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The action mechanism of fenoxasulfone was studied by examining the inhibitory effects of this herbicide on the biosynthesis of very-long-chain fatty acids (VLCFAs). Fenoxasulfone treatment decreased VLCFAs, such as C20:0, C20:1, C22:0, C24:0, C24:1 and C26:0 fatty acids, in barnyard millet cultured cells, and increased long-chain-fatty acids and medium-chain-fatty acids, such as C18:0 and C15:0, which are precursors of VLCFAs. Fenoxasulfone potently inhibited activities of VLCFA elongase (VLCFAE) in the microsomal fraction of etiolated barnyard millet seedlings, which catalyzed the elongation steps from C22:0 to C24:0 and C24:0 to C26:0, respectively. These results strongly suggested that fenoxasulfone is a potent inhibitor of plant VLCFAEs and should be categorized within the K3 group of the Herbicide Resistance Action Committee. VLCFAE activity of recombinant Fatty Acid Elongation 1 (FAE1) of Arabidopsis that catalyzes the elongation step from C18:1 to C20:1, was inhibited by fenoxasulfone in a time-dependent manner, which has been shown in the inhibition of VLCFAEs by other well-known VLCFAE-inhibiting herbicides. On the other hand, VLCFAE activity of the microsomal fraction of etiolated barnyard millet seedlings, which catalyzes the elongation step from C24:0 to C26:0, was inhibited by fenoxasulfone in a time-independent manner. This time-independent inhibition raised a new inhibition mechanism of VLCFAE by fenoxasulfone likely with pyroxasulfone, which is classified in the same chemical class as fenoxasulfone.

Target-site and non-target-site herbicide tolerance are caused by the prevention of herbicide binding to the target enzyme and the reduction to a nonlethal dose of herbicide reaching the target enzyme, respectively. There is little information on the molecular mechanisms involved in non-target-site herbicide tolerance, although it poses the greater threat in the evolution of herbicide-resistant weeds and could potentially be useful for the production of herbicide-tolerant crops because it is often involved in tolerance to multiherbicides. Bispyribac sodium (BS) is an herbicide that inhibits the activity of acetolactate synthase. Rice (Oryza sativa) of the indica variety show BS tolerance, while japonica rice varieties are BS sensitive. Map-based cloning and complementation tests revealed that a novel cytochrome P450 monooxygenase, CYP72A31, is involved in BS tolerance. Interestingly, BS tolerance was correlated with CYP72A31 messenger RNA levels in transgenic plants of rice and Arabidopsis (Arabidopsis thaliana). Moreover, Arabidopsis overexpressing CYP72A31 showed tolerance to bensulfuron-methyl (BSM), which belongs to a different class of acetolactate synthase-inhibiting herbicides, suggesting that CYP72A31 can metabolize BS and BSM to a compound with reduced phytotoxicity. On the other hand, we showed that the cytochrome P450 monooxygenase CYP81A6, which has been reported to confer BSM tolerance, is barely involved, if at all, in BS tolerance, suggesting that the CYP72A31 enzyme has different herbicide specificities compared with CYP81A6. Thus, the CYP72A31 gene is a potentially useful genetic resource in the fields of weed control, herbicide development, and molecular breeding in a broad range of crop species.

Transgenic crops such as maize (Zea mays L.), soybeans (Glycine max L. Merr.), canola (Brassica napus L.) and cotton (Gossypium hirsutum L.) have been widely used in the field. In 2009, transgenic crops were cultivated in approximately 134 million hectares in 25 countries, mainly USA, Brazil, Argentina, India, Canada and China (http://www.isaaa.org). The adoption of transgenic crops has steadily increased since 1996 because of their many benefits for farmers. Herbicide resistance is one of the most important agronomic traits conferred onto transgenic crops. Herbicide-resistant crops comprise 62 percent of all transgenic crops (http://www.isaaa.org), and are produced by the introduction of herbicide-resistant genes using genetic transformation. Herbicide resistance can be used as an efficient tool to allow easier weed management. It facilitates control of weed species and contributes to reducing costs, labor, and the waste of chemical spray. Herbicide resistance can also facilitate the selection of transgenic cells from non-transgenic cells as a selectable marker in genetic transformation. Acetolactate synthase (ALS)-inhibiting herbicides are widely used around the world. ALSinhibiting herbicide-resistant weeds were first found in kochia (Kochia scoparia L. Shrad) (Primiani et al., 1990) and prickly lettuce (Lactuca serriola L.) (Mallory-Smith et al., 1990). Subsequently, plants and cultured cells resistant to ALS-inhibiting herbicides have been generated using both conventional mutation breeding and somatic cell selection. Since then, the ALS genes have been cloned and characterized. In most cases, resistance to ALSinhibiting herbicides has been found to be conferred by single or double amino-acid mutations at a particular position in ALS. Mutated ALS genes can be used not only for the generation of herbicide-resistant crops, but also as selectable markers. We are now producing transgenic tall fescue (Festuca arundinacea Schreb.) and maize that are resistant to ALS-inhibiting herbicides using novel mutated ALS genes. This chapter focuses on mutated ALS genes and their application to the production of herbicide-resistant crops and selection of transgenic cells as selectable markers.

Three metabolites of indole-3-acetic acid (IAA), N-(6-hydroxyindol-3-ylacetyl)-phenylalanine (6-OH-IAA-Phe), N-(6-hydroxyindol-3-ylacetyl)-valine (6-OH-IAA-Val), and 1-O-(2-oxoindol-3-ylacetyl)-beta-d-glucopyranose (OxIAA-Glc), were found by a liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS)-based search for oxidative IAA metabolites during the vegetative growth of Arabidopsis. Their structures were confirmed by making a comparison of chromatographic characteristics and mass spectra between naturally occurring compounds and synthetic standards. An incorporation study using deuterium-labeled compounds showed that 6-OH-IAA-Phe and 6-OH-IAA-Val were biosynthesized from IAA-Phe and IAA-Val, respectively, which strongly suggested the formation of these amino acid conjugates of IAA in plants. Both 6-OH-IAA-Phe and 6-OH-IAA-Val were inactive as auxins, as indicated by no significant root growth inhibition in Arabidopsis. Quantitative analysis demonstrated that OxIAA-Glc was present in the largest amount among the metabolites of IAA in Arabidopsis, suggesting that the conversion into OxIAA-Glc represents the main metabolic process regarding IAA in Arabidopsis.