Positional differences of intronic transposons in pAMT affect the pungency level in chili pepper through altered splicing efficiency

Yoshiyuki Tanaka, Takaya Asano, Yorika Kanemitsu, Tanjuro Goto, Yuichi Yoshida, Kenichiro Yasuba, Yuki Misawa, Sachie Nakatani, Kenji Kobata

Research output: Contribution to journalArticle

Abstract

Capsaicinoids are unique compounds that give chili pepper fruits their pungent taste. Capsaicinoid levels vary widely among pungent cultivars, which range from low pungency to extremely pungent. However, the molecular mechanisms underlying this quantitative variation have not been elucidated. Our previous study identified various loss-of-function alleles of the pAMT gene which led to low pungency. The mutations in these alleles are commonly defined by Tcc transposon insertion and its footprint. In this study, we identified two leaky pamt alleles (pamtL1 and pamtL2) with different levels of putative aminotransferase (pAMT) activity. Notably, both alleles had a Tcc transposon insertion in intron 3, but the locations of the insertions within the intron were different. Genetic analysis revealed that pamtL1, pamtL2 and a loss-of-function pamt allele reduced capsaicinoid levels to about 50%, 10% and less than 1%, respectively. pamtL1 and pamtL2 encoded functional pAMT proteins, but they exhibited lower transcript levels than the functional type. RNA sequencing analysis showed that intronic transposons disrupted splicing in intron 3, which resulted in simultaneous expression of functional pAMT mRNA and non-functional splice variants containing partial sequences of Tcc. The non-functional splice variants were more dominant in pamtL2 than in pamtL1. This suggested that the difference in position of the intronic transposons could alter splicing efficiency, leading to different pAMT activities and reducing capsaicinoid content to different levels. Our results provide a striking example of allelic variations caused by intronic transposons; these variations contribute to quantitative differences in secondary metabolite contents.

Original languageEnglish
JournalPlant Journal
DOIs
Publication statusAccepted/In press - Jan 1 2019

Fingerprint

Capsicum
hot peppers
transaminases
Transaminases
transposons
Alleles
alleles
Introns
introns
RNA Sequence Analysis
secondary metabolites
genetic techniques and protocols
Fruit
sequence analysis
mutation
Messenger RNA
Mutation
fruits
cultivars
Genes

Keywords

  • capsaicinoid
  • Capsicum chinense
  • capsinoid
  • chili pepper
  • hAT transposon

ASJC Scopus subject areas

  • Genetics
  • Plant Science
  • Cell Biology

Cite this

Positional differences of intronic transposons in pAMT affect the pungency level in chili pepper through altered splicing efficiency. / Tanaka, Yoshiyuki; Asano, Takaya; Kanemitsu, Yorika; Goto, Tanjuro; Yoshida, Yuichi; Yasuba, Kenichiro; Misawa, Yuki; Nakatani, Sachie; Kobata, Kenji.

In: Plant Journal, 01.01.2019.

Research output: Contribution to journalArticle

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abstract = "Capsaicinoids are unique compounds that give chili pepper fruits their pungent taste. Capsaicinoid levels vary widely among pungent cultivars, which range from low pungency to extremely pungent. However, the molecular mechanisms underlying this quantitative variation have not been elucidated. Our previous study identified various loss-of-function alleles of the pAMT gene which led to low pungency. The mutations in these alleles are commonly defined by Tcc transposon insertion and its footprint. In this study, we identified two leaky pamt alleles (pamtL1 and pamtL2) with different levels of putative aminotransferase (pAMT) activity. Notably, both alleles had a Tcc transposon insertion in intron 3, but the locations of the insertions within the intron were different. Genetic analysis revealed that pamtL1, pamtL2 and a loss-of-function pamt allele reduced capsaicinoid levels to about 50{\%}, 10{\%} and less than 1{\%}, respectively. pamtL1 and pamtL2 encoded functional pAMT proteins, but they exhibited lower transcript levels than the functional type. RNA sequencing analysis showed that intronic transposons disrupted splicing in intron 3, which resulted in simultaneous expression of functional pAMT mRNA and non-functional splice variants containing partial sequences of Tcc. The non-functional splice variants were more dominant in pamtL2 than in pamtL1. This suggested that the difference in position of the intronic transposons could alter splicing efficiency, leading to different pAMT activities and reducing capsaicinoid content to different levels. Our results provide a striking example of allelic variations caused by intronic transposons; these variations contribute to quantitative differences in secondary metabolite contents.",
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AU - Tanaka, Yoshiyuki

