Functional Characterization of the Arabidopsis thaliana ARG1 Gene Involved in Gravity Signal Transduction

拟南芥 ARG1 基因参与重力信号转导的功能表征

• 批准号: 9905675
• 负责人: Patrick Masson
• 金额: $ 30万
• 依托单位: University of Wisconsin-Madison
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-08-15 至 2002-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9905675&HistoricalAwards=false
• 关键词: Functional; Characterization; Arabidopsis; thaliana; ARG1

Plant organs use the gravity vector as an environmental cue to direct their growth. Gravitropism allows roots to grow downward into the soil where they take up the water and mineral ions required for plant growth and development. It also forces shoots to grow upward toward light, allowing them to photosynthesize. This important plant growth response to its environment also allows plant shoots to resume upward growth after having been prostrated by the action of wind and rain, and roots to resume downward growth after having overcome an obstacle placed in their growth path.A change in the orientation of a specific plant organ within the gravity field is perceived mainly by the sedimentation of dense amyloplasts in the cytoplasm of specialized cells named statocytes. Amyloplast sedimentation activates a signal transduction pathway that result in the production of a physiological signal, believed to be an auxin and/or apoplastic Ca2+ gradient across the gravistimulated organ. That signal is then transmitted to the site of response where it promotes a differential cellular growth responsible for the development of curvature. In roots, gravity sensing is thought to occur in the columella cells of the root cap, while the differential growth response occurs in the distal and central elongation zones.Very little is known about the molecular mechanisms allowing plant organs to convert the physical information derived from amyloplast sedimentation into a physiological signal. A molecular genetic approach using Arabidopsis thaliana now allows investigations that should help to better understand that process. Mutations in the ARG1 gene result in a specific defect in root and hypocotyl gravitropism. Mutant organs show wild-type growth rates, growth sensitivities to phytohormones and develop wild-type kinetics of phototropic response, strongly suggesting a specific defect in the early phases of gravity signal transduction. ARG1 encodes a novel dnaJ-like protein containing a putative coiled coil domain with amino acid similarity with coiled coils found in a number of proteins known to interact with the cytoskeleton. Interestingly, the cytoskeleton has been proposed to be implicated in the sensing phase, but not in the curvature response phase, of gravitropism. Based on these data, it is hypothesized that the ARG1 protein interacts with the cytoskeleton in the statocytes, facilitating the transduction of physical forces derived from amyloplast sedimentation into a physiological signal. To test this model the pattern of ARG1 expression will be analyzed in Arabidopsis thaliana seedlings, minimal spatial ARG1 expression requirements will be defined for complete restoration of gravitropism in transgenic arg1 plants, and the regions of ARG1 that are required for its function in gravitropism will be investigated. In situ immunofluorescence strategies will be used to localize the putative ARG1 protein within the columella cells of the root cap and in hypocotyl cells, and to compare its localization with the distribution of microtubules and microfilaments in statocytes. Proteins that interact with the various domains of ARG1 will be identified, and the genes encoding them will be cloned and characterized. Finally, ARG1 paralogs will be identified and functionally characterized. This combination of genetic, physiological and immunocytological studies will help define the molecular function(s) of ARG1 and ARG1 paralogs. Interestingly, a highly conserved ARG1 ortholog was found in Coenorhabditis elegans. This observation suggests an important role for the corresponding protein in the regulation of plant and animal growth, development and/or responses to the environment. Thus, it is expected that research will provide the foundation for a better understanding of the functions played by this novel type of dnaJ-like protein in higher eukaryotes.

植物器官利用重力矢量作为环境线索来指导它们的生长。 向重力性允许根向下生长到土壤中，在那里它们吸收植物生长和发育所需的水和矿物质离子。 它还迫使嫩芽朝着光线向上生长，使它们能够进行光合作用。 这种重要的植物生长对环境的反应也允许植物枝条在受到风雨的作用后恢复向上生长，在重力场中，一种特定植物器官的方向变化主要是通过在特化细胞的细胞质中沉积致密的淀粉体来感知的，平衡细胞 淀粉体沉降激活导致生理信号产生的信号转导途径，该生理信号被认为是生长素和/或跨重力刺激的器官的质外体Ca 2+梯度。 然后，该信号被传递到响应部位，在那里它促进了负责曲率发展的差异细胞生长。在根中，重力感应被认为发生在根冠的柱细胞中，而差异生长反应发生在远端和中央伸长区。关于植物器官将淀粉体沉积的物理信息转化为生理信号的分子机制知之甚少。 一种利用拟南芥的分子遗传学方法现在允许调查，应该有助于更好地理解这一过程。 ARG 1基因的突变导致根和下胚轴向地性的特定缺陷。 突变体器官显示野生型生长速率，生长敏感性植物激素和发展野生型动力学的向光性反应，强烈表明在重力信号转导的早期阶段的特定缺陷。 ARG 1编码一种新的dnaJ样蛋白，含有一个假定的卷曲螺旋结构域，与已知与细胞骨架相互作用的许多蛋白质中发现的卷曲螺旋具有氨基酸相似性。有趣的是，细胞骨架被认为与向地性的感知阶段有关，但与向地性的曲率响应阶段无关。基于这些数据，假设ARG 1蛋白与平衡细胞中的细胞骨架相互作用，促进将淀粉体沉降产生的物理力转导为生理信号。 为了测试该模型，将在拟南芥幼苗中分析ARG 1表达的模式，将定义在转基因arg 1植物中完全恢复向地性的最小空间ARG 1表达要求，并将研究ARG 1在向地性中的功能所需的区域。 原位免疫荧光策略将被用来本地化的根冠和下胚轴细胞的柱状细胞内的推定ARG 1蛋白，并比较其定位与微管和微丝在平衡细胞中的分布。将鉴定与ARG 1的各个结构域相互作用的蛋白质，并克隆和表征编码它们的基因。最后，将鉴定ARG 1旁系同源物并对其功能进行表征。 这种遗传、生理和免疫细胞学研究的结合将有助于确定ARG 1和ARG 1旁系同源物的分子功能。 有趣的是，在秀丽隐杆线虫中发现了高度保守的ARG 1直系同源物。 这一观察结果表明，相应的蛋白质在调节植物和动物的生长，发育和/或对环境的反应中起着重要作用。 因此，它是预期的研究将提供更好地理解这种新型的dnaJ样蛋白在高等真核生物中发挥的功能的基础。