Comparative global profiling of Perilla leaf and stem via transcriptomics and metabolomics.

Perilla (Perilla frutescens L.) is a time-honored herbal plant with widespread applications in both medicine and culinary practices around the world. Profiling the essential organs and tissues with medicinal significance on a global scale offers valuable insights for enhancing the yield of desirable compounds in Perilla and other medicinal plants. In the present study, genome-wide RNA-sequencing (RNA-seq) and assessing the global spectrum of metabolites were carried out in the two major organs/tissues of stem (PfST) and leaf (PfLE) in Perilla. The results showed a total of 18,490 transcripts as the DEGs (differentially expressed genes) and 144 metabolites as the DAMs (differentially accumulated metabolites) through the comparative profiling of PfST vs PfLE, and all the DEGs and DAMs exhibited tissue-specific trends. An association analysis between the transcriptomics and metabolomics revealed 14 significantly enriched pathways for both DEGs and DAMs, among which the pathways of Glycine, serine and threonine metabolism (ko00260), Glyoxylate and dicarboxylate metabolism (ko00630), and Glucagon signaling pathway (ko04922) involved relatively more DEGs and DAMs. The results of qRT-PCR assays of 18 selected DEGs confirmed the distinct tissue-specific characteristics of all identified DEGs between PfST and PfLE. Notably, all eight genes associated with the flavonoid biosynthesis/metabolism pathways exhibited significantly elevated expression levels in PfLE compared to PfST. This observation suggests a heightened accumulation of metabolites related to flavonoids in Perilla leaves. The findings of this study offer a comprehensive overview of the organs and tissues in Perilla that have medicinal significance.

Integrated analysis of miRNA, transcriptome, and degradome sequencing provides new insights into lipid metabolism in perilla seed.

MicroRNAs (miRNA) are small noncoding RNAs that play a crucial as molecular regulators in lipid metabolism in various oil crops. Perilla (Perilla frutescens) is a specific oil crop known for its high alpha-linolenic acid (C18:3n3, ALA) content (>65 %) in their seed oils. In view of the regulatory mechanism of miRNAs in perilla remains unclear, we conducted miRNAs and transcriptome sequencing in two cultivars with distinct lipid compositions. A total of 525 unique miRNAs, including 142 differentially expressed miRNAs was identified in perilla seeds. The 318 miRNAs targeted 7,761 genes. Furthermore, we identified 112 regulated miRNAs and their 610 target genes involved in lipid metabolism. MiR159b and miR167a as the core nodes to regulate the expression of genes in oil biosynthesis (e.g., KAS, FATB, GPAT, FAD, DGK, LPAAT) and key regulatory TFs (e.g., MYB, ARF, DOF, SPL, NAC, TCP, and bHLH). The 1,219 miRNA-mRNA regulation modules were confirmed through degradome sequencing. Notably, pf-miR159b-MYBs and pf-miR167a-ARFs regulation modules were confirmed. They exhibited significantly different expression levels in two cultivars and believed to play important roles in oil biosynthesis in perilla seeds. This provides valuable insights into the functional analysis of miRNA-regulated lipid metabolism in perilla seeds.

Assessing the Potential for Genome-Assisted Breeding in Red Perilla Using Quantitative Trait Locus Analysis and Genomic Prediction.

Red perilla is an important medicinal plant used in Kampo medicine. The development of elite varieties of this species is urgently required. Medicinal compounds are generally considered target traits in medicinal plant breeding; however, selection based on compound phenotypes (i.e., conventional selection) is expensive and time consuming. Here, we propose genomic selection (GS) and marker-assisted selection (MAS), which use marker information for selection, as suitable selection methods for medicinal plants, and we evaluate the effectiveness of these methods in perilla breeding. Three breeding populations generated from crosses between one red and three green perilla genotypes were used to elucidate the genetic mechanisms underlying the production of major medicinal compounds using quantitative trait locus analysis and evaluating the accuracy of genomic prediction (GP). We found that GP had a sufficiently high accuracy for all traits, confirming that GS is an effective method for perilla breeding. Moreover, the three populations showed varying degrees of segregation, suggesting that using these populations in breeding may simultaneously enhance multiple target traits. This study contributes to research on the genetic mechanisms of the major medicinal compounds of red perilla, as well as the breeding efficiency of this medicinal plant.

