The powerful potential of amino acid menthyl esters for anti-inflammatory and anti-obesity therapies.

Our newly developed menthyl esters of valine and isoleucine exhibit anti-inflammatory properties beyond those of the well-known menthol in macrophages stimulated by lipopolysaccharide (LPS) and in a mouse model of colitis induced by sodium dextran sulfate. Unlike menthol, which acts primarily through the cold-sensitive TRPM8 channel, these menthyl esters displayed unique mechanisms that operate independently of this receptor. They readily penetrated target cells and efficiently suppressed LPS-stimulated tumour necrosis factor-alpha (Tnf) expression mediated by liver X receptor (LXR), a key nuclear receptor that regulates intracellular cholesterol and lipid balance. The menthyl esters showed affinity for LXR and enhanced the transcriptional activity through their non-competitive and potentially synergistic agonistic effect. This effect can be attributed to the crucial involvement of SCD1, an enzyme regulated by LXR, which is central to lipid metabolism and plays a key role in the anti-inflammatory response. In addition, we discovered that the menthyl esters showed remarkable efficacy in suppressing adipogenesis in 3T3-L1 adipocytes at the mitotic clonal expansion stage in an LXR-independent manner as well as in mice subjected to diet-induced obesity. These multiple capabilities of our compounds establish them as formidable allies in the fight against inflammation and obesity, paving the way for a range of potential therapeutic applications.

Novel Potential of Rose Essential Oil as a Powerful Plant Defense Potentiator.

Terpenoids, natural compounds released by plants, function to enhance plant defense. The aim of this study was to investigate the effects of terpenoid-enriched essential oils (EOs) on tomato plants. From the application of a highly diluted solution of 11 different EOs to potted tomato soil, our study showed that rose essential oil (REO), rich in ß-citronellol, played a crucial role in activating defense genes in tomato leaves. As a result, leaf damage caused by herbivores, such as Spodoptera litura and Tetranychus urticae, was significantly reduced. In addition, our results were validated in field trials, providing evidence that REO is an effective biostimulant for enhancing plant defense against pests. Notably, the REO solution also had the added benefit of attracting herbivore predators, such as Phytoseiulus persimilis. Our findings suggest a practical approach to promote organic tomato production that encourages environmentally friendly and sustainable practices.

Long-term viable chimeric nephrons generated from progenitor cells are a reliable model in cisplatin-induced toxicity.

Kidney organoids have shown promise as evaluation tools, but their in vitro maturity remains limited. Transplantation into adult mice has aided in maturation; however, their lack of urinary tract connection limits long-term viability. Thus, long-term viable generated nephrons have not been demonstrated. In this study, we present an approachable method in which mouse and rat renal progenitor cells are injected into the developing kidneys of neonatal mice, resulting in the generation of chimeric nephrons integrated with the host urinary tracts. These chimeric nephrons exhibit similar maturation to the host nephrons, long-term viability with excretion and reabsorption functions, and cisplatin-induced renal injury in both acute and chronic phases, as confirmed by single-cell RNA-sequencing. Additionally, induced human nephron progenitor cells differentiate into nephrons within the neonatal kidneys. Collectively, neonatal injection represents a promising approach for in vivo nephron generation, with potential applications in kidney regeneration, drug screening, and pathological analysis.

Dialysis-related Amyloidosis Presenting as a Fever of Unknown Origin: Symptoms and Management.

A 74-year-old woman with a 34-year history of hemodialysis presented with an intermittent fever, which later coincided with recurrent bilateral shoulder and hip joint pain. Imaging studies suggested amyloid arthropathy, which was histologically confirmed by a synovial biopsy. Increasing ß2-microglobulin clearance during dialysis alone attenuated the intermittent fever and joint pain, but the symptoms did not disappear until the administration of prednisolone 10 mg/day. Reported cases of dialysis-related amyloidosis with a fever imply that changing to blood purification methods with high ß2-microglobulin clearance is crucial for controlling the condition long-term, whereas concurrent use of anti-inflammatory agents promptly alleviates the symptoms.

