The pungent-variable sweet chili pepper 'Shishito' (Capsicum annuum) provides insights regarding the relationship between pungency, the number of seeds, and gene expression involving capsaicinoid biosynthesis.

Pungent traits caused by capsaicinoids are characteristic of chili peppers (Capsicum spp.), and the pungent-variable sweet chili pepper 'Shishito' (Capsicum annuum) is unique in being known for the pungency in fruits with few seeds. In the present study, we tried to clarify the relationship between the number of seeds and pungency in 'Shishito'. First, we investigated the pungency of 'Shishito' by simple sensory evaluations and quantifications of capsaicinoids by high-performance liquid chromatography. As a result, few-seeded fruits had a larger fluctuation of capsaicinoid content than many-seeded ones. This indicates that the number of seeds, in particular a decrease of the seeds, has some sort of connection with the pungency of 'Shishito'. Then, we analyzed the relationship between pungency and gene expression involving capsaicinoid biosynthesis at the individual fruit level. We vertically separated the placental septum in which capsaicinoids are synthesized and performed quantitative reverse transcription-polymerase chain reaction (qRT-PCR) for 18 genes involved in capsaicinoid biosynthesis. We also used the placental septum for capsaicinoid quantification so that the gene expression levels and capsaicinoid contents in the same fruits were obtained, and their correlations were analyzed using 20 biological replicates. Among the 18 genes, expression levels of 11 genes (WRKY9, CaMYB31, AT3, BCAT, BCKDH, KAS I, KAS III, ACL, CaKR1, FAT, and pAMT) had a significant positive correlation with the capsaicinoid concentration, and they were considered to upregulate capsaicinoid biosynthesis. These results provide new insights regarding the environmental variation of the pungency traits in chili peppers and the relationship between pungency, the number of seeds, and gene expression involved in capsaicinoid biosynthesis.

Kondo F ，Hatakeyama K ，Sakai A ，Minami M ，Nemoto K ，Matsushima K ... -  《-》

Genetic analysis of pungency deficiency in Japanese chili pepper 'Shishito' (Capsicum annuum) revealed its unique heredity and brought the discovery of two genetic loci involved with the reduction of pungency.

The sensation of pungency generated by capsaicinoids is a characteristic trait of chili peppers (Capsicum spp.), and the presence or absence of pungency is central in determining its usage as a spice or a vegetable. In the present study, we aimed to clarify the heredity and genetic factors involved in the deficiency of pungency (quite low pungency) that is uniquely observed in the Japanese chili pepper 'Shishito' (Capsicum annuum). First, the F population ('Shishito' × pungent variety 'Takanotsume') was used for segregation analysis, and pungency level was investigated using capsaicinoid quantification with high-performance liquid chromatography. Also, restriction site associated DNA sequencing of the F population was performed, and genetic map construction and quantitative trait locus (QTL) mapping were implemented. The results indicated that the F population showed varying capsaicinoid content and two major QTLs were detected, Shql3 and Shql7, which explained 39.8 and 19.7% of the genetic variance, respectively. According to these results, the quite low pungency of 'Shishito' was a quantitative trait that involved at least the two loci. Further, this trait was completely separate from general non-pungent traits controlled by individual recessive genes, as described in previous studies. The present study is the first report to investigate the genetic mechanism of pungency deficiency in Japanese chili peppers, and our results provide new insights into the genetic regulation of pungency in chili pepper.

Kondo F ，Umeda K ，Sudasinghe SP ，Yamaguchi M ，Aratani S ，Kumanomido Y ，Nemoto K ，Nagano AJ ，Matsushima K ... -  《-》

Difference in capsaicinoid biosynthesis gene expression in the pericarp reveals elevation of capsaicinoid contents in chili peppers (Capsicum chinense).