AU - Asano, Takaya

AU - Kanemitsu, Yorika

AU - Goto, Tanjuro

AU - Yoshida, Yuichi

AU - Yasuba, Kenichiro

AU - Misawa, Yuki

AU - Nakatani, Sachie

AU - Kobata, Kenji

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N2 - Capsaicinoids are unique compounds that give chili pepper fruits their pungent taste. Capsaicinoid levels vary widely among pungent cultivars, which range from low pungency to extremely pungent. However, the molecular mechanisms underlying this quantitative variation have not been elucidated. Our previous study identified various loss-of-function alleles of the pAMT gene which led to low pungency. The mutations in these alleles are commonly defined by Tcc transposon insertion and its footprint. In this study, we identified two leaky pamt alleles (pamtL1 and pamtL2) with different levels of putative aminotransferase (pAMT) activity. Notably, both alleles had a Tcc transposon insertion in intron 3, but the locations of the insertions within the intron were different. Genetic analysis revealed that pamtL1, pamtL2 and a loss-of-function pamt allele reduced capsaicinoid levels to about 50%, 10% and less than 1%, respectively. pamtL1 and pamtL2 encoded functional pAMT proteins, but they exhibited lower transcript levels than the functional type. RNA sequencing analysis showed that intronic transposons disrupted splicing in intron 3, which resulted in simultaneous expression of functional pAMT mRNA and non-functional splice variants containing partial sequences of Tcc. The non-functional splice variants were more dominant in pamtL2 than in pamtL1. This suggested that the difference in position of the intronic transposons could alter splicing efficiency, leading to different pAMT activities and reducing capsaicinoid content to different levels. Our results provide a striking example of allelic variations caused by intronic transposons; these variations contribute to quantitative differences in secondary metabolite contents.

AB - Capsaicinoids are unique compounds that give chili pepper fruits their pungent taste. Capsaicinoid levels vary widely among pungent cultivars, which range from low pungency to extremely pungent. However, the molecular mechanisms underlying this quantitative variation have not been elucidated. Our previous study identified various loss-of-function alleles of the pAMT gene which led to low pungency. The mutations in these alleles are commonly defined by Tcc transposon insertion and its footprint. In this study, we identified two leaky pamt alleles (pamtL1 and pamtL2) with different levels of putative aminotransferase (pAMT) activity. Notably, both alleles had a Tcc transposon insertion in intron 3, but the locations of the insertions within the intron were different. Genetic analysis revealed that pamtL1, pamtL2 and a loss-of-function pamt allele reduced capsaicinoid levels to about 50%, 10% and less than 1%, respectively. pamtL1 and pamtL2 encoded functional pAMT proteins, but they exhibited lower transcript levels than the functional type. RNA sequencing analysis showed that intronic transposons disrupted splicing in intron 3, which resulted in simultaneous expression of functional pAMT mRNA and non-functional splice variants containing partial sequences of Tcc. The non-functional splice variants were more dominant in pamtL2 than in pamtL1. This suggested that the difference in position of the intronic transposons could alter splicing efficiency, leading to different pAMT activities and reducing capsaicinoid content to different levels. Our results provide a striking example of allelic variations caused by intronic transposons; these variations contribute to quantitative differences in secondary metabolite contents.

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