Abstract
WRKY transcription factors (TFs) are essential in regulating plant secondary metabolism, yet their role in the saponin biosynthesis of Eleutherococcus senticosus remains unclear. This study aimed to identify and characterize EsWRKY TFs associated with saponin biosynthesis. Bioinformatics analyses and correlation studies indicated the strongest correlation of EsWRKY7 , EsWRKY35 , EsWRKY40 , and EsWRKY88 with saponin production. Subcellular localization confirmed their nuclear distribution; regulatory activity was influenced by light and DNA methylation, with Es WRKY7, Es WRKY35, and Es WRKY40 possessing transcriptional autoactivation ability. Overexpression experiments revealed that Es WRKY7 and Es WRKY88 negatively regulated saponin biosynthesis genes, reducing saponin content, while Es WRKY35 and Es WRKY40 acted as positive regulators, promoting saponin accumulation. Through the application of Y2H and BiFC assays, a yeast library screening uncovered several proteins that interact with Es WRKYs. Furthermore, three regulatory mechanisms mediated by Es WRKY in saponin biosynthesis were elucidated: direct promoter binding, transcriptional regulation via DNA methylation-sensitive elements, and regulation through protein complex formation. Molecular dynamics simulations provided insights into the variations in the regulatory function of Es WRKYs upon protein interaction. These findings unveil the multifaceted modulatory network of saponin biosynthesis in E. senticosus and lay a foundation for enhancing saponin accumulation through metabolic engineering. • Four core EsWRKYs transcription factors were identified to oppositely regulate saponin biosynthesis. • Es WRKYs directly bind promoters of saponin synthases ( EsFPS / EsSS / EsSE ) to control their expression. • A novel bidirectional interplay between Es WRKYs and DNA methylation fine-tunes promoter binding. • Protein interactions modulate Es WRKYs' DNA-binding stability, thereby affecting transcriptional activity. • A triple-mechanism framework (direct binding, methylation sensing, protein interactome) is proposed.
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Claims 12
Claim · model-written summary
EsWRKY7, EsWRKY35, EsWRKY40, and EsWRKY88 had the strongest correlations with saponin production.
- Taxon
- Eleutherococcus senticosus
Eleutherococcus senticosuse1 - Gene
- EsWRKY7 , EsWRKY35 , EsWRKY40 , and EsWRKY88e0
- Outcome
- saponin productione0
Not reportedDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“the strongest correlation of EsWRKY7 , EsWRKY35 , EsWRKY40 , and EsWRKY88 with saponin production”
e1“Eleutherococcus senticosus”
Claim · model-written summary
The four studied EsWRKY factors showed nuclear localization.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse2
- Gene
- EsWRKY7 , EsWRKY35 , EsWRKY40 , and EsWRKY88e1
- Outcome
- nuclear distributione0
Not reportedDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Subcellular localization confirmed their nuclear distribution”
e1“EsWRKY7 , EsWRKY35 , EsWRKY40 , and EsWRKY88”
e2“Eleutherococcus senticosus”
Claim · model-written summary
Light and DNA methylation influenced regulatory activity of EsWRKY factors.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Treatment
- light and DNA methylatione0
- Outcome
- regulatory activitye0
Not reportedDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“regulatory activity was influenced by light and DNA methylation”
e1“Eleutherococcus senticosus”
Claim · model-written summary
Es WRKY7, Es WRKY35, and Es WRKY40 possessed transcriptional autoactivation ability.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Gene
- Es WRKY7, Es WRKY35, and Es WRKY40e0
- Outcome
- transcriptional autoactivation abilitye0
Not reportedDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Es WRKY7, Es WRKY35, and Es WRKY40 possessing transcriptional autoactivation ability”
e1“Eleutherococcus senticosus”
Claim · model-written summary
Overexpressed Es WRKY7 negatively regulated saponin biosynthesis genes and reduced saponin content.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Treatment
- Overexpressione0
- Gene
- Es WRKY7e0
- Outcome
- saponin contente0
- Direction
- reducinge0
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Overexpression experiments revealed that Es WRKY7 and Es WRKY88 negatively regulated saponin biosynthesis genes, reducing saponin content”
e1“Eleutherococcus senticosus”
Claim · model-written summary
Overexpressed Es WRKY88 negatively regulated saponin biosynthesis genes and reduced saponin content.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Treatment
- Overexpressione0
- Gene
- Es WRKY88e0
- Outcome
- saponin contente0
- Direction
- reducinge0
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Overexpression experiments revealed that Es WRKY7 and Es WRKY88 negatively regulated saponin biosynthesis genes, reducing saponin content”
e1“Eleutherococcus senticosus”
Claim · model-written summary
Es WRKY35 promoted saponin accumulation as a positive regulator.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Gene
- Es WRKY35e0
- Outcome
- saponin accumulatione0
- Direction
- promotinge0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Es WRKY35 and Es WRKY40 acted as positive regulators, promoting saponin accumulation”
e1“Eleutherococcus senticosus”
Claim · model-written summary
Es WRKY40 promoted saponin accumulation as a positive regulator.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Gene
- Es WRKY40e0
- Outcome
- saponin accumulatione0
- Direction
- promotinge0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Es WRKY35 and Es WRKY40 acted as positive regulators, promoting saponin accumulation”
e1“Eleutherococcus senticosus”
Claim · model-written summary
Yeast screening identified several proteins interacting with Es WRKYs.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Gene
- Es WRKYse0
- Outcome
- several proteins that interact with Es WRKYse0
Not reportedDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“a yeast library screening uncovered several proteins that interact with Es WRKYs”
e1“Eleutherococcus senticosus”
Claim · model-written summary
Es WRKYs directly bound promoters of saponin synthases to control expression.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Gene
- Es WRKYse0
- Outcome
- their expressione0
- Qualifier
- directly bind promoters of saponin synthases ( EsFPS / EsSS / EsSE )e0
Not reportedDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Es WRKYs directly bind promoters of saponin synthases ( EsFPS / EsSS / EsSE ) to control their expression”
e1“Eleutherococcus senticosus”
Claim · model-written summary
DNA methylation and Es WRKYs interacted bidirectionally to tune promoter binding.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Gene
- Es WRKYse0
- Outcome
- promoter bindinge0
- Qualifier
- bidirectional interplay between Es WRKYs and DNA methylatione0
Not reportedDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“A novel bidirectional interplay between Es WRKYs and DNA methylation fine-tunes promoter binding”
e1“Eleutherococcus senticosus”
Claim · model-written summary
Protein interactions modulated Es WRKY DNA binding stability and transcriptional activity.
- Taxon
- Eleutherococcus senticosusEleutherococcus senticosuse1
- Gene
- Es WRKYse0
- Outcome
- transcriptional activitye0
- Qualifier
- Protein interactionse0
Not reportedDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Protein interactions modulate Es WRKYs' DNA-binding stability, thereby affecting transcriptional activity”
e1“Eleutherococcus senticosus”