The Genome of the Korean Island-Originated Perilla citriodora 'Jeju17' Sheds Light on Its Environmental Adaptation and Fatty Acid and Lipid Production Pathways.

Perilla is a key component of Korean food. It contains several plant-specialized metabolites that provide medical benefits. In response to an increased interest in healthy supplement food from the public, people are focusing on the properties of Perilla. Nevertheless, unlike rice and soybeans, there are few studies based on molecular genetics on Perilla, so it is difficult to systematically study the molecular breed. The wild Perilla, Perilla citriodora 'Jeju17', was identified a decade ago on the Korean island of Jeju. Using short-reads, long-reads, and Hi-C, a chromosome-scale genome spanning 676 Mbp, with high contiguity, was assembled. Aligning the 'Jeju17' genome to the 'PC002' Chinese species revealed significant collinearity with respect to the total length. A total of 31,769 coding sequences were predicted, among which 3331 were 'Jeju17'-specific. Gene enrichment of the species-specific gene repertoire highlighted environment adaptation, fatty acid metabolism, and plant-specialized metabolite biosynthesis. Using a homology-based approach, genes involved in fatty acid and lipid triacylglycerol biosynthesis were identified. A total of 22 fatty acid desaturases were found and comprehensively characterized. Expression of the FAD genes in 'Jeju17' was examined at the seed level, and hormone signaling factors were identified. The results showed that the expression of FAD genes in 'Jeju17' at the seed level was high 25 days after flowering, and their responses of hormones and stress were mainly associated with hormone signal transduction and abiotic stress via cis-elements patterns. This study presents a chromosome-level genome assembly of P. citriodora 'Jeju17', the first wild Perilla to be sequenced from the Korean island of Jeju. The analyses provided can be useful in designing ALA-enhanced Perilla genotypes in the future.

Genome-wide comprehensive characterization and transcriptomic analysis of AP2/ERF gene family revealed its role in seed oil and ALA formation in perilla (Perilla frutescens).

Perilla (Perilla frutescens) is a potential specific oilseed crop with an extremely high α-linolenic acid (ALA) content in its seeds. AP2/ERF transcription factors (TFs) play important roles in multiple biological processes. However, limited information is known about the regulatory mechanism of the AP2/ERF family in perilla's oil accumulation. In this research, we identified 212 AP2/ERF family members in the genome of perilla, and their domain characteristics, collinearity, and sub-genome differentiation were comprehensively analyzed. Transcriptome sequencing revealed that genes encoding key enzymes involved in oil biosynthesis (e.g., ACCs, KASII, GPAT, PDAT and LPAAT) were up-regulated in the high-oil variety. Moreover, the endoplasmic reticulum-localized FAD2 and FAD3 were significantly up-regulated in the high-ALA variety. To investigate the roles of AP2/ERFs in lipid biosynthesis, we conducted a correlation analysis between non-redundant AP2/ERFs and key lipid metabolism genes using WGCNA. A significant correlation was found between 36 AP2/ERFs and 90 lipid metabolism genes. Among them, 12 AP2/ERFs were identified as hub genes and showed significant correlation with lipid synthase genes (e.g., FADs, GPAT and ACSL) and key regulatory TFs (e.g., LEC2, IAA, MYB, UPL3). Furthermore, gene expression analysis identified three AP2/ERFs (WRI, ABI4, and RAVI) potentially playing an important role in the regulation of oil accumulation in perilla. Our study suggests that PfAP2/ERFs are important regulatory TFs in the lipid biosynthesis pathway, providing a foundation for the molecular understanding of oil accumulation in perilla and other oilseed crops.

Development of an SNP marker set for marker-assisted backcrossing using genotyping-by-sequencing in tetraploid perilla.