Evaluation of the ability of human induced nephron progenitor cells to form chimeric renal organoids using mouse embryonic renal progenitor cells.

The number of patients with end-stage renal failure is increasing annually worldwide and the problem is compounded by a shortage of renal transplantation donors. In our previous research, we have shown that transplantation of renal progenitor cells into the nephrogenic region of heterologous fetuses can induce the development of nephrons. We have also developed transgenic mice in which specific renal progenitor cells can be removed by drugs. By combining these two technologies, we have succeeded in generating human-mouse chimeric kidneys in fetal mice. We hope to apply these technologies to regenerative medicine. The quality of nephron progenitor cells (NPCs) derived from human pluripotent stem cells is important for the generation of chimeric kidneys, but there is currently no simple evaluation system for the chimerogenic potential of human NPCs. In this study, we focused on the fact that the re-aggregation of mouse renal progenitor cells can be used for nephron formation, even when merged into single cells. First, we examined the conditions under which nephron formation is likely to occur in mice during re-aggregation. Next, to improve the differentiation potential of human NPCs derived from pluripotent stem cells, NPCs were sorted using Integrin subunit alpha 8 (ITGA8). Finally, we demonstrated chimera formation between different species by mixing mouse cells with purified, selectively-induced human NPCs under optimum conditions. We observed these chimeric organoids at different time points to learn about these human-mouse chimeric structures at various stages of renal development. We found that the rate of chimera formation was affected by the purity of the human NPCs and the cell ratios used. We demonstrated that chimeric nephrons can be generated using a simple model, even between distant species. We believe that this admixture of human and mouse renal progenitor cells is a promising technology with potential application for the evaluation of the chimera formation abilities of NPCs.

Biosynthesis of dillapiole/apiole in dill (Anethum graveolens): characterization of regioselective phenylpropene O-methyltransferase.

The phenylpropene volatiles dillapiole and apiole impart one of the characteristic aromas of dill (Anethum graveolens) weeds. However, very few studies have been conducted to investigate the chemical composition of volatile compounds from different developmental stages and plant parts of A. graveolens. In this study, we examined the distribution of volatile phenylpropenes, including dillapiole, in dill plants at various developmental stages. We observed that young dill seedlings accumulate high levels of dillapiole and apiole, whereas a negligible proportion was found in the flowering plants and dry seeds. Based on transcriptomics and co-expression approaches with phenylpropene biosynthesis genes, we identified dill cDNA encoding S-adenosyl-L-methionine-dependent O-methyltransferase 1 (AgOMT1), an enzyme that can convert 6- and 2-hydroxymyristicin to dillapiole and apiole, respectively, via the methylation of the ortho-hydroxy group. The AgOMT1 protein shows an apparent Km value of 3.5 µm for 6-hydroxymyristicin and is 75% identical to the anise (Pimpinella anisum) O-methyltransferase (PaAIMT1) that can convert isoeugenol to methylisoeugenol via methylation of the hydroxy group at the para-position of the benzene ring. AgOMT1 showed a preference for 6-hydroxymyristicin, whereas PaAIMT1 displayed a large preference for isoeugenol. In vitro mutagenesis experiments demonstrated that substituting only a few residues can substantially affect the substrate specificity of these enzymes. Other plants belonging to the Apiaceae family contained homologous O-methyltransferase (OMT) proteins highly similar to AgOMT1, converting 6-hydroxymyristicin to dillapiole. Our results indicate that apiaceous phenylpropene OMTs with ortho-methylating activity evolved independently of phenylpropene OMTs of other plants and the enzymatic function of AgOMT1 and PaAIMT1 diverged recently.

Generation of functional chimeric kidney containing exogenous progenitor-derived stroma and nephron via a conditional empty niche.

