Reliable detection and quantification of plasmodesmal callose in Nicotiana benthamiana leaves during defense responses.
Reliable detection and quantification of plasmodesmal callose in Nicotiana benthamiana leaves during defense responses.
复制标题
在防御反应过程中对本塞姆氏烟草叶片中胞间连丝胼胝质进行可靠的检测和定量。
DOI：
10.1101/2023.09.30.560305
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Burch-Smith,TessaM
中科院分区：
文献类型：
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作者：
Sankoh,AmieF;Adjei,Joseph;Roberts,DanielM;Burch-Smith,TessaM
胼胝质是一种β-（1，3）-D-葡聚糖聚合物，对于调节通过胞间连丝（PD）的细胞间运输至关重要。病原体操纵PD定位的蛋白质，使细胞间的运输，通过去除胼胝质在PD，或相反，通过增加胼胝质积累在PD限制感染期间的细胞间运输。植物防御激素如水杨酸调节PD定位蛋白，以控制PD和先天免疫防御反应（如系统获得性抗性）期间的细胞间运输。因此，测量植物中PD处的胼胝质沉积已经成为用于评估植物免疫期间细胞间运输活性的流行参数。尽管这个指标很受欢迎，但对于如何进行这些测量并没有标准。在这项研究中，三种常用的方法来识别和定量PD胼胝质苯胺蓝染色进行了评估，以确定最有效的烟草叶片模型。结果表明，最可靠的方法是苯胺蓝染色和荧光显微镜测量胼胝质沉积在固定的组织。还比较了用于图像分析的手动或半自动工作流程，发现产生了类似的结果，尽管半自动工作流程产生了更广泛的数据点分布。
Callose, a beta-(1,3)-D-glucan polymer, is essential for regulating intercellular trafficking via plasmodesmata (PD). Pathogens manipulate PD-localized proteins to enable intercellular trafficking by removing callose at PD, or conversely by increasing callose accumulation at PD to limit intercellular trafficking during infection. Plant defense hormones like salicylic acid regulate PD-localized proteins to control PD and intercellular trafficking during innate immune defense responses such as systemic acquired resistance. Measuring callose deposition at PD in plants has therefore emerged as a popular parameter for assessing the intercellular trafficking activity during plant immunity. Despite the popularity of this metric there is no standard for how these measurements should be made. In this study, three commonly used methods for identifying and quantifying PD callose by aniline blue staining were evaluated to determine the most effective in the Nicotiana benthamiana leaf model. The results reveal that the most reliable method used aniline blue staining and fluorescent microscopy to measure callose deposition in fixed tissue. Manual or semi-automated workflows for image analysis were also compared and found to produce similar results although the semi-automated workflow produced a wider distribution of data points.