Lenzi Luisa, Caruso Carla, Bianchedi Pier Luigi, Pertot Ilaria, Perazzolli Michele
Research and Innovation Center, Fondazione Edmund Mach (FEM), Via E. Mach, 1, 38010 S. Michele all'Adige, Italy Department of Ecological and Biological Sciences, University of Tuscia, Via San Camillo de Lellis, 01100 Viterbo, Italy.
Department of Ecological and Biological Sciences, University of Tuscia, Via San Camillo de Lellis, 01100 Viterbo, Italy.
Plant Cell Physiol. 2016 Jan;57(1):69-81. doi: 10.1093/pcp/pcv166. Epub 2015 Nov 6.
Grapevine is one of the most important fruit crops in the world, and it is highly susceptible to downy mildew caused by the biotrophic oomycete Plasmopara viticola. Gene expression profiling has been used extensively to investigate the regulation processes of grapevine-P. viticola interaction, but all studies to date have involved the use of whole leaves. However, only a small fraction of host cells is in contact with the pathogen, so highly localized transcriptional changes of infected cells may be masked by the large portion of non-infected cells when analyzing the whole leaf. In order to understand the transcriptional regulation of the plant reaction at the sites of pathogen infection, we optimized a laser microdissection protocol and analyzed the transcriptional changes in stomata cells and surrounding areas of grapevine leaves at early stages of P. viticola infection. The results indicate that the expression levels of seven P. viticola-responsive genes were greater in microdissected cells than in whole leaves, highlighting the site-specific transcriptional regulation of the host response. The gene modulation was restricted to the stomata cells and to the surrounding areas of infected tissues, indicating that the host response is mainly located at the infection sites and that short-distance signals are implicated. In addition, due to the high sensitivity of the laser microdissection technique, significant modulations of three genes that were completely masked in the whole tissue analysis were detected. The protocol validated in this study could greatly increase the sensitivity of further transcriptomic studies of the grapevine-P. viticola interaction.
葡萄是世界上最重要的水果作物之一，极易受到活体营养卵菌葡萄生单轴霉引起的霜霉病的影响。基因表达谱已被广泛用于研究葡萄与葡萄生单轴霉相互作用的调控过程，但迄今为止所有研究都使用了完整叶片。然而，只有一小部分宿主细胞与病原体接触，因此在分析整片叶子时，感染细胞高度局部化的转录变化可能会被大部分未感染细胞掩盖。为了了解病原体感染部位植物反应的转录调控，我们优化了一种激光显微切割方案，并分析了葡萄生单轴霉感染早期葡萄叶片气孔细胞及其周围区域的转录变化。结果表明，七个葡萄生单轴霉响应基因在显微切割细胞中的表达水平高于完整叶片，突出了宿主反应的位点特异性转录调控。基因调节仅限于气孔细胞和受感染组织的周围区域，表明宿主反应主要位于感染部位，并且涉及短距离信号。此外，由于激光显微切割技术的高灵敏度，检测到了在全组织分析中完全被掩盖的三个基因的显著调节。本研究中验证的方案可以大大提高葡萄与葡萄生单轴霉相互作用的进一步转录组学研究的灵敏度。
