Methods for Detection of Protein Interactions with Plasmodesmata-Localized Reticulons.
Methods for Detection of Protein Interactions with Plasmodesmata-Localized Reticulons.
复制标题
检测蛋白质与胞连丝局部网状相互作用的方法。
DOI：
10.1007/978-1-0716-2132-5_13
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发表时间：
2022
期刊：
影响因子：
--
通讯作者：
Kriechbaumer V
中科院分区：
文献类型：
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作者：
Kriechbaumer V
植物网状蛋白家族蛋白（RTN）通过二聚化和寡聚化使内质网微管化，产生局部内质网膜张力，导致膜弯曲。在胞间连丝（PD）蛋白质组中发现了两种RTN ER成形蛋白，它们可能有助于形成桥连管，桥连管是一种ER衍生的结构，它穿过原发性PD并物理连接两个细胞的ER。在这里，我们描述了两种用于鉴定位于烟草（Nicotiana tabacum）胞质分裂的原发性PD中的两种PD驻留网状蛋白RTN 3和RTN 6的伴侣的方法：使用GFP-Trap®_A珠的免疫沉淀以寻找新的相互作用伴侣，以及FRET-FLIM以测试和定量植物中的直接蛋白质-蛋白质相互作用。
Plant reticulon family proteins (RTN) tubulate the ER by dimerization and oligomerization, creating localized ER membrane tensions that result in membrane curvature. Two RTN ER-shaping proteins have been found in the plasmodesmata (PD) proteome which could potentially contribute to the formation of the desmotubule, an ER-derived structure that crosses primary PD and physically connects the ER of two cells. Here we describe two methods used to identify partners of two PD-resident reticulon proteins, RTN3 and RTN6 that are located in primary PD at cytokinesis in tobacco (Nicotiana tabacum): immunoprecipitations using GFP-Trap®_A beads to find novel interaction partners and FRET-FLIM to test for and quantify direct protein-protein interactionsin planta.