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Browsing by Subject "Indole-3-acetic acid"

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    New analytical methodologies in the study of auxin biochemistry
    (2014-08) Yu, Peng
    Auxin is the essential plant hormone that regulates many aspects of plant growth and development. Plants typically possess highly complex biochemical networks to regulate the homeostasis of the active hormone, through the regulation of biosynthesis, degradation, transport and conjugation. Biosynthesis, among other processes, has been of particular importance and warranted extensive studies over the decades of auxin research. A number of pathways were proposed and some enzymes potentially involved have been characterized. Stable isotope labeling and turnover studies have proven very useful in these investigative efforts. With the advancement of analytical and computational technologies, it is now feasible to concurrently analyze the turnover patterns of all the precursors in the entire auxin biosynthesis network. I devoted chapter two of the dissertation to establish LC-MS methods to concurrently quantify most of the auxin precursors and deployed different isotopic labeling strategies for turnover studies in Arabidopsis thaliana. Preliminary results showed that indole-3-pyruvate (IPyA) and indole-3-acetaldehyde (IAAld) were among the fastest to be labeled and a key regulatory step existed between IPyA and indole-3-acetic acid (IAA). Chapter three reported the discrepancy and the study of the amount of the total IAA determined by alkaline hydrolysis and that of the summation of all known forms of IAA conjugates in Arabidopsis. My results indicated that chemical artifacts induced by harsh chemical treatments were responsible for a significant portion of the unknown putative IAA conjugates. In chapter four, I described a facile way to directly survey a plant extract for its indole profile, notably IAA conjugates, based on high resolution and accurate mass (HR/AM) liquid chromatography-mass spectrometry (LC-MS). The method was successfully applied to Glycine max, Solanum lycopersicum, Cocos nucifera, and Ginkgo biloba. Together, these investigations and developments have led to an improved understanding of auxin metabolism and now provide useful tools for subsequent studies.
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    Roles of multiple mechanisms in regulating auxin levels during plant growth and development.
    (2012-04) Liu, Xing
    Auxins, primarily indole-3-acetic acid (IAA), are endogenous plant hormones well known as key regulators of plant growth and development. Both genetic and biochemical studies have demonstrated that plants have developed a complex system to regulate the level of IAA, including biosynthesis of IAA from Trp-dependent and Trp-independent pathways, polar auxin transport, conjugation and hydrolysis of auxin. To accurately measure changes in IAA levels and identify pathways that contribute to the changes, I developed methods for quantitative analyses of auxin levels, auxin biosynthesis, and polar auxin transport. Using radioisotope labeling and stable-isotope dilution, I found that in etiolated tomato seedlings, a brief light exposure increased both IAA biosynthesis in the upper tissue sections and polar IAA transport in hypocotyls in a phytochrome-dependent manner, leading to unchanged free IAA levels in the top section and increased free IAA levels in the lower hypocotyl regions. In addition, using stable-isotope labeling and stable-isotope dilution, I quantified polar auxin transport in Arabidopsis hypocotyls, and I found that the transport of indole-3-butyric acid (IBA), another endogenous auxin, was much lower than IAA and that its transport mechanism was distinct from IAA transport. I also found that a small amount of IBA metabolic products, such as ester-linked IBA and IAA, was transported, while the majority of transported IAA remained as free IAA, suggesting that the polar transport of IAA could directly change the level of IAA while the transport of IBA could be an additional means to regulate IAA. In summary, my studies provide comprehensive views of auxin regulation in plants under different physiological conditions, showing that multiple mechanisms cooperatively regulate local auxin levels.