This research reveals that the up-regulated expression of multiple capsaicinoid biosynthetic genes in pericarp tissue leads to the elevation of total capsaicinoid content in chili pepper fruit. Capsaicinoids are health-functional compounds that are produced uniquely in chili pepper fruits. A high capsaicinoid level is one of the major parameters determining the commercial quality and health-promoting properties of chili peppers. To investigate the mechanisms responsible for its high contents, we compared an extremely pungent cultivar 'Trinidad Moruga Scorpion Yellow' (MY) with other cultivars of different pungency levels (Fushimi-amanaga, Takanotsume, Red Habanero). Capsaicinoid concentrations were markedly higher in MY fruit (23.9 mg/g DW) than in other pungent cultivars including 'Red Habanero' (HB) fruit (14.3 mg/g DW). Comparative analysis of MY and HB reveals that both cultivars accumulated similar capsaicinoid concentrations in the placental septum, with that in the HB pericarp (1.8 mg/g DW) being markedly lower than that in the placental septum (69.1 mg/g DW). The capsaicinoid concentration in HB fruit is dependent on the placental septum, as reported in other accessions. Therefore, even though placental septum tissue contains high capsaicinoid concentrations, those in the pericarp and seeds attenuated its total content. In contrast, the MY pericarp exhibited a markedly higher concentration (23.2 mg/g DW). A qRT-PCR analysis revealed that multiple capsaicinoid biosynthetic pathway genes (Pun1, pAMT, KAS, and BCAT) were strongly up-regulated in placental septum of pungent cultivars. The genes were expressed exclusively in the MY pericarp, but were barely detected in the pericarps of other pungent cultivars. Collectively, the present study indicates that the up-regulated expression of these genes not only in placental septum but also in pericarp plays an important role in driving capsaicinoid accumulation in the whole fruit.

Tanaka Y ，Nakashima F ，Kirii E ，Goto T ，Yoshida Y ，Yasuba KI ... -  《-》

Molecular biology of capsaicinoid biosynthesis in chili pepper (Capsicum spp.).

Capsicum species produce fruits that synthesize and accumulate unique hot compounds known as capsaicinoids in placental tissues. The capsaicinoid biosynthetic pathway has been established, but the enzymes and genes participating in this process have not been extensively studied or characterized. Capsaicinoids are synthesized through the convergence of two biosynthetic pathways: the phenylpropanoid and the branched-chain fatty acid pathways, which provide the precursors phenylalanine, and valine or leucine, respectively. Capsaicinoid biosynthesis and accumulation is a genetically determined trait in chili pepper fruits as different cultivars or genotypes exhibit differences in pungency; furthermore, this characteristic is also developmentally and environmentally regulated. The establishment of cDNA libraries and comparative gene expression studies in pungent and non-pungent chili pepper fruits has identified candidate genes possibly involved in capsaicinoid biosynthesis. Genetic and molecular approaches have also contributed to the knowledge of this biosynthetic pathway; however, more studies are necessary for a better understanding of the regulatory process that accounts for different accumulation levels of capsaicinoids in chili pepper fruits.

Aza-González C ，Núñez-Palenius HG ，Ochoa-Alejo N 《-》

Mutation in the putative ketoacyl-ACP reductase CaKR1 induces loss of pungency in Capsicum.

A putative ketoacyl-ACP reductase (CaKR1) that was not previously known to be associated with pungency of Capsicum was identified from map-based cloning and functional characterization. The pungency of chili pepper fruits is due to the presence of capsaicinoids, which are synthesized through the convergence of the phenylpropanoid and branched-chain fatty acid pathways. The extensive, global use of pungent and non-pungent peppers underlines the importance of understanding the genetic mechanism underlying capsaicinoid biosynthesis for breeding pepper cultivars. Although Capsicum is one of the earliest domesticated plant genera, the only reported genetic causes of its loss of pungency are mutations in acyltransferase (Pun1) and putative aminotransferase (pAMT). In this study, a single recessive gene responsible for the non-pungency of pepper No.3341 (C. chinense) was identified on chromosome 10 using an F population derived from a cross between Habanero and No.3341. Five candidate genes were identified in the target region, within a distance of 220 kb. A candidate gene, a putative ketoacyl-ACP reductase (CaKR1), of No.3341 had an insertion of a 4.5-kb transposable element (TE) sequence in the first intron, resulting in the production of a truncated transcript missing the region coding the catalytic domain. Virus-induced gene silencing of CaKR1 in pungent peppers resulted in the decreased accumulation of capsaicinoids, a phenotype consistent with No.3341. Moreover, GC-MS analysis of 8-methyl-6-nonenoic acid, which is predicted to be synthesized during the elongation cycle of branched-chain fatty acid biosynthesis, revealed that its deficiency in No.3341. Genetic, genomic, transcriptional, silencing, and biochemical precursor analyses performed in combination provide a solid ground for the conclusion that CaKR1 is involved in capsaicinoid biosynthesis and that its disruption results in a loss of pungency.

Koeda S ，Sato K ，Saito H ，Nagano AJ ，Yasugi M ，Kudoh H ，Tanaka Y ... -  《-》