA-B2 and AtVHA-B3 (coding for isoforms of subunit B of V-ATPase), and TIP1;1 and TIP1;2 (coding for γ-TIP proteins) genes were analysed. RT-qPCR revealed significant downregulation of AtVHA-B2, TIP1;1 and TIP1;2 at the examined stages of syncytium development compared to uninfected roots. Expression of VHA-B1 and VHA-B3 decreased at 3 dpi but reached the level of control at 7 dpi. These results were confirmed for TIP1;1 by monitoring At-γ-TIP-YFP reporter construct expression. Infection test conducted on tip1;1 mutant plants showed formation of larger syncytia and higher numbers of females in comparison to wild-type plants indicating that reduced levels or lack of TIP1;1 protein promote nematode development.
植物寄生性胞囊线虫在寄主根中诱导形成特定的高代谢合胞体营养细胞结构。合胞体的一个特征是缺乏中央液泡，并形成许多大小不一的小泡。我们发现，这些结构是在合胞体的整个发育过程中通过内质网池的扩张从头形成的，而在合胞体发育的后期，较大的液泡也是通过围绕无细胞器前液泡区域的小泡/小管融合形成的。合胞体的免疫金透射电子显微镜观察将液泡标记物液泡H-腺苷三磷酸酶（V-ATPase）复合物的E亚基和液泡膜内在蛋白（γ-TIP1;1）主要定位在合胞体小泡周围的膜中，因此表明这些结构是液泡，并且其中一些具有溶酶体特征。为了研究合胞体液泡的功能，分析了AtVHA-B1、AtVHA-B2和AtVHA-B3（编码V-ATPase B亚基的同工型）以及TIP1;1和TIP1;2（编码γ-TIP蛋白）基因表达的变化。RT-qPCR显示，与未感染的根相比，在合胞体发育的检测阶段，AtVHA-B2、TIP1;1和TIP1;2显著下调。VHA-B1和VHA-B3的表达在3 dpi时下降，但在7 dpi时达到对照水平。通过监测At-γ-TIP-YFP报告构建体的表达，对TIP1;1的这些结果进行了验证。对tip1;1突变体植物进行的感染试验表明，与野生型植物相比，形成了更大的合胞体和更多的雌虫，这表明TIP1;（此处原文似乎有误，推测是TIP1;1）蛋白水平降低或缺失促进了线虫的发育。
