Feeding Cells Induced by Phytoparasitic Nematodes Require γ-Tubulin Ring Complex for Microtubule Reorganization

Reorganization of the microtubule network is important for the fast isodiametric expansion of giant-feeding cells induced by root-knot nematodes. The efficiency of microtubule reorganization depends on the nucleation of new microtubules, their elongation rate and activity of microtubule severing factors. New microtubules in plants are nucleated by cytoplasmic or microtubule-bound γ-tubulin ring complexes. Here we investigate the requirement of γ-tubulin complexes for giant feeding cells development using the interaction between Arabidopsis and Meloidogyne spp. as a model system. Immunocytochemical analyses demonstrate that γ-tubulin localizes to both cortical cytoplasm and mitotic microtubule arrays of the giant cells where it can associate with microtubules. The transcripts of two Arabidopsis γ-tubulin (TUBG1 and TUBG2) and two γ-tubulin complex proteins genes (GCP3 and GCP4) are upregulated in galls. Electron microscopy demonstrates association of GCP3 and γ-tubulin as part of a complex in the cytoplasm of giant cells. Knockout of either or both γ-tubulin genes results in the gene dose-dependent alteration of the morphology of feeding site and failure of nematode life cycle completion. We conclude that the γ-tubulin complex is essential for the control of microtubular network remodelling in the course of initiation and development of giant-feeding cells, and for the successful reproduction of nematodes in their plant hosts.

微管网络的重组对于根结线虫诱导形成的取食巨细胞的快速等径扩张至关重要。微管重组的效率取决于新生微管的成核过程、其延伸速率以及微管切割因子的活性。植物中的新生微管由细胞质结合或微管结合的γ微管蛋白环复合物（γ-tubulin ring complexes）介导成核。本研究以拟南芥与根结线虫属（Meloidogyne spp.）物种的互作为模式系统，探究γ微管蛋白复合物在取食巨细胞发育过程中的必要性。免疫细胞化学分析显示，γ微管蛋白定位于取食巨细胞的细胞质皮层与有丝分裂微管阵列，并可与微管相结合。拟南芥的两个γ微管蛋白基因（TUBG1与TUBG2）以及两个γ微管蛋白复合物蛋白基因（GCP3与GCP4）的转录本在根结中表达上调。电子显微镜观察证实，GCP3与γ微管蛋白以复合物形式存在于取食巨细胞的细胞质中。单独敲除或同时敲除任一或两个γ微管蛋白基因，均会导致取食位点形态发生基因剂量依赖性改变，并使线虫无法完成生活史。本研究最终表明，γ微管蛋白复合物对于取食巨细胞起始与发育过程中的微管网络重塑调控，以及线虫在植物宿主体内的成功繁殖均不可或缺。