The pungency-variable sweet chili pepper 'Shishito' (Capsicum annuum) is widely cultivated in Japan. While 'Shishito' is generally used as a vegetable because of its relatively low pungency, it sometimes exhibits high pungency depending on cultivation conditions. Although the occurrence of pungent 'Shishito' fruits is a problem in retail distribution and utilization, the responsible mechanism is largely unknown. As one approach to clarify the mechanism, we focused on the effects of

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... (resulting in seedless fruit) on the pungency traits of 'Shishito' fruits. In the present study, artificial parthenocarpic 'Shishito' fruits were prepared by treatment with 2,4-dichlorophenoxyacetic acid (2,4-D), and the pungency level was investigated by quantification of capsaicinoids, components responsible for the pungency. For comparison, two controls were used: naturally pollinated fruits and 2,4-D-treated pollinated fruits which were exposed to 2,4-D treatment without parthenocarpy. The results indicated that the capsaicinoid content in parthenocarpic fruits tended to be higher compared to the controls. This suggests that the alterations in pungency in 'Shishito' were associated with parthenocarpy. Further, these relationships were assessed using a molecular method, and gene expression analysis using qRT-PCR was conducted on 16 capsaicinoid biosynthesis genes. The results showed that eight capsaicinoid biosynthesis genes (Pun1, pAMT, KAS, CaMYB31, BCAT, CaKR1, ACL, and FAT) exhibited parthenocarpy-specific high expression, suggesting that these genes influence capsaicinoid biosynthesis and the pungency levels in parthenocarpic fruits. This is a novel report that carefully investigated the parthenocarpy-dependent changes in the pungency traits of chili pepper. We anticipate that the data will add to horticultural knowledge and help control the pungency of 'Shishito' fruits during cultivation.