Isolation and Characterization of Arabidopsis Mutants with Altered Response to Auxin (Picloram)
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The phyto-hormone auxin is vital for the plant growth and development throughout the plant’s lifecycle. At the molecular level, auxin rapidly modulates the expression of auxin responsive genes by inducing degradation of a family of negative regulators known as Aux/IAA proteins. Besides the major natural auxin indoleacetic acid (IAA), there are several other natural and many synthetic auxins. Picloram and 2,4-dichlorophenoxyacetic acid (2,4-D) are two synthetic auxins that are commonly used as herbicides. Our initial results suggest that picloram may function somewhat differently from other commonly used auxins such as IAA, 2, 4-D and 1-naphthalene acetic acid (1-NAA). To understand the molecular mechanisms of auxin responses in plants, we isolated a range of mutants that exhibit altered response to picloram. The goal of my research was to characterize two of these mutants, pic11 and pic32. The mutant pic32 is hypersensitive to picloram but not to other commonly known auxins. pic32 mutation was mapped to a 62 kb region that includes 17 genes in chromosome V of the Arabidopsis genome. The mutant pic11 is resistant to both picloram and 2,4-D. Compared to wild type,pic11 exhibits many growth and development defects. It produces fewer and shorter root hairs compared to wild type and no lateral roots. Several auxins such as 2,4-D, IAA and IBA stimulate lateral root initiation in pic11 at higher concentrations, suggesting that pic11 mutation does not affect the ability to produce lateral roots, but may diminish sensitivity to auxin. Picloram also induces few lateral roots in pic11; however, all lateral roots initiate at the same point on the root just below to the hypocotyl suggesting that picloram response is severely compromised by the pic11 mutation. While pic11 is also defective in root hair development, it can be rescued by 1-aminocyclopropane-carboxylic acid (ACC), the immediate precursor of ethylene, suggesting that PIC11 functions upstream of ethylene synthesis and signaling. Using map based cloning, the pic11 mutation was identified as an allele (iaa28) of the IAA28 gene that encodes an Aux/IAA protein (IAA28), a repressor of auxin induced gene transcription. pic11/iaa28 causes a gain of function mutation in domain II of IAA28 protein. While the wild type IAA28 interacts with the auxin receptor protein AFB1 in an auxindependent manner, mutation of domain II interferes with this interaction, probably leading to the stabilization of the mutant pic11/iaa28 protein. Results of this study along with previous studies by others suggest that the stabilized mutant protein may directly affect the regulation of other auxin-inducible genes that are important in growth and development.