High-quality molecular markers are essential for marker-assisted selection to accelerate breeding progress. Compared with diploid species, recently diverged polyploid crop species tend to have highly similar homeologous subgenomes, which is expected to limit the development of broadly applicable locus-specific single-nucleotide polymorphism (SNP) assays. Furthermore, it is particularly challenging to make genome-wide marker sets for species that lack a reference genome. Here, we report the development of a genome-wide set of kompetitive allele specific PCR (KASP) markers for marker-assisted recurrent selection (MARS) in the tetraploid minor crop perilla. To find locus-specific SNP markers across the perilla genome, we used genotyping-by-sequencing (GBS) to construct linkage maps of two F2 populations. The two resulting high-resolution linkage maps comprised 2326 and 2454 SNP markers that spanned a total genetic distance of 2133 cM across 16 linkage groups and 2169 cM across 21 linkage groups, respectively. We then obtained a final genetic map consisting of 22 linkage groups with 1123 common markers from the two genetic maps. We selected 96 genome-wide markers for MARS and confirmed the accuracy of markers in the two F2 populations using a high-throughput Fluidigm system. We confirmed that 91.8% of the SNP genotyping results from the Fluidigm assay were the same as the results obtained through GBS. These results provide a foundation for marker-assisted backcrossing and the development of new varieties of perilla.

Overexpression of the Purple Perilla (Perilla frutescens (L.)) FAD3a Gene Enhances Salt Tolerance in Soybean.

The increasingly serious trend of soil salinization inhibits the normal growth and development of soybeans, leading to reduced yields and a serious threat to global crop production. Microsomal ω-3 fatty acid desaturase encoded by the FAD3 gene is a plant enzyme that plays a significant role in α-linolenic acid synthesis via regulating the membrane fluidity to better accommodate various abiotic stresses. In this study, PfFAD3a was isolated from perilla and overexpressed in soybeans driven by CaMV P35S, and the salt tolerance of transgenic plants was then evaluated. The results showed that overexpression of PfFAD3a increased the expression of PfFAD3a in both the leaves and seeds of transgenic soybean plants, and α-linolenic acid content also significantly increased; hence, it was shown to significantly enhance the salt tolerance of transgenic plants. Physiological and biochemical analysis showed that overexpression of PfFAD3a increased the relative chlorophyll content and PSII maximum photochemical efficiency of transgenic soybean plants under salt stress; meanwhile, a decreased accumulation of MDA, H2O2, and O2•-, increased the activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbic acid peroxidase (APX), as well as the production of proline and soluble sugar. In summary, the overexpression of PfFAD3a may enhance the salt tolerance in transgenic soybean plants through enhanced membrane fluidity and through the antioxidant capacity induced by C18:3.

The revealing of a novel double bond reductase related to perilla ketone biosynthesis in Perilla frutescens.

BACKGROUND: Perilla frutescens is widely used as both a medicine and a food worldwide. Its volatile oils are its active ingredients, and, based on the different volatile constituents, P. frutescens can be divided into several chemotypes, with perilla ketone (PK) being the most common. However, the key genes involved in PK biosynthesis have not yet been identified. RESULTS: In this study, metabolite constituents and transcriptomic data were compared in leaves of different levels. The variation in PK levels was the opposite of that of isoegoma ketone and egoma ketone in leaves at different levels. Based on transcriptome data, eight candidate genes were identified and successfully expressed in a prokaryotic system. Sequence analysis revealed them to be double bond reductases (PfDBRs), which are members of the NADPH-dependent, medium-chain dehydrogenase/reductase (MDR) superfamily. They catalyze the conversion of isoegoma ketone and egoma ketone into PK in in vitro enzymatic assays. PfDBRs also showed activity on pulegone, 3-nonen-2-one, and 4-hydroxybenzalacetone. In addition, several genes and transcription factors were predicted to be associated with monoterpenoid biosynthesis, and their expression profiles were positively correlated with variations in PK abundance, suggesting their potential functions in PK biosynthesis. CONCLUSIONS: The eight candidate genes encoding a novel double bond reductase related to perilla ketone biosynthesis were identified in P. frutescens, which carries similar sequences and molecular features as the MpPR and NtPR from Nepeta tenuifolia and Mentha piperita, respectively. These findings not only reveal the pivotal roles of PfDBR in exploring and interpreting PK biological pathway but also contribute to facilitating future studies on this DBR protein family.

A highly contiguous genome assembly of red perilla (Perilla frutescens) domesticated in Japan.

