Genetic Analysis of Indole-3-Butyric Acid Function in Plants

本文概述了阐明吲哚-3-丁酸（IBA）在模式植物拟南芥中的功能的实验。虽然IBA是植物生长激素生长素的一种天然形式，在商业上被用于促进许多物种的生根，但其作用的分子机制在很大程度上是未知的。有人认为，IBA转化为吲哚-3-乙酸（IAA）的反应类似于脂肪酸降解（b-氧化），这是植物中的过氧化物酶体过程。已分离出一组IBA应答突变体，并将其暂时分为两类：1)过氧化物酶体生物生成或一般氧化缺陷突变体，可能无法有效地将IBA转化为IAA； 2)过氧化物酶体功能明显正常的突变体，可能在IBA感知或信号传导方面存在缺陷。IBA通过在过氧化物酶体中转化为IAA在生长素信号传导中起作用的假设将通过对这些突变体的特征进行验证。将进行以下实验：1)在iba应答突变体中检测与生长素和过氧化物酶体功能相关的表型。2)对两个已确定致病基因的iba反应突变体的特异性缺陷进行研究。一个突变体在过氧化物酶体靶向受体中存在缺陷。过氧化物酶体的形态将被可视化，过氧化物酶体蛋白的亚细胞分布将被检查。第二个突变体的缺陷基因序列表明，编码的蛋白在IBA链缩短后水解IAA- coa释放游离IAA。该酶的底物特异性将被测试。3)其他几种iba反应突变体中的缺陷基因将被分离，重点是那些具有明显正常过氧化物酶体功能的基因。这些基因和突变体将进一步表征以阐明它们在IBA作用中的作用。4)将分离更多的IBA反应突变体，以鉴定更完整的IBA功能相关基因。为了了解生长素的作用，必须确定植物中发现的各种生长素的功能意义。鉴定参与IBA到IAA转化的基因是了解这种转化调控的第一步。这些知识对于确定IAA代谢的各个方面对活性IAA池的贡献是必不可少的。阐明遗传易感植物中IBA作用的分子机制可能为IBA在农业和园艺中的应用提供见解。鉴定和鉴定将IBA转化为IAA的特异性同工酶，有助于在IBA应用通常无效的难根品种中对其进行改良。这项研究还将揭示对植物过氧化物酶体的功能和生物发生至关重要的基因，这些基因在哺乳动物或酵母过氧化物酶体中没有发现。最后，所提出的实验将扩大对植物氧化作用的理解，这对于种子储存脂质的利用和其他底物链的缩短都是必不可少的，包括茉莉酸，一种参与防御反应的植物激素。

This proposal outlines experiments to elucidate the function of indole-3-butyric acid (IBA) in the model plant Arabidopsis thaliana. Although IBA, a naturally occurring form of the plant growth hormone auxin, is used commercially to promote rooting in many species, the molecular mechanisms by which it acts are largely unknown. It has been suggested that IBA is converted into indole-3-acetic acid (IAA) using reactions analogous to those of fatty acid degradation (b-oxidation), a peroxisomal process in plants. A set of IBA-response mutants has been isolated and provisionally grouped into two classes: 1) mutants defective in peroxisome biogenesis or general boxidation, which may inefficiently convert IBA to IAA, and 2) mutants with apparently normal peroxisome function, which may be defective in IBA perception or signaling. The hypothesis that IBA functions in auxin signaling through its conversion to IAA in peroxisomes will be tested by characterizing these mutants. The following experiments will be conducted: 1) Phenotypes related to auxin and peroxisomal functions will be examined in the IBA-response mutants. 2) Specific defects of two IBA-response mutants for which the responsible genes have been identified will be investigated. One mutant is defective in a peroxisomal targeting receptor. Peroxisome morphology will be visualized and the subcellular distribution of peroxisomal proteins will be examined. The sequence of the gene defective in the second mutant suggests that the encoded protein hydrolyzes IAA-CoA to release free IAA following IBA chain shortening. The substrate specificity of this enzyme will be tested. 3) The genes defective in several other IBA-response mutants will be isolated, with emphasis on those having apparently normal peroxisomal function. These genes and mutants will be further characterized to elucidate their role in IBA action. 4) Additional IBA-response mutants will be isolated to identify a more complete set of genes involved in IBA function.To understand auxin action, the functional significance of the various auxins found in plants must be determined. Identifying genes involved in IBA to IAA conversion is a first step to understanding the regulation of this conversion. This knowledge is essential in determining the contributions of the various facets of IAA metabolism to the active IAA pool. Elucidating of the molecular mechanisms of IBA action in a genetically tractable plant may provide insights for agricultural and horticultural IBA uses. Identifying and characterizing the specific isozymes that convert IBA to IAA may facilitate their modification in difficult-to-root cultivars where IBA application is normally ineffective. This research will also reveal genes critical for the function and biogenesis of plant peroxisomes, which carry out functions not found in mammalian or yeast peroxisomes. Finally, the proposed experiments will expand understanding of plant boxidation, which is essential both for the utilization of seed storage lipids and for chain shortening of other substrates, including jasmonic acid, a plant hormone involved in defense responses.