Generation of new kidneys can be useful in various research fields, including organ transplantation. However, generating renal stroma, an important component tissue for structural support, endocrine function, and kidney development, remains difficult. Organ generation using an animal developmental niche can provide an appropriate in vivo environment for renal stroma differentiation. Here, we generate rat renal stroma with endocrine capacity by removing mouse stromal progenitor cells (SPCs) from the host developmental niche and transplanting rat SPCs. Furthermore, we develop a method to replace both nephron progenitor cells (NPCs) and SPCs, called the interspecies dual replacement of the progenitor (i-DROP) system, and successfully generate functional chimeric kidneys containing rat nephrons and stroma. This method can generate renal tissue from progenitors and reduce xenotransplant rejection. Moreover, it is a safe method, as donor cells do not stray into nontarget organs, thus accelerating research on stem cells, chimeras, and xenotransplantation.

Acquired aplastic anemia complicated with anti-glomerular basement membrane disease successfully treated with immunosuppressive therapy: a case report.

BACKGROUND: Aplastic anemia (AA) is a rare but fatal disorder characterized by pancytopenia due to bone marrow hypoplasia. Anti-glomerular basement membrane disease (anti-GBM disease) is an immune complex small-vessel vasculitis that presents as rapidly progressive glomerulonephritis and/or pulmonary hemorrhage. Although both involve autoreactive T cells that are partially triggered by human leukocyte antigen (HLA)-DR15, there have been no reports of their co-existence and the treatment strategy is not well understood. CASE PRESENTATION: A 67-year-old woman presented with fever, malaise, and acute kidney injury with proteinuria and hematuria requiring hemodialysis. She was diagnosed with anti-GBM antibody disease based on high serum anti-GBM antibody titer and crescentic glomerulonephritis on a renal biopsy. Pulse administration of methylprednisolone (MP), oral prednisolone (PSL), and plasmapheresis were performed. Only 2 weeks after the diagnosis of anti-GBM disease, the patient developed pancytopenia requiring frequent blood transfusions. The blood cell count did not recover even 1 month after discontinuing the drugs that could cause pancytopenia. Bone marrow examination showed hypocellularity without abnormal infiltrates or fibrosis, which led to the diagnosis of severe acquired AA. Further HLA phenotyping revealed that she had HLA-DR15. Increased dose of PSL with the secondary MP pulse and the addition of cyclosporine improved pancytopenia. Although she remained dialysis-dependent, anti-GBM disease and pancytopenia did not recur for more than 2 years. CONCLUSIONS: We report the first case of acquired AA complicated with anti-GBM disease in an elderly woman with HLA-DR15, which was successfully treated with immunosuppressive therapy (IST). This report is valuable not only because it shows they may co-occur, but also because it provides a therapeutic option for this complex condition. It was also suggested that pancytopenia in patients with anti-GBM disease recalls serious hematologic diseases including AA that require immediate treatment based on bone marrow examination.

Recurrent hemorrhagic shock from hemorrhagic cystitis due to neurogenic bladder.

INTRODUCTION: Hemorrhagic cystitis is characterized by gross hematuria, with hemorrhagic shock a rare complication. However, to our knowledge, its exact frequency has not been reported. CASE PRESENTATION: We report a case of an 86-year-old woman who showed repeated hemorrhagic cystitis with massive bleeding and hemorrhagic shock. The hemorrhagic cystitis was supposedly caused by the administration of aspirin and a neurogenic bladder. A urethral catheter was indwelled and hemorrhagic cystitis subsequently ceased. CONCLUSION: A review of patients with hemorrhagic cystitis at our hospital showed that only 3.3% experienced hemorrhagic shock. This case was even rarer because the patient experienced recurrent hemorrhagic shocks. A neurogenic bladder, which reduces the bladder's ability to function as a uroepithelial barrier against recurrent bacterial infections, caused the condition in this case. This report highlights how hemorrhagic cystitis can sometimes cause hemorrhagic shock.