Perilla frutescens (Lamiaceae) is an important herbal plant with hundreds of bioactive chemicals, among which perillaldehyde and rosmarinic acid are the two major bioactive compounds in the plant. The leaves of red perilla are used as traditional Kampo medicine or food ingredients. However, the medicinal and nutritional uses of this plant could be improved by enhancing the production of valuable metabolites through the manipulation of key enzymes or regulatory genes using genome editing technology. Here, we generated a high-quality genome assembly of red perilla domesticated in Japan. A near-complete chromosome-level assembly of P. frutescens was generated contigs with N50 of 41.5 Mb from PacBio HiFi reads. 99.2% of the assembly was anchored into 20 pseudochromosomes, among which seven pseudochromosomes consisted of one contig, while the rest consisted of less than six contigs. Gene annotation and prediction of the sequences successfully predicted 86,258 gene models, including 76,825 protein-coding genes. Further analysis showed that potential targets of genome editing for the engineering of anthocyanin pathways in P. frutescens are located on the late-stage pathways. Overall, our genome assembly could serve as a valuable reference for selecting target genes for genome editing of P. frutescens.

Genetic variation of seed oil characteristics in native Korean germplasm of Perilla crop (Perilla frutescens L.) using SSR markers.

BACKGROUND: In order to maximize the use of valuable native Perilla germplasm in South Korea, knowledge of the Perilla seed oil content and genetic variation among native Perilla germplasm resources is very important for the conservation and development of new Perilla seed oil varieties using the native Perilla germplasm accessions preserved from the Rural Development Administration Genebank (RDA-Genebank) collection from South Korea. OBJECTIVES: In this study, we studied population structure and association mapping to identify Perilla SSR markers (PSMs) associated with the five fatty acid contents and two seed characteristics of the native Korean Perilla germplasm accessions of cultivated var. frutescens of the RDA-Genebank collected in South Korea. METHODS: For an association mapping analysis to find PSMs associated with the five fatty acid contents and two seed characteristics of the Perilla germplasm accessions of cultivated var. frutescens, we evaluated the content of five fatty acids of 280 native Korean Perilla germplasm accessions and used 29 Perilla SSR primer sets to measure the genetic diversity and relationships, population structure, and association mapping of the native Korean Perilla germplasm accessions of the RDA-Genebank collected in South Korea. RESULTS: Five fatty acids of 280 native Korean Perilla accessions were identified as follows: palmitic acid (PA) (5.30-8.66%), stearic acid (SA) (1.60-4.19%), oleic acid (OA) (9.60-22.5%), linoleic acid (LA) (8.38-25.4%), and linolenic acid (LNA) (52.7-76.4%). In a correlation analysis among the five fatty acids and two seed characteristics of the 280 Perilla accessions, the combinations of PA and SA (0.794**) and SA and OA (0.724**) showed a particularly high positive correlation coefficients compare to other combinations. By using an association analysis of the 29 PSMs and the five fatty acids in the 280 Perilla accessions, we found 17 PSMs (KNUPF1, KNUPF2, KNUPF4, KNUPF10, KNUPF16, KNUPF25, KNUPF26, KNUPF28, KNUPF37, KNUPF55, KNUPF62, KNUPF71, KNUPF74, KNUPF77, KNUPF85, KNUPF89, and KNUPF118) associated with the content of the five fatty acid components and two seed characteristics. CONCLUSIONS: These PSMs are considered to be useful molecular markers related to five fatty acid components and two seed characteristics for selecting accessions from the germplasm accessions of the Perilla crop and their related weedy types through association mapping analysis and marker-assisted selection (MAS) breeding programs.

Incipient diploidization of the medicinal plant Perilla within 10,000 years.

Perilla is a young allotetraploid Lamiaceae species widely used in East Asia as herb and oil plant. Here, we report the high-quality, chromosome-scale genomes of the tetraploid (Perilla frutescens) and the AA diploid progenitor (Perilla citriodora). Comparative analyses suggest post Neolithic allotetraploidization within 10,000 years, and nucleotide mutation in tetraploid is 10% more than in diploid, both of which are dominated by G:C → A:T transitions. Incipient diploidization is characterized by balanced swaps of homeologous segments, and subsequent homeologous exchanges are enriched towards telomeres, with excess of replacements of AA genes by fractionated BB homeologs. Population analyses suggest that the crispa lines are close to the nascent tetraploid, and involvement of acyl-CoA: lysophosphatidylcholine acyltransferase gene for high α-linolenic acid content of seed oil is revealed by GWAS. These resources and findings provide insights into incipient diploidization and basis for breeding improvement of this medicinal plant.

