An acyltransferase-like gene obtained by differential gene expression profiles of quinolizidine alkaloid-producing and nonproducing cultivars of Lupinus angustifolius

Bunsupa, Somnuk; Okada, Taketo; Saito, Kazuki; Yamazaki, Mami

doi:10.5511/plantbiotechnology.10.1109b

Cited by 26 publications

(40 citation statements)

References 17 publications

(7 reference statements)

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“…A BLASTN search was conducted against the NLL Tanjil gene annotation (Hane et al, ) using the following GenBank accessions: LaL/ODC ; AB560664, LaHMT/HLT ; EF381744 and LaAT ; and AB581532 (Bunsupa et al, , ; Okada et al, ; e‐value <0.0001 and identity >70%). A single gene was returned for both LaL/ODC ( Lup009726.1 ) and LaAT ( Lup021586.1 ), whereas two genes were returned for LaHMT/HLT ( Lup022250.1 and Lup022251.1 ), which were 13 Kb apart on the same scaffold.…”

Section: Methodsmentioning

confidence: 99%

Characterization of the genetic factors affecting quinolizidine alkaloid biosynthesis and its response to abiotic stress in narrow‐leafed lupin (Lupinus angustifoliusL.)

et al. 2018

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Quinolizidine alkaloids (QAs) are toxic secondary metabolites that complicate the end use of narrow-leafed lupin (NLL; Lupinus angustifolius L.) grain, as levels sometimes exceed the industry limit for its use as a food and feed source. The genotypic and environmental influences on QA production in NLL are poorly understood. Here, the expression of QA biosynthetic genes was analysed in vegetative and reproductive tissues of bitter (high QA) and sweet (low QA) accessions. It was demonstrated that sweet accessions are characterized by lower QA biosynthetic gene expression exclusively in leaf and stem tissues than bitter NLL, consistent with the hypothesis that QAs are predominantly produced in aerial tissues and transported to seeds, rather than synthesized within the seed itself. This analysis informed our identification of additional candidate genes involved in QA biosynthesis. Drought and temperature stress are two major abiotic stresses that often occur during NLL pod set. Hence, we assessed the effect of drought, increased temperature, and their combination, on QA production in three sweet NLL cultivars. A cultivar-specific response to drought and temperature in grain QA levels was observed, including the identification of a cultivar where alkaloid levels did not change with these stress treatments.

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Section: Methodsmentioning

confidence: 99%

Characterization of the genetic factors affecting quinolizidine alkaloid biosynthesis and its response to abiotic stress in narrow‐leafed lupin (Lupinus angustifoliusL.)

et al. 2018

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“…Therefore, the expression of L/ODC, and thus the subsequent biosynthesis of QAs themselves, may be regulated at a steady state mRNA level by transcription factor(s). By PCR-select subtraction, the fragments of the candidate transcription factors that might regulate the expression of the genes in the QA biosynthesis were obtained; these include DNA binding protein EREBP-3, Leu zipper protein, and coronatine-insensitive 1 (Bunsupa et al, 2011). Since the enzyme activity and the transcript level of an acyltransferase committed in the terminal step of QA biosynthesis do not differ in bitter and sweet cultivars (Hirai et al, 2000;Okada et al, 2005), this finding is interesting in terms of the regulation of the whole biosynthetic pathway.…”

Section: Discussion Physiological Roles Of Ldc In Qa-producing Plantsmentioning

confidence: 99%

“…To isolate cDNAs involved in QA biosynthesis, we conducted a differential PCR-select subtraction analysis using cDNAs from the seedlings of two different Lupinus angustifolius cultivars: a QA-producing bitter cultivar (cv Fest) and a sweet cultivar that does not produce QA (cv Uniharvest) (Bunsupa et al, 2011). The bitter and sweet cultivars are similar in flowering time and pod shattering but differ in alkaloid contents and the colors of the flower and seed coat (Oram, 1983).…”

Section: Cloning Of Cdnas Encoding Ldc From Qa-producing Plantsmentioning

confidence: 99%

Lysine Decarboxylase Catalyzes the First Step of Quinolizidine Alkaloid Biosynthesis and Coevolved with Alkaloid Production in Leguminosae

