Institute of Biology, Biotechnology and Environmental Protection, Faculty of Natural Sciences, University of Silesia in Katowice, Katowice, Poland.
Methods Mol Biol. 2022;2457:351-365. doi: 10.1007/978-1-0716-2132-5_24.
Plasmodesmata (PD) are membraneous channels that span cell walls of adjacent cells to establish the symplasm. These connections are unique to plants and enable the cell-to-cell exchange of information via the symplasm. However, not every plant cell is connected to its neighbor. Absence of PD and lack of communication (symplasmic isolation) are important regulators of cell differentiation. To determine cell-to-cell symplasmic connectivity, the distribution of fluorescent tracers can be analyzed. Here, we describe in detail the entire procedure for conducting such analysis using fluorescence and confocal microscopy to study molecular fluxes in fluorescence recovery after photobleaching (FRAP) experiments. Studies using fluorochromes and fluorescent-labeled dextrans successfully inform the degree of symplasmic connectivity between cells in zygotic and somatic embryos. Small molecules, such as water and ions, travel through PD but also transcription factors and different types of RNA. Studies of symplasmic communication are important to determine the spatio-temporal correlation between cell differentiation and the exchange of information between cells. This information is necessary to determine the role of symplasmic communication during embryogenesis, which is a very important stage in plant development and morphogenesis.
胞间连丝（PD）是跨越相邻细胞细胞壁的膜状通道，建立共质体。这些连接是植物特有的，使细胞通过共质体进行信息交换。然而，并非每个植物细胞都与其相邻细胞相连。PD 的缺失和缺乏通讯（共质体隔离）是细胞分化的重要调节因子。为了确定细胞间的共质体连接性，可以分析荧光示踪剂的分布。在这里，我们详细描述了使用荧光和共焦显微镜进行此类分析的整个过程，以研究荧光漂白后恢复（FRAP）实验中的分子通量。使用荧光染料和荧光标记的葡聚糖的研究成功地告知了合子和体细胞胚中细胞之间共质体连接的程度。小分子，如水和离子，通过 PD 运输，但也有转录因子和不同类型的 RNA。共质体通讯的研究对于确定细胞分化和细胞间信息交换之间的时空相关性非常重要。这些信息对于确定胚胎发生过程中共质体通讯的作用是必要的，胚胎发生是植物发育和形态发生的一个非常重要的阶段。