Genetic diversity and population structure of Perilla frutescens collected from Korea and China based on simple sequence repeats (SSRs).

BACKGROUND: Perilla frutescens (L.) Britt. is divided into two varieties based on morphology and use. One is P. frutescens var. frutescens, which is used both as a leafy vegetable and as an oil obtained from the seeds. The other variety is P. frutescens var. crispa, a Chinese medicine or spice vegetable crop. In addition, weedy types of var. frutescens and var. crispa are occasionally grown as relict forms and are easy to find on roadsides, in waste areas and around farmers' fields or farmhouses. SSR markers have been successfully used to examine the genetic diversity and relationships of cultivated and weedy types of Perilla in many regions. OBJECTIVES: In this study, we used 25 simple sequence repeat (SSR) markers were used to assess the genetic diversity and population structure of 90 Perilla accessions from Korea and China. METHODS: A total of 90 accessions of Perilla were collected in Korea and China included 45 accessions from each of Korea and China. We selected 25 SSR markers representing the polymorphism of and adequately amplifying all the Perilla accessions. RESULTS: A total of 153 alleles were identified, with an average of 6.12 alleles per locus. The GD level and PIC value for cultivated and weedy types of P. frutescens from China were higher than those for accessions from Korea. Weedy accessions had higher GD and PIC values than cultivated accessions. In the population structure analysis using the model-based method, the 90 Perilla accessions were divided into two main group and an admixed group based on a membership probability threshold of 0.8. Based on the distance-based unweighted pair group method with the arithmetic mean (UPGMA), all accessions were classified into four major groups with a genetic similarity of 32.8%. CONCLUSION: Finally, the findings of this study will provide useful theoretical knowledge for further study of the population structure and genetic diversity of Perilla species and benefit Perilla crop breeding and germplasm conservation in Korea and China.

Identification of quantitative trait loci associated with flowering time in perilla using genotyping-by-sequencing.

Understanding the transition to the reproductive period is important for crop breeding. This information can facilitate the production of novel varieties that are better adapted to local environments or changing climatic conditions. Here, we report the development of a high-density linkage map based on genotyping-by-sequencing (GBS) for the genus perilla. Through GBS library construction and Illumina sequencing of an F2 population, a total of 9607 single-nucleotide polymorphism (SNP) markers were developed. The ten-group linkage map of 1309.39 cM contained 2518 markers, with an average marker density of 0.56 cM per linkage group (LG). Using this map, a total of six QTLs were identified. These quantitative trait loci (QTLs) are associated with three traits related to flowering time: days to visible flower bud, days to flowering, and days to maturity. Ortholog analysis conducted with known genes involved in the regulation of flowering time among different crop species identified GI, CO and ELF4 as putative perilla orthologs that are closely linked to the QTL regions associated with flowering time. These results provide a foundation that will be useful for future studies of flowering time in perilla using fine mapping, and marker-assisted selection for the development of new varieties of perilla.

Genetic diversity and population structure analysis in Perilla crop and their weedy types from northern and southern areas of China based on simple sequence repeat (SSRs).