et al. 2012

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Lysine decarboxylase (LDC) catalyzes the first-step in the biosynthetic pathway of quinolizidine alkaloids (QAs), which form a distinct, large family of plant alkaloids. A cDNA of lysine/ornithine decarboxylase (L/ODC) was isolated by differential transcript screening in QA-producing and nonproducing cultivars of Lupinus angustifolius. We also obtained L/ODC cDNAs from four other QA-producing plants, Sophora flavescens, Echinosophora koreensis, Thermopsis chinensis, and Baptisia australis. These L/ODCs form a phylogenetically distinct subclade in the family of plant ornithine decarboxylases. Recombinant L/ODCs from QA-producing plants preferentially or equally catalyzed the decarboxylation of L-lysine and L-ornithine. L. angustifolius L/ODC (La-L/ODC) was found to be localized in chloroplasts, as suggested by the transient expression of a fusion protein of La-L/ODC fused to the N terminus of green fluorescent protein in Arabidopsis thaliana. Transgenic tobacco (Nicotiana tabacum) suspension cells and hairy roots produced enhanced levels of cadaverine-derived alkaloids, and transgenic Arabidopsis plants expressing (La-L/ODC) produced enhanced levels of cadaverine, indicating the involvement of this enzyme in lysine decarboxylation to form cadaverine. Site-directed mutagenesis and protein modeling studies revealed a structural basis for preferential LDC activity, suggesting an evolutionary implication of L/ODC in the QAproducing plants. INTRODUCTIONAlkaloids are one of the most diverse groups of natural products and are found in ;20% of plant species. Many of the ;12,000 known alkaloids produced by plants display potent pharmacological activities and several are widely used as pharmaceuticals (Croteau et al., 2000;De Luca and St Pierre, 2000). Alkaloids are derived from the products of primary metabolism with amino acids such as Phe, Tyr, Trp, Orn, and Lys serving as their main precursors (Facchini, 2001). Quinolizidine alkaloids (QAs) are derived from cadaverine, and several hundred structurally related compounds have been identified that are distributed mostly within the Leguminosae (Ohmiya et al., 1995;Michael, 2008). Some QAs exhibit beneficial pharmacological properties, such as cytotoxic, antiarrhythmic, oxytocic, hypoglycemic, and antipyretic activities, and thus can be used as drugs (Ohmiya et al., 1995). They can therefore serve as potential starting compounds in the development of new drugs and in pest control for plants .QAs are synthesized through the cyclization of the cadaverine unit (Golebiewski and Spenser, 1988), which is produced through the action of Lys decarboxylase (LDC; EC 4.1.1.18) (Figure 1), via a postulated enzyme-bound intermediate . The various skeletons of QAs [e.g., bicyclic alkaloids such as (2)-lupinine and (+)-epilupinine or tetracyclic alkaloids such as (+)-multiflorine, (+)-lupanine, and (+)-matrine] are then further modified by tailoring reactions (e.g., dehydrogenation, oxygenation, esterification, and glycosylation) to yield hundreds of structurally related alkaloids (Ohmiya ...

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“…Isolation of differentially expressed genes between HR and CSC using PCR-select™ cDNA subtraction (Clontech, Japan) was performed according to previously described by Bunsupa et al (2011). HR-specific AP2/ERF unigenes were selected from de novo transcriptome assembly (Yamazaki et al, 2013) using log 2 reads per kilobase of transcript per million mapped reads (RPKM) of HR/CSC ≥1 as a cut-off value.…”

Section: Methodsmentioning

confidence: 99%

Function of AP2/ERF Transcription Factors Involved in the Regulation of Specialized Metabolism in Ophiorrhiza pumila Revealed by Transcriptomics and Metabolomics

et al. 2016

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The hairy roots (HR) of Ophiorrhiza pumila produce camptothecin (CPT), a monoterpenoid indole alkaloid used as a precursor in the synthesis of chemotherapeutic drugs. O. pumila HR culture is considered as a promising alternative source of CPT, however, the knowledge about the biosynthetic pathway and regulatory mechanism is still limited. In this study, five genes that encode AP2/ERF transcription factors, namely OpERF1–OpERF5, were isolated from HR of O. pumila. Phylogenetic analysis of AP2/ERF protein sequences suggested the close evolutionary relationship of OpERF1 with stress-responsive ERF factors in Arabidopsis and of OpERF2 with ERF factors reported to regulate alkaloid production, such as ORCA3 in Catharanthus roseus, NIC2 locus ERF in tobacco, and JRE4 in tomato. We generated the transgenic HR lines of O. pumila, ERF1i and ERF2i, in which the expression of OpERF1 and OpERF2, respectively, was suppressed using RNA interference technique. The transcriptome and metabolome of these suppressed HR were analyzed for functional characterization of OpERF1 and OpERF2. Although significant changes were not observed in the metabolome, including CPT and related compounds, the suppression of OpERF2 resulted in reduced expression of genes in the 2-C-methyl-d-erythritol 4-phosphate and secologanin-strictosidine pathways, which supply a precursor, strictosidine, for CPT biosynthesis. Furthermore, while it was not conclusive for OpERF1, enrichment analysis of differentially expressed genes in the suppressed HR showed that the gene ontology terms for oxidation-reduction, presumably involved in secondary metabolite pathways, were enriched in the ERF2i downregulated gene set. These results suggest a positive role of OpERF2 in regulating specialized metabolism in O. pumila.

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An acyltransferase-like gene obtained by differential gene expression profiles of quinolizidine alkaloid-producing and nonproducing cultivars of Lupinus angustifolius

Cited by 26 publications

References 17 publications

Characterization of the genetic factors affecting quinolizidine alkaloid biosynthesis and its response to abiotic stress in narrow‐leafed lupin (Lupinus angustifoliusL.)

Characterization of the genetic factors affecting quinolizidine alkaloid biosynthesis and its response to abiotic stress in narrow‐leafed lupin (Lupinus angustifoliusL.)

Lysine Decarboxylase Catalyzes the First Step of Quinolizidine Alkaloid Biosynthesis and Coevolved with Alkaloid Production in Leguminosae

Function of AP2/ERF Transcription Factors Involved in the Regulation of Specialized Metabolism in Ophiorrhiza pumila Revealed by Transcriptomics and Metabolomics

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