Growth Enhancement of Arabidopsis (Arabidopsis thaliana) and Onion (Allium cepa) With Inoculation of Three Newly Identified Mineral-Solubilizing Fungi in the Genus Aspergillus Section Nigri.

Some soil fungi play an important role in supplying elements to plants by the solubilizing of insoluble minerals in the soil. The present study was conducted to isolate the mineral-solubilizing fungi from rhizosphere soil in some agricultural areas in northern Thailand. Seven fungal strains were obtained and identified using a polyphasic taxonomic approach with multilocus phylogenetic and phenotypic (morphology and extrolite profile) analyses. All obtained fungal strains were newly identified in the genus Aspergillus section Nigri, Aspergillus chiangmaiensis (SDBR-CMUI4 and SDBR-CMU15), Aspergillus pseudopiperis (SDBR-CMUI1 and SDBR-CMUI7), and Aspergillus pseudotubingensis (SDBR-CMUO2, SDBR-CMUO8, and SDBR-CMU20). All fungal strains were able to solubilize the insoluble mineral form of calcium, copper, cobalt, iron, manganese, magnesium, zinc, phosphorus, feldspar, and kaolin in the agar plate assay. Consequently, the highest phosphate solubilization strains (SDBR-CMUI1, SDBR-CMUI4, and SDBR-CMUO2) of each fungal species were selected for evaluation of their plant growth enhancement ability on Arabidopsis and onion in laboratory and greenhouse experiments, respectively. Plant disease symptoms were not found in any treatment of fungal inoculation and control. All selected fungal strains significantly increased the leaf number, leaf length, dried biomass of shoot and root, chlorophyll content, and cellular inorganic phosphate content in both Arabidopsis and onion plants under supplementation with insoluble mineral phosphate. Additionally, the inoculation of selected fungal strains also improved the yield and quercetin content of onion bulb. Thus, the selected strains reveal the potential in plant growth promotion agents that can be applied as a biofertilizer in the future.

Production of raspberry ketone by redirecting the metabolic flux to the phenylpropanoid pathway in tobacco plants.

Raspberry ketone is one of the characteristic flavors of raspberry fruits, and it is an important and expensive ingredient in the flavor and fragrance industries. It is present at low levels in plant tissues, and its occurrence is limited to a few taxa. In this context, the stable production of nature-identical raspberry ketone using heterologous synthesis in plants hosts has recently garnered the attention of plant biochemists. In this study, we demonstrate the rational switching of the metabolic flow from anthocyanin pigments to volatile phenylbutanoid production via the phenylpropanoid pathway. This shift led to the efficient and stable production of raspberry ketone and its glycosides via heterologous expression of the biosynthetic enzymes benzalacetone synthase (BAS) and raspberry ketone/zingerone synthase 1 (RZS1) in the transgenic tobacco (Nicotiana tabacum 'Petit Havana SR-1'). Additionally, we achieved improved product titers by activating the phenylpropanoid pathway with the transcriptional factor, production of anthocyanin pigment 1 (PAP1), from Arabidopsis thaliana. We further demonstrated another metabolic-flow switching by RNA interference (RNAi)-mediated silencing of chalcone synthase (CHS) to increase pathway-intermediate p-coumaroyl-CoA in transgenic tobacco for raspberry-ketone production. The redirection of metabolic flux resulted in transgenic lines producing 0.45 µg/g of raspberry ketone and 4.5 µg/g, on the fresh weight basis, of its glycosides in the flowers. These results suggest that the intracellular enforcement of endogenous substrate supply is an important factor while engineering the phenylpropanoid pathway. This strategy might be useful for the production of other phenylpropanoids/polyketides that are produced via the pathway-intermediate p-coumaroyl-CoA, in tobacco plants.

Renal hemosiderosis presenting with acute kidney Injury and macroscopic hematuria in Immunoglobulin A nephropathy: a case report.

