Functional characterization of AtRUG8, a protein that modulates plasmodesmal movement of macromolecules, and of three 3 closely related proteins

AtRUG8（一种调节大分子胞间连丝运动的蛋白质）以及 3 个密切相关的蛋白质的功能表征

• 批准号: 121466635
• 负责人: Professor Dr. Norbert Sauer
• 金额: --
• 依托单位: Department Biologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2009
• 资助国家: 德国
• 起止时间: 2008-12-31 至 2012-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/121466635?language=en
• 关键词: Functional; characterization; AtRUG8; protein; modulates

We have shown that soluble GFP that is synthesized in Arabidopsis or tobacco (Nicotiana tabacum) companion cells (CCs) under the control of the Arabidopsis AtSUC2 (SUCROSE TRANSPORTER 2) promoter moves from CCs into sieve elements (SEs), traffics with the photoassimilates from source to sink, and is eventually symplastically unloaded into different terminal sink tissues, such as anthers, young leaves or root tips. Screening of more than 7,000 T-DNA insertion lines that we generated in plants with an AtSUC2-promoter::GFP transgene, identified eight mutants with significantly reduced cell-to-cell mobility of GFP. These mutants were named “reduced unloading of GFP” (rug). So far, one of these mutants, rug8, has been partly characterized on the molecular level. In two independent mutants (rug8-1 & rug8-2) the functionally uncharacterized gene At1g17930 was shown to be responsible for the observed phenotype, and complementation with the wild type sequence restored the mobility of soluble GFP. RUG8 has an N-terminal domain of unknown function (DUF1723) and a predicted Cterminal nuclear localization sequence (NLS). In fact, analyses of the subcellular localization confirmed that RUG8 localizes to the nucleus and a yeast 2-hybrid screen revealed interaction of RUG8 with the AP2/ERF-type transcription factor ERF38, an Arabidopsis protein that regulates the expression of genes involved in cell wall thickening. In the proposed project we will study the physiological role of the RUG8 protein and the effect of rug8 mutations on the size exclusion limit (SEL) of plasmodesmata. Moreover, we will screen for additional RUG8 interactors, search for altered gene expression in rug8 mutants, and study the mode of interaction between RUG8 and ERF38. Eventually we will extend these analyses to the three closely related Arabidopsis genes At1g48120, At2g04865 and At2g25010.

我们已经证明，在拟南芥AtSUC 2（蔗糖转运蛋白2）启动子的控制下，拟南芥或烟草（烟草）伴胞（CC）中合成的可溶性GFP从CC移动到筛元件（SE），与源到库的光合同化物一起运输，并最终共质体卸载到不同的终端库组织，如花药，幼叶或根尖。我们在带有AtSUC 2启动子：：GFP转基因的植物中筛选了7,000多个T-DNA插入系，鉴定出8个GFP细胞间迁移率显著降低的突变体。这些突变体被命名为“GFP卸载减少”（rug）。到目前为止，这些突变体之一，rug 8，已部分特征在于在分子水平上。在两个独立的突变体（rug 8 -1和rug 8 -2）的功能不明的基因At 1g 17930被证明是负责观察到的表型，与野生型序列互补恢复可溶性GFP的流动性。RUG 8具有一个功能未知的N端结构域（DUF 1723）和一个预测的C端核定位序列（NLS）。事实上，亚细胞定位的分析证实RUG 8定位于细胞核，酵母双杂交筛选揭示RUG 8与AP 2/ERF型转录因子ERF 38的相互作用，ERF 38是一种调节细胞壁增厚相关基因表达的拟南芥蛋白。在拟议的项目中，我们将研究RUG 8蛋白的生理作用和rug 8突变对胞间连丝的大小排阻限制（SEL）的影响。此外，我们将筛选额外的RUG 8相互作用，寻找rug 8突变体中改变的基因表达，并研究RUG 8和ERF 38之间的相互作用模式。最终，我们将这些分析扩展到三个密切相关的拟南芥基因At 1g 48120，At 2g 04865和At 2g 25010。