Biology subjects

Nakabayashi, R.

Publications and source records attributed to Nakabayashi, R..

4 recordsLinked to original sources

bioRxiv2022

Genome-wide association studies of seed metabolites identify loci controlling specialized metabolites in Arabidopsis thaliana

Plants synthesize specialized metabolites to facilitate environmental and ecological interactions. During evolution, plants diversified in their potential to synthesize these metabolites. Quantitative differences in metabolite levels of natural Arabidopsis thaliana accessions can be employed to unravel the genetic basis for metabolic traits using genome-wide association studies (GWAS). Here, we performed metabolic GWAS (mGWAS) on seeds of a panel of 315 A. thaliana natural accessions, including the reference genotypes C24 and Col-0, for polar and semi-polar seed metabolites using untargeted ultra-performance liquid chromatography-mass spectrometry. As a complementary approach, we performed quantitative trait locus (QTL) mapping of near-isogenic introgression lines between C24 and Col-0 for specific seed specialized metabolites. Besides common QTL between seeds and leaves, GWAS revealed seed-specific QTL for specialized metabolites indicating differences in the genetic architecture of seeds and leaves. In seeds, aliphatic methylsulfinylalkyl and methylthioalkyl glucosinolates associated with the GS-ALK and GS-OHP locus on chromosome 4 containing alkenyl hydroxyalkyl producing 2 (AOP2) and 3 (AOP3) and/or with the GS-ELONG locus on chromosome 5 containing methylthioalkyl malate synthase (MAM1) and MAM3. We detected two unknown sulfur-containing compounds that were also mapped to these loci. In GWAS, some of the annotated flavonoids (kaempferol 3-O-rhamnoside-7-O-rhamnoside, quercetin 3-O-rhamnoside-7-O-rhamnoside) were mapped to transparent testa 7 (AT5G07990), encoding a cytochrome P450 75B1 monooxygenase. Three additional mass signals corresponding to quercetin-containing flavonols were mapped to UGT78D2 (AT5G17050). The association of the loci and associating metabolic features were functionally verified in knockdown mutant lines. By performing GWAS and QTL mapping, we were able to leverage variation of natural populations and parental lines to study seed specialized metabolism. The GWAS data set generated here is a high-quality resource that can be interrogated in further studies.

plant biology↗

bioRxiv2021

Tandem mass spectrum similarity-based network analysis using 13C-labeled and non-labeled metabolome data to identify the biosynthesis pathway of the blood pressure-lowering asparagus metabolite asparaptine A

Asparaptine, a conjugate of arginine and asparagusic acid, was found in asparagus (Asparagus officinalis) as a naturally occurring inhibitor of angiotensin-converting enzyme (ACE) in vitro. The biosynthetic pathway to asparaptine is largely unknown; however, it is suggested that asparagusic acid may be biosynthesized from valine. To determine which metabolites are involved in the asparaptine biosynthetic pathway, we performed tandem mass spectrometry similarity-based metabolome network analysis using 13C labeled and non-labeled valine-fed asparagus calluses. We determined that valine is used as a starting material, S(2-carboxy-n-propyl)-cysteine as an intermediate, and two new metabolites as asparaptine analogs, lysine- and histidine-type conjugates, are involved in the pathway. Asparaptine was therefore renamed asparaptine A (arginine type), and the two analogs were named asparaptines B (lysine type) and C (histidine type). Oral feeding of asparaptine A to a hypertensive mouse species showed that this metabolite lowers both blood pressure and heart rate within two hours and both of which were back to normal two days later. These results suggest that asparaptine A may not only have effects as an ACE inhibitor, but also has {beta}-antagonistic effects, which are well-known to be preventive for cardiovascular diseases.

plant biology↗

bioRxiv2021

Spatial metabolomics using imaging mass spectrometry to identify the localization of asparaptine in Asparagus officinalis

Spatial metabolomics uses imaging mass spectrometry (IMS) to localize metabolites within tissue section. Here, we performed matrix-assisted laser desorption/ionization-Fourier transform ion cyclotron resonance-IMS (MALDI-FTICR-IMS) to identify the localization of asparaptine, a naturally occurring inhibitor of angiotensin-converting enzyme, in green spears of asparagus (Asparagus officinalis). Spatial metabolome data were acquired with an untargeted manner. Segmentation analysis using the data characterized tissue-type-dependent and - independent distribution patterns in cross-sections of asparagus spears. Moreover, asparaptine accumulated at high levels in developing lateral shoot tissues. Quantification of asparaptine in lateral shoots using liquid chromatography-tandem mass spectrometry (LC-MS/MS) validated the IMS analysis. These results provide valuable information for understanding the function of asparaptine in asparagus, and identify the lateral shoot as a potential region of interest for multiomic studies to examine gene-to-metabolite associations in asparaptine biosynthesis.

plant biology↗

bioRxiv2021

Multi-metabolomics using imaging mass spectrometry and liquid chromatography-tandem mass spectrometry for spatially characterizing monoterpene indole alkaloids secreted from roots

Plants release specialized (secondary) metabolites from their roots to communicate with other organisms, including soil microorganisms. The spatial behavior of such metabolites around these roots can help us understand roles for the communication; however, currently they are unclear because soil-based studies are complex. Here, we established a multi-metabolomics approach using imaging mass spectrometry (IMS) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) to spatially assign metabolites under laboratory conditions using agar. In a case study using Catharanthus roseus, we showed that 58 nitrogen (N)-containing metabolites are released from the roots into the agar. For the metabolite assignment, we used 15N-labeled and nonlabeled LC-MS/MS data, previously reported. Four metabolite ions were identified using authentic standard compounds as derived from monoterpene indole alkaloids (MIAs) such as ajmalicine, catharanthine, serpentine, and yohimbine. An alkaloid network analysis using dot products and spinglass methods characterized five clusters to which the 58 ions belong. The analysis clustered ions from the indolic skeleton-type MIAs to a cluster, suggesting that other communities may represent distinct metabolite groups. For future chemical assignments of the serpentine community, key fragmentation patterns were characterized using the 15N-labeled and nonlabeled MS/MS spectra.

plant biology↗