BACKGROUND: Macroscopic hematuria-associated acute kidney injury (AKI) is a well-known complication of immunoglobulin A (IgA) nephropathy. In such cases, intratubular obstruction by red blood cell (RBC) casts and acute tubular necrosis are mainly observed pathologically. Herein, we report the case of a patient with IgA nephropathy presenting with AKI following an episode of macrohematuria. The patient presented with severe renal tubular hemosiderosis and acute tubular necrosis and without any obvious obstructive RBC casts. CASE PRESENTATION: A 68-year-old woman, who was diagnosed with IgA nephropathy on renal biopsy 6 years ago, was admitted to our hospital after an episode of macroscopic glomerular hematuria and AKI following upper respiratory tract infection. Renal biopsy showed mesangial proliferation of the glomeruli, including crescent formation in 17 % of the glomeruli, and acute tubular necrosis without obvious hemorrhage or obstructive RBC casts. The application of Perls' Prussian blue stain showed hemosiderin deposition in the renal proximal tubular cells. Immunofluorescence showed granular mesangial deposits of IgA and C3. Based on these findings, she was diagnosed with acute tubular necrosis with a concurrent IgA nephropathy flare-up. Moreover, direct tubular injury by heme and iron was considered to be the cause of AKI. She was treated with intravenous pulse methylprednisolone followed by oral prednisolone. Thereafter, the gross hematuria gradually faded, and her serum creatinine levels decreased. CONCLUSIONS: IgA nephropathy presenting with acute kidney injury accompanied by macrohematuria may cause renal hemosiderosis and acute tubular necrosis without obstructive RBC casts. Hemosiderosis may be a useful indicator for determining the pathophysiology of macroscopic hematuria-associated AKI. However, renal hemosiderosis may remain undiagnosed. Thus, Perls' Prussian blue iron staining should be more widely used in patients presenting with hematuria.

In vivo regeneration of neo-nephrons in rodents by renal progenitor cell transplantation.

Renal progenitor cells induced from pluripotent stem cells have attracted attention as a cell source for organ regeneration. Here, we report an in vivo protocol for the regeneration of urine-producing nephrons, i.e., neo-nephrons, in mice. We outline steps to transplant exogenous renal progenitor cells into the nephrogenic zone of transgenic mice and subsequently analyze these neo-nephrons. For complete details on the use and execution of this protocol, please refer to Fujimoto et al. (2020).

Fungal-Type Terpene Synthases in Marchantia polymorpha Are Involved in Sesquiterpene Biosynthesis in Oil Body Cells.

The liverwort Marchantia polymorpha possesses oil bodies in idioblastic oil body cells scattered in its thallus. Oil bodies are subcellular organelles in which specific sesquiterpenes and bisbibenzyls are accumulated. Therefore, a specialized system for the biosynthesis and accumulation of these defense compounds specifically in oil bodies has been implied. A recent study on M. polymorpha genome sequencing revealed 10 genes that shared high similarities with fungal-type terpene synthases (TPSs). Eight of these fungal-type TPS-like genes in M. polymorpha (MpFTPSL1-6, -9 and -10) are located within a 376-kb stretch on chromosome 6 and share similarities of over 94% at the nucleotide level. Therefore, these genes have likely originated from recent gene duplication events. The expression of a subset of MpFTPSLs was induced under non-axenic growth on vermiculite, which increased the amounts of sesquiterpenes and number of oil bodies. The tdTomato fluorescent protein-based in-fusion reporter assay with MpFTPSL2 promoter revealed fluorescent signals specifically in oil body cells of the thallus, indicating that MpFTPSL2 functions in oil body cells. Recombinant MpFTPSL2 expression in Escherichia coli led to sesquiterpene synthesis from farnesyl pyrophosphate. Moreover, suppression of a subset of MpFTPSLs through RNA interference reduced sesquiterpene accumulation in thalli grown on vermiculite. Taken together, these results suggest that at least a subset of MpFTPSLs is involved in sesquiterpene synthesis in oil body cells.

Study Profile of the Japan Multi-institutional Collaborative Cohort (J-MICC) Study.