INTRODUCTION: Identification of genetic variation is an essential ability for the long-term success of breeding programs and maximizes the use of germplasm resources. In East Asia, China has a long history of the cultivation of Perilla crop, but there has been little research on the genetic diversity and genetic relationships among accessions of Perilla crop and their weedy types. OBJECTIVES: To better understand the genetic variations of the cultivated and weedy types of Perilla crop in China, the 91 accessions were evaluated for genetic diversity by 21 simple sequence repeat (SSR) markers. METHODS: SSR amplifications were conducted in a total volume of 20 µL, consisting of 20 ng genomic DNA, 1X PCR buffer, 0.5 µM forward and reverse primers, 0.2 mM dNTPs, and 1 U Taq polymerase. Power Marker version 3.25 was applied to obtain the information on the number of alleles, allele frequency, major allele frequency, gene diversity (GD), and polymorphic information content (PIC). The similarity matrix was used to construct an unweighted pair group method with arithmetic mean dendrogram by the application of SAHN-Clustering from NTSYS-pc.V.2.1. RESULTS: A total of 147 alleles were identified with an average of 7 alleles per locus. The average values of PIC and GD were 0.577 and 0.537, respectively. The genetic diversity level of accessions from Northern China was lower than accessions from Southern China. The genetic diversity level and PIC values for accessions of var. crispa were the highest. For accessions of cultivated var. frutescens, genetic diversity in Southern China was higher than that in Northern China. CONCLUSION: Most cultivated Perilla accessions were clearly separated from weedy Perilla accessions, but there was no clear geographic structure between cultivated Perilla crop and weedy types based on their regional distribution. This study demonstrated the utility of SSR analysis for performing genetic and population analysis of cultivated and weedy types of Perilla accessions in China.

RNA Sequencing and Coexpression Analysis Reveal Key Genes Involved in α-Linolenic Acid Biosynthesis in Perilla frutescens Seed.

Perillafrutescen is used as traditional food and medicine in East Asia. Its seeds contain high levels of α-linolenic acid (ALA), which is important for health, but is scarce in our daily meals. Previous reports on RNA-seq of perilla seed had identified fatty acid (FA) and triacylglycerol (TAG) synthesis genes, but the underlying mechanism of ALA biosynthesis and its regulation still need to be further explored. So we conducted Illumina RNA-sequencing in seven temporal developmental stages of perilla seeds. Sequencing generated a total of 127 million clean reads, containing 15.88 Gb of valid data. The de novo assembly of sequence reads yielded 64,156 unigenes with an average length of 777 bp. A total of 39,760 unigenes were annotated and 11,693 unigenes were found to be differentially expressed in all samples. According to Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, 486 unigenes were annotated in the "lipid metabolism" pathway. Of these, 150 unigenes were found to be involved in fatty acid (FA) biosynthesis and triacylglycerol (TAG) assembly in perilla seeds. A coexpression analysis showed that a total of 104 genes were highly coexpressed (r > 0.95). The coexpression network could be divided into two main subnetworks showing over expression in the medium or earlier and late phases, respectively. In order to identify the putative regulatory genes, a transcription factor (TF) analysis was performed. This led to the identification of 45 gene families, mainly including the AP2-EREBP, bHLH, MYB, and NAC families, etc. After coexpression analysis of TFs with highly expression of FAD2 and FAD3 genes, 162 TFs were found to be significantly associated with two FAD genes (r > 0.95). Those TFs were predicted to be the key regulatory factors in ALA biosynthesis in perilla seed. The qRT-PCR analysis also verified the relevance of expression pattern between two FAD genes and partial candidate TFs. Although it has been reported that some TFs are involved in seed development, more direct evidence is still needed to verify their function. However, these findings can provide clues to reveal the possible molecular mechanisms of ALA biosynthesis and its regulation in perilla seed.

The complete chloroplast genome sequence of Perilla citriodora (Makino) Nakai.

Perilla citriodora is a diploid and an aromatic herb species belonging to the family Lamiaceae and widely distributed in East Asia. The complete chloroplast genome sequence of P. citriodora was generated by de novo assembly using whole genome next-generation sequences. The assembled chloroplast genome of P. citriodora was 152 588 bp in length and structurally divided into four distinct regions; large single copy region (83 699 bp), small single copy region (17 537 bp), and a pair of inverted repeat regions (25 676 bp). A total of 115 genes were annotated including 81 protein-coding genes, 30 tRNA genes, and four rRNA genes. Phylogenetic relationship with previously reported chloroplast genomes revealed that P. citriodora is most closely related to P. frutescens, a tetraploid Perilla species.

The complete chloroplast genome sequence of Perilla frutescens (L.).

