Cellular Mechanism of Auxin-Induced Proton Excretion

生长素诱导质子排泄的细胞机制

• 批准号: 9316947
• 负责人: Elizabeth Van Volkenburgh
• 金额: $ 21万
• 依托单位: University of Washington
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-01-01 至 1998-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9316947&HistoricalAwards=false
• 关键词: Cellular; Mechanism; Auxin; Induced; Proton

9316947 VanVolkenburgh The PI, Dr. Elizabeth Van Volkenburgh proposes to investigate the cellular mechanism of auxin-induced proton excretion in protoplasts from Avena coleoptiles and tobacco plants. The goal of this research is to determine how the hormone, auxin, activates the ATP-dependent proton-pump in plasma membranes of growing cells. This hormone stimulates cell enlargement in plants, and plays a fundamental role in regulating development during the plants life cycle. It is hypothesized that this ligand initiates cascades of cellular events that lead to pump activation, proton excretion, membrane hyperpolarization, solute accumulation and cell growth. Electrophysiological techniques, primarily whole cell patch clamp methods will be used to determine the possible roles of suspected components of signal transduction cascades leading to changes in ion flux across the plasma membrane. Cytoplasmic contents will be equilibrated with the solution in the patch pipette containing either substances that might initiate ion flux in the absence of auxin, or substances that might block the response to auxin. The pH and pCa of the cytoplasm will be manipulated to determine the effect of these ions of the auxin response. These experiments will provide direct evidence concerning the role of cytoplasmic components in auxin induced proton excretion. %%% Protoplasts will be isolated from the known auxin-sensitive tissue, the Avena coleoptile, and from tobacco mesophyll which reportedly shows an unusually rapid response to auxin. Protoplasts will be characterized for responsiveness to auxin by measurement of auxin-induced protoplasts swelling. The relationship between swelling and proton excretion will be determined. The effect of auxin on the membrane potential of tobacco leaf cells will be compared to that of coleoptiles using microelectrode techniques. Both cell types will be investigated further with patch clamp experiments, and the results will confirm or reject the reports of auxin effects on tobacco protoplasts. The results of these experiments will serve as foundation for further biotechnological manipulation of vascular plants. ***

小行星9316947 主要研究者Elizabeth货车Volkenburgh博士建议研究生长素诱导燕麦胚芽鞘和烟草原生质体质子分泌的细胞机制。 这项研究的目的是确定激素，生长素，如何激活生长细胞质膜中ATP依赖的质子泵。 这种激素刺激植物细胞的扩大，并在调节植物生命周期中的发育中发挥重要作用。 据推测，这种配体启动级联的细胞事件，导致泵激活，质子排泄，膜超极化，溶质积累和细胞生长。 电生理学技术，主要是全细胞膜片钳方法将用于确定信号转导级联的可疑成分的可能作用，导致跨质膜离子通量的变化。 细胞质内容物将用贴片移液管中的溶液平衡，该溶液含有在不存在生长素的情况下可能引发离子通量的物质或可能阻断对生长素的反应的物质。 将操纵细胞质的pH和pCa以确定这些离子对生长素响应的影响。 这些实验将为细胞质组分在生长素诱导的质子分泌中的作用提供直接证据。 原生质体将从已知的对生长素敏感的组织燕麦胚芽鞘和烟草叶肉中分离出来，据报道烟草叶肉对生长素反应异常迅速。 通过测量生长素诱导的原生质体膨胀来表征原生质体对生长素的响应性。 将确定肿胀和质子排泄之间的关系。 利用微电极技术比较了生长素对烟草叶片细胞和胚芽鞘细胞膜电位的影响。 这两种细胞类型将进一步研究与膜片钳实验，结果将证实或拒绝生长素对烟草原生质体的影响的报告。 这些实验结果将为进一步的维管植物生物技术操作奠定基础。 ***