BACKGROUND: The Japan Multi-institutional Collaborative Cohort (J-MICC) study was launched in 2005 to examine gene-environment interactions in lifestyle-related diseases, including cancers, among the Japanese. This report describes the study design and baseline profile of the study participants. METHODS: The participants of the J-MICC Study were individuals aged 35 to 69 years enrolled from respondents to study announcements in specified regions, inhabitants attending health checkup examinations provided by local governments, visitors at health checkup centers, and first-visit patients at a cancer hospital in Japan. At the time of the baseline survey, from 2005 to 2014, we obtained comprehensive information regarding demographics, education, alcohol consumption, smoking, sleeping, exercise, food intake frequency, medication and supplement use, personal and family disease history, psychological stress, and female reproductive history and collected peripheral blood samples. RESULTS: The baseline survey included 92,610 adults (mean age: 55.2 [standard deviation, 9.4] years, 44.1% men) from 14 study regions in 12 prefectures. The participation rate was 33.5%, with participation ranging from 19.7% to 69.8% in different study regions. The largest number of participants was in the age groups of 65-69 years for men and 60-64 years for women. There were differences in body mass index, educational attainment, alcohol consumption, smoking, and sleep duration between men and women. CONCLUSIONS: The J-MICC Study collected lifestyle and clinical data and biospecimens from over 90,000 participants. This cohort is expected to be a valuable resource for the national and international scientific community in providing evidence to support longer healthy lives.

Peritoneal dialysis-related peritonitis complicated with nonocclusive mesenteric ischemia.

Peritoneal dialysis (PD)-related peritonitis is a common complication of PD. Nonocclusive mesenteric ischemia (NOMI) is a rare complication of PD-related peritonitis, has a high mortality rate, and therefore should be detected early once it occurs. We describe a case of a 70-year-old woman on PD presented with moderate abdominal pain and low blood pressure, which contributed to the early diagnosis of PD-related peritonitis complicated with NOMI. Increased white cell count of 7150/µL (neutrophil, 84%) in dialysate effluent was diagnostic of PD-related peritonitis, which was later found to be caused by Pseudomonas putida. Computed tomography with contrast performed after administering crystalloids revealed hepatic portal venous gas, pneumatosis intestinalis in the ascending colon, and normal enhancement of the bowel wall and mesenteric arteries, which suggested a reperfusion of the previously ischemic ascending colon. Colonoscopy on hospital day seventeen revealed mucosal hemorrhage and ulcers in the entire right colon and the terminal ileum while the remaining colon was normal. These findings are compatible with the consequence of NOMI. Increased peak systolic velocity of the superior mesenteric artery (SMA) implied its stenosis. Past studies show that ischemia of the colon in patients with chronic kidney disease commonly occurs in the right colon. Arteriosclerosis of the SMA due to the long history of chronic kidney disease and diabetes might have caused its vulnerability to low blood pressure. Abdominal complications including NOMI should be screened for when a patient presents with low blood pressure and strong abdominal pain. This is the first case report that shows colonoscopy images of the colonic ulcers post-NOMI and PD-related peritonitis.

Techniques of orthotopic renal transplantation. II. Size-matched porcine grafts in monkey recipients

ABSTRACT Purpose As a classical xenotransplantation model, porcine kidneys have been transplanted into the lower abdomen of non-human primates. However, we have improved upon this model by using size-matched grafting in the orthotopic position. The beneficial aspects and surgical details of our method are reported herein. Methods Donors were two newborn pigs (weighting 5 to 6 kg) and recipients were two cynomolgus monkeys (weighting, approximately, 7 kg). After bilateral nephrectomy, kidneys were cold-transported in Euro-Collins solution. The porcine kidney was transplanted to the site of a left nephrectomy and fixed to the peritoneum. Results Kidneys transplanted to the lower abdomen by the conventional method were more susceptible to torsion of the renal vein (two cases). In contrast, early-stage blood flow insufficiency did not occur in orthotopic transplants of theleft kidney. Conclusions Size-matched porcine-primate renal grafting using our method of transplanting tothe natural position of the kidneys contributes to stable post-transplant blood flow to the kidney.

