Tea plant leaf droopiness traced to one-base mutation regulating CsTPR

Mechanical harvesting is replacing labor-intensive hand-plucking to meet global tea production needs. However, many traditional tea cultivars have soft, droopy leaves that break easily during machine cutting, leading to yield loss. Previous work identified brassinosteroid signaling and transcription factor CsBES1.2 as key promoters of droopiness, yet the negative regulators of this trait remained largely unknown. Understanding gene networks behind leaf architecture is essential for breeding machine-friendly cultivars. Despite the importance of TPR proteins in hormone pathways, their role in tea droopiness has not been clarified. Based on these challenges, further research was needed to identify regulatory genes and mutation sites controlling leaf drooping traits.

Researchers from the Tea Research Institute, Chinese Academy of Agricultural Sciences, together with collaborators, reported (DOI: 10.1093/hr/uhaf098) a new on March 25, 2025 in Horticulture Research. The team discovered that a single-base mutation in the promoter of CsTPR enhances transcriptional repression by CsBES1.2, intensifying brassinosteroid-induced leaf drooping. By integrating genome-wide association study (GWAS), RNA sequencing (RNA-seq), assay for transposase-accessible chromatin using sequencing (ATAC-seq), whole-genome sequencing (WGS), promoter cloning, ChIP-seq, EMSA, and transient assays, the study reveals how promoter variation shifts gene expression and reshapes leaf morphology, providing a breeding target for mechanized tea production.

Through genome-wide association analysis of 130 tea accessions, the team identified a droopiness-related QTL q10.3 containing CsTPR. RNA-seq and ATAC-seq demonstrated higher chromatin openness and expression of CsTPR in straight-leaf varieties than in droopy types. Functional assays showed that silencing CsTPR increased droopiness severity and reduced vascular tissue length by ~25%, confirming its suppressive role. Whole-genome sequencing further revealed a single T→A mutation at –53 bp in the promoter region, located precisely within the CsBES1.2-binding E-box. ChIP-seq and ChIP-qPCR indicated that CsBES1.2 binds this site more strongly in mutant genotypes, and EMSA validated the enhanced affinity in vitro. Reporter assays confirmed that CsBES1.2 significantly reduced CsTPRpro:LUC activity, with stronger inhibition observed in mutant promoter lines.

These results demonstrate that CsTPR represses droopiness, while the promoter mutation increases CsBES1.2-mediated inhibition, lowering CsTPR expression and accelerating leaf bending. The mechanism establishes a genetic basis for leaf morphology variation and highlights promoter sequences as key regulatory nodes in tea breeding.

“This discovery fills a missing link in the droopiness regulatory network,” said the study authors. “We now understand how a single nucleotide change can amplify transcription factor suppression and reshape plant architecture. CsTPR acts as a counterforce to brassinosteroid-induced drooping, and altering its promoter sensitivity directly affects leaf mechanical strength. This work provides a precise molecular target for future breeding and demonstrates the value of integrating multi-omics with genetic validation.”

The identified mutation and regulatory module (CsTPR—CsBES1.2—E-box) offer promising genetic markers for developing tea cultivars compatible with mechanical harvesting. By selecting alleles with weaker CsBES1.2 binding or enhancing CsTPR expression, breeders may create straighter leaves with reduced breakage. This strategy can improve harvesting efficiency, reduce labor dependence, and increase economic returns for large-scale plantations. Future applications may extend to CRISPR promoter editing and marker-assisted breeding. The work also sheds light on transcription factor–promoter dynamics, offering reference for studying similar traits in other perennial crops.

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References

DOI

10.1093/hr/uhaf098

Original Source URL

https://doi.org/10.1093/hr/uhaf098

Funding information

This work was supported by the grants from Zhejiang Provincial Natural Science Foundation of China (LQ23C160010), Jiangsu Seed Industry Revitalization Project JBGS[2021]081, Fundamental Research Fund for Tea Research Institute of the Chinese Academy of Agricultural Sciences (grant number 1610212022002), Jiangxi Province Talent Plan (jxsq2023102020), China Agriculture Research System of MOF and MARA (CARS-19), Yunnan Province of Science and Technology Department (202449CE340010), and National Key Laboratory for Tea Plant Germplasm Innovation and Resource Utilization (SQ2024SKL03104).

About Horticulture Research

Horticulture Research is an open access journal of Nanjing Agricultural University and ranked number one in the Horticulture category of the Journal Citation Reports ™ from Clarivate, 2023. The journal is committed to publishing original research articles, reviews, perspectives, comments, correspondence articles and letters to the editor related to all major horticultural plants and disciplines, including biotechnology, breeding, cellular and molecular biology, evolution, genetics, inter-species interactions, physiology, and the origination and domestication of crops.