Perilla frutescens (L.) is a traditionally medical herb of East Asia. The complete chloroplast genome of P. frutescens (L.) Britton var. frutescens was assembled in this study. Total chloroplast genome size of Perilla was 153,666 bp in length, containing a pair of inverted repeats (IRs) of 25,677 bp, separated by large single copy (LSC) and small single copy (SSC) of 84,288 bp and 17,925 bp, respectively. Overall GC contents of the genome were 37.6%. The chloroplast genome harbored 127 annotated genes, including 89 protein-coding genes, 29 tRNA genes and 8 rRNA genes. Eleven genes contained one or two introns.

Two types of alcohol dehydrogenase from Perilla can form citral and perillaldehyde.

Studies on the biosynthesis of oil compounds in Perilla will help in understanding regulatory systems of secondary metabolites and in elucidating reaction mechanisms for natural product synthesis. In this study, two types of alcohol dehydrogenases, an aldo-keto reductase (AKR) and a geraniol dehydrogenase (GeDH), which are thought to participate in the biosynthesis of perilla essential oil components, such as citral and perillaldehyde, were isolated from three pure lines of perilla. These enzymes shared high amino acid sequence identity within the genus Perilla, and were expressed regardless of oil type. The overall reaction from geranyl diphosphate to citral was performed in vitro using geraniol synthase and GeDH to form a large proportion of citral and relatively little geraniol as reaction products. The biosynthetic pathway from geranyl diphosphate to citral, the main compound of citral-type perilla essential oil, was established in this study.

Effects of SYJN, a Chinese herbal formula, on chronic unpredictable stress-induced changes in behavior and brain BDNF in rats.

AIM OF THE STUDY: Suyu-Jiaonang (SYJN) is a Chinese herbal formula that contains four herbs: Bupleurum chinense DC, Curcuma aromatica Salisb., Perilla frutescens (Linn.) Britt., and Acorus tatarinowii Schott. Previous studies conducted in our laboratory have revealed an antidepressant-like effect of the formula in various mouse models of behavioral despair. The present study aimed to investigate whether SYJN could produce antidepressant-like effects in chronic unpredictable stress (CUS)-induced depression model in rats and its possible mechanism(s). MATERIALS AND METHODS: Rats were subjected to an experimental setting of CUS. The effect of SYJN treatment on CUS-induced depression was examined using behavioral tests including the sucrose consumption and open field tests. The mechanism underlying the antidepressant-like action of SYJN was examined by measuring brain-derived neurotrophic factor (BDNF) protein and mRNA expression in brain tissues of CUS-exposed rats. RESULTS: Exposure to CUS for 4 weeks caused depression-like behavior in rats, as indicated by significant decreases in sucrose consumption and locomotor activity (assessed in the open field test). In addition, it was found that BDNF protein and mRNA levels in the hippocampus and frontal cortex were lower in CUS-treated rats, as compared to controls. Daily intragastric administration of SYJN (1300 or 2600 mg/kg) during the 4-week period of CUS significantly suppressed behavioral changes and attenuated the CUS-induced decrease in BDNF protein and mRNA levels in the hippocampus and frontal cortex. CONCLUSION: The results suggest that SYJN alleviates depression induced by CUS. The antidepressant-like activity of SYJN is likely mediated by the increase in BDNF expression in brain tissues.

Determination of toxic perilla ketone, secondary plant metabolites and antioxidative capacity in five Perilla frutescens L. varieties.

Perilla frutescens L. is a traditional Asian crop with multiple uses. Several varieties exist but only little data is available about the content of the toxic perilla ketone and secondary plant metabolites of those genotypes. To estimate the nutritional value of this new vegetable more information about those components is necessary. We investigated five genotypes of P. frutescens L. to determine their content of PK, phenolics, carotenoids and AC. AC was examined using ABTS-decolorization assay and lipid peroxidation assay. Carotenoids were identified and quantified by HPLC analysis, phenolics were quantified by means of Folin-Ciocalteu and PK was identified by GC/MS. Two genotypes were found to contain PK, a potent lung toxin, and are therefore not suitable for consumption. The phenolic content and corresponding antioxidative capacity of all genotypes is considerably high compared to other vegetables. All genotypes moreover contain notably high concentrations of carotenoids with contents up to fivefold higher than in other carotenoid rich vegetables. The results indicate that there are several genotypes which are not suitable for consumption due to their content of PK. However PK free genotypes are rich sources of natural antioxidants, and may therefore be considered as a novel vegetable with health promoting properties.