ADP-ribosylation factor (ARF) family proteins, one type of small guanine-nucleotide-binding (G) proteins, play a central role in regulating vesicular traffic and organelle structures in eukaryotes. The Arabidopsis (Arabidopsis thaliana) genome contains more than 21 ARF proteins, but relatively little is known about the functional heterogeneity of ARF homologs in plants. Here, we characterized the function of a unique ARF protein, ARFD1B, in Arabidopsis. ARFD1B exhibited both cytosol and punctate localization patterns, colocalizing with a Golgi marker in protoplasts and transgenic plants. Distinct from other ARF1 homologs, overexpression of a dominant-negative mutant form of ARFD1B did not alter the localization of the Golgi marker mannosidase I (ManI)-RFP in Arabidopsis cells. Interestingly, the ARFD1 artificial microRNA knockdown mutant arfd1 displayed a deleterious growth phenotype, while this phenotype was restored in complemented plants. Further, confocal imaging and transmission electron microscopy analyses of the arfd1 mutant revealed defective cell plate formation and abnormal Golgi morphology. Pull-down and liquid chromatography-tandem mass spectrometry analyses identified Coat Protein I (COPI) components as interacting partners of ARFD1B, and subsequent bimolecular fluorescence complementation, yeast (Saccharomyces cerevisiae) two-hybrid, and co-immunoprecipitation assays further confirmed these interactions. These results demonstrate that ARFD1 is required for cell plate formation, maintenance of Golgi morphology, and plant growth in Arabidopsis.

中文翻译：

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ADP-核糖基化因子 D1 调节拟南芥中的高尔基体形态、细胞板形成和植物生长。

ADP-核糖基化因子 (ARF) 家族蛋白是一种小鸟嘌呤核苷酸结合 (G) 蛋白，在调节真核生物中的囊泡运输和细胞器结构中发挥着核心作用。拟南芥（Arabidopsis thaliana）基因组含有超过21种ARF蛋白，但人们对植物中ARF同源物的功能异质性知之甚少。在这里，我们表征了拟南芥中独特的 ARF 蛋白 ARFD1B 的功能。ARFD1B 表现出胞质和点状定位模式，与原生质体和转基因植物中的高尔基体标记共定位。与其他 ARF1 同源物不同，ARFD1B 显性失活突变体的过度表达不会改变拟南芥细胞中高尔基体标记物甘露糖苷酶 I (ManI)-RFP 的定位。有趣的是，ARFD1人工microRNA敲低突变体arfd1表现出有害的生长表型，而这种表型在互补植物中得到恢复。此外，arfd1突变体的共焦成像和透射电子显微镜分析揭示了细胞板形成缺陷和高尔基体形态异常。Pull-down 和液相色谱-串联质谱分析确定了外壳蛋白 I (COPI) 成分是 ARFD1B 的相互作用伙伴，随后的双分子荧光互补、酵母（酿酒酵母）双杂交和免疫共沉淀分析进一步证实了这些相互作用。这些结果表明 ARFD1 是拟南芥细胞板形成、高尔基体形态维持和植物生长所必需的。