Techniques of fragile renal organoids transplantation in mice

Purpose: This study aimed to develop a microsurgical technique to transplant extremely fragile renal organoids in vivo, created by in-vitro reaggregation of metanephros from fetal mice. These organoids in reaggregation and development were examined histologically after transplantation under the renal capsule. Methods: Initially, metanephros from fetal mice were enzymatically treated to form single cells, and spheroids were generated in vitro. Under a microscope, the renal capsule was detached to avoid bleeding, and the outer cylinder of the indwelling needle was inserted to detach the renal parenchyma from the renal capsule using water pressure. The reaggregated spheroid was aspirated from the culture plate using a syringe with an indwelling needle outer cylinder and carefully extruded under the capsule. Pathological analysis was performed to evaluate changes in reaggregated spheroids over time and the effects of co-culture of spinal cord and subcapsular implantation on maturation. Results: In vitro, the formation of luminal structures became clearer on day 5. These fragile organoids were successfully implanted without tissue crapes under the renal capsule and formed glomerular. The effect of spinal cord co-transplant was not obvious histrionically. Conclusions: A simple and easy method to transplant fragile spheroids and renal under the renal capsule without damage was developed.

Biochemical characterization of the jasmonic acid methyltransferase gene from wasabi (Eutrema japonicum).

Methyl jasmonate and jasmonic acid play important roles as signaling molecules in regulating plant development and stress-related responses. Previous studies have shown that jasmonic acid carboxyl methyltransferase (JMT), which belongs to the SABATH methyltransferase gene family, catalyzes the transfer of methyl groups from S-adenosyl-L-methionine to the carboxyl groups of jasmonic acid. In the present study, we used RNA-seq analysis to identify a putative JMT gene, EujJMT, in wasabi (Eutrema japonicum). The EujJMT proteins showed the highest similarity (89% identity) to JMT proteins of Brassica rapa. Functional characterization of a recombinant EujJMT protein expressed in Escherichia coli showed the highest level of activity with jasmonic acid among the different carboxylic acids tested. The apparent Km value of EujJMT using jasmonic acid as substrate was 62.6 µM, which is comparable to the values of known JMTs. Phylogenetic analysis suggested that EujJMT shares a common ancestor with the JMTs of Arabidopsis and Brassica species and that the strict substrate specificity toward jasmonic acid is conserved among Brassicaceae JMTs.

Molecular cloning and functional characterization of an O-methyltransferase catalyzing 4'-O-methylation of resveratrol in Acorus calamus.

Resveratrol and its methyl ethers, which belong to a class of natural polyphenol stilbenes, play important roles as biologically active compounds in plant defense as well as in human health. Although the biosynthetic pathway of resveratrol has been fully elucidated, the characterization of resveratrol-specific O-methyltransferases remains elusive. In this study, we used RNA-seq analysis to identify a putative aromatic O-methyltransferase gene, AcOMT1, in Acorus calamus. Recombinant AcOMT1 expressed in Escherichia coli showed high 4'-O-methylation activity toward resveratrol and its derivative, isorhapontigenin. We purified a reaction product enzymatically formed from resveratrol by AcOMT1 and confirmed it as 4'-O-methylresveratrol (deoxyrhapontigenin). Resveratrol and isorhapontigenin were the most preferred substrates with apparent Km values of 1.8 µM and 4.2 µM, respectively. Recombinant AcOMT1 exhibited reduced activity toward other resveratrol derivatives, piceatannol, oxyresveratrol, and pinostilbene. In contrast, recombinant AcOMT1 exhibited no activity toward pterostilbene or pinosylvin. These results indicate that AcOMT1 showed high 4'-O-methylation activity toward stilbenes with non-methylated phloroglucinol rings.