Showing papers by "Shin-ichi Ayabe published in 2006"

Cytochrome P450s in flavonoid metabolism

Abstract: In this review, cytochrome P450s characterized at the molecular level catalyzing aromatic hydroxylations, aliphatic hydroxylations and skeleton formation in the flavonoid metabolism are surveyed. They are involved in the biosynthesis of anthocyanin pigments and condensed tannin (CYP75, flavonoid 3′,5′-hydroxylase and 3′-hydroxylase), flavones [CYP93B, (2S)-flavanone 2-hydroxylase and flavone synthase II], and leguminous isoflavonoid phytoalexins [CYP71D9, flavonoid 6-hydroxylase; CYP81E, isoflavone 2′-hydroxylase and 3′-hydroxylase; CYP93A, 3,9-dihydroxypterocarpan 6a-hydroxylase; CYP93C, 2-hydroxyisoflavanone synthase (IFS)]. Other P450s of the flavonoid metabolism include methylenedioxy bridge forming enzyme, cyclases producing glyceollins, flavonol 6-hydroxylase and 8-dimethylallylnaringenin 2′-hydroxylase. Mechanistic studies on the unusual aryl migration by CYP93C, regulation of IFS expression in plant organs and its biotechnological applications are introduced, and flavonoid metabolisms by non-plant P450s are also briefly discussed.

Expression of the 1-aminocyclopropane-1-carboxylic acid deaminase gene requires symbiotic nitrogen-fixing regulator gene nifA2 in Mesorhizobium loti MAFF303099.

Abstract: Many soil bacteria contain 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase, which degrades ACC, a precursor of the phytohormone ethylene. In order to examine the regulation of the acdS gene encoding ACC deaminase in Mesorhizobium loti MAFF303099 during symbiosis with the host legume Lotus japonicus, we introduced the β-glucuronidase (GUS) gene into acdS so that GUS was expressed under control of the acdS promoter, and we also generated disruption mutants with mutations in a nitrogen fixation regulator gene, nifA. The histochemical GUS assay showed that there was exclusive expression of acdS in mature root nodules. Two homologous nifA genes, mll5857 and mll5837, were found in the symbiosis island of M. loti and were designated nifA1 and nifA2, respectively. Quantitative reverse transcription-PCR demonstrated that nifA2 disruption resulted in considerably diminished expression of acdS, nifH, and nifA1 in bacteroid cells. In contrast, nifA1 disruption slightly enhanced expression of the acdS transcripts and suppressed nifH to some extent. These results indicate that the acdS gene and other symbiotic genes are positively regulated by the NifA2 protein, but not by the NifA1 protein, in M. loti. The mode of gene expression suggests that M. loti acdS participates in the establishment and/or maintenance of mature nodules by interfering with the production of ethylene, which induces negative regulation of nodulation.

Plant Lanosterol Synthase : Divergence of the Sterol and Triterpene Biosynthetic Pathways in Eukaryotes

Abstract: Sterols, essential eukaryotic constituents, are biosynthesized through either cyclic triterpenes, lanosterol (fungi and animals) or cycloartenol (plants). The cDNA for OSC7 of Lotus japonicus was shown to encode lanosterol synthase (LAS) by the complementation of a LAS-deficient mutant yeast and structural identification of the accumulated lanosterol. A double site-directed mutant of OSC7, in which amino acid residues crucial for the reaction specificity were changed to the cycloartenol synthase (CAS) type, produced parkeol and cycloartenol. The multiple amino acid sequence alignment of a conserved region suggests that the LAS of different eukaryotic lineages emerged from the ancestral CAS by convergent evolution.

A single-base deletion in soybean flavonol synthase gene is associated with magenta flower color

Abstract: The Wm locus of soybean [Glycine max (L.) Merr.] controls flower color. Dominant Wm and recessive wm allele of the locus produce purple and magenta flower, respectively. A putative full-length cDNA of flavonol synthase (FLS), gmfls1 was isolated by 5' RACE and end-to-end PCR from a cultivar Harosoy with purple flower (WmWm). Sequence analysis revealed that gmfls1 consisted of 1,208 nucleotides encoding 334 amino acids. It had 59-72% homology with FLS proteins of other plant species. Conserved dioxygenase domains A and B were found in the deduced polypeptide. Sequence comparison between Harosoy and Harosoy-wm (magenta flower mutant of Harosoy; wmwm) revealed that they differed by a single G deletion in the coding region of Harosoy-wm. The deletion changed the subsequent reading frame resulting in a truncated polypeptide consisting of 37 amino acids that lacked the dioxygenase domains A and B. Extracts of E. coli cells expressing gmfls1 of Harosoy catalyzed the formation of quercetin from dihydroquercetin, whereas cell extracts expressing gmfls1 of Harosoy-wm had no FLS activity. Genomic Southern analysis suggested the existence of three to four copies of the FLS gene in the soybean genome. CAPS analysis was performed to detect the single-base deletion. Harosoy and Clark (WmWm) exhibited longer fragments, while Harosoy-wm had shorter fragments due to the single-base deletion. The CAPS marker co-segregated with genotypes at Wm locus in a F(2) population segregating for the locus. Linkage mapping using SSR markers revealed that the Wm and gmfls1 were mapped at similar position in the molecular linkage group F. The above results strongly suggest that gmfls1 represents the Wm gene and that the single-base deletion may be responsible for magenta flower color.

Isolation and characterization of a cDNA encoding polyketide reductase in Lotus japonicus

Abstract: Leguminous plants have a unique pathway for production of 5-deoxy-type flavonoids and isoflavonoids that is distinct from the general flavonoid pathways. 5-Deoxy(iso)flavonoids are believed to play important eco-physiological roles as antimicrobial compounds and symbiotic signals toward rhizobia. The branching point of the 5-deoxyflavonoid pathway is the formation of the deoxy-type chalcone (isoliquiritigenin), which is catalyzed by the co-action of chalcone synthase and polyketide reductase (PKR). In the course of the comparative genomics of legume-specific genes, we cloned a putative cDNA for PKR (cPKR1) of a model legume Lotus japonicus (Regel) K. Larsen. Genomic Southern analysis showed that L. japonicus has a gene family composed of two to four paralogous PKR genes. The overexpression of cPKR1 in a red- flowered cultivar of petunia, "Polo Red Target", reduced anthocyanin accumulation and caused the formation of isoliquiritigenin and its putative derivatives. These results suggested that PKR1 encodes a PKR that functions in planta.

DNAs coding for flavone synthases, methods of using flavone synthase DNAs, and plants, flowers, and vectors containing flavone synthase DNAs

Abstract: DNA obtained, for example, from snapdragon or torenia, encoding an enzyme that can convert flavanones directly to flavones, and its uses; the DNA and amino acid sequences for enzymes encoded thereby are listed as SEQ.ID. No. 1 & 2 and 3 & 4, for example. Introduction of the genes into plants can, for example, alter the flower colors of the plants.

Flavone synthases, methods of using flavone synthases, and plants expressing flavone synthases

Abstract: A [DNA encoding a] flavone synthase that synthesizes flavones from flavanones, vectors containing said DNAs encoding said flavone synthases, and plants expressing same are disclosed. Methods of producing the protein which synthesizes flavones from flavanones, methods of altering the composition of flavonoids or the amount of flavonoids in a plant, methods of altering flower color and plant photosensitivity, and methods of controlling interactions between plants expressing the protein and microorganisms are also disclosed.