Grafting, one of agriculture’s oldest propagation techniques, has been used for millennia to improve crop resilience and productivity. In cucurbits like watermelon, retaining at least one cotyledon on the rootstock is essential for survival, suggesting its crucial physiological role. Cotyledons serve as the seedling’s early life-support system, producing and releasing nutrients and hormones such as auxin that guide tissue regeneration. In model plants, auxin has been shown to promote cell proliferation and vascular bridge formation at the graft junction. However, how watermelon cotyledons coordinate auxin transport and which molecular components regulate this process remain unclear. Addressing these questions, researchers sought to reveal the genetic and hormonal basis of watermelon graft healing.
Researchers from Huazhong Agricultural University and Nagoya University have now identified the molecular link between watermelon cotyledons and graft healing. Their study (DOI: 10.1093/hr/uhae329) , published on March 1, 2025, in Horticulture Research, uncovers the role of ClPIN1a, an auxin efflux carrier that directs hormone flow from the rootstock cotyledon to the graft interface, enabling tissue regeneration and vascular reconnection. By integrating a newly developed seed-soaking VIGS system with cotyledon grafting, the team demonstrated how controlling auxin movement through ClPIN1a determines graft success—a discovery poised to advance cucurbit breeding and propagation technologies.
The team compared watermelon seedlings grafted with or without rootstock cotyledons. Seedlings lacking cotyledons exhibited slower growth, reduced callus formation, and delayed reconnection of xylem and phloem by nearly three days. Quantitative assays revealed that removing cotyledons sharply decreased indole-3-acetic acid (IAA) levels in rootstocks, while exogenous auxin application restored graft healing efficiency.
To explore the mechanism, the scientists developed a cotyledon grafting system with an 85% survival rate, enabling the tracing of hormone movement between grafted organs. In parallel, they introduced an innovative seed-soaking VIGS technique that achieved an 83% infection rate and up to 75% gene-silencing efficiency—nearly doubling the success of traditional cotyledon injection. When ClPIN1a was silenced in rootstock cotyledons, auxin accumulated locally but failed to reach the graft junction, resulting in persistent necrotic layers and poor survival. These findings identify ClPIN1a as a key regulator that channels auxin from the cotyledon toward the graft interface, effectively powering tissue regeneration and vascular bridge formation during graft healing.
“Our study provides direct genetic evidence that cotyledon-derived auxin transport is essential for graft healing,” said Professor Yuan Huang, the corresponding author of the study. “By identifying ClPIN1a as the central auxin transporter in this process, we connected the ancient practice of grafting with the modern molecular understanding of plant regeneration. The seed-soaking VIGS method we established not only accelerates functional genomics in cucurbits but also offers a practical platform to study hormonal signaling and developmental coordination in early-stage seedlings.”
This discovery opens new avenues for improving grafting efficiency and plant survival in watermelon and other cucurbit crops. Manipulating ClPIN1a expression or applying tailored auxin treatments could enhance callus formation and vascular integration, ensuring more consistent graft performance in commercial production. Beyond its agricultural value, the study introduces a scalable genetic tool for early functional gene validation, accelerating research on plant regeneration and compatibility. By bridging classical horticultural wisdom with molecular biology, this work advances sustainable crop propagation practices in an era of climate and resource challenges.
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References
DOI
10.1093/hr/uhae329
Original Source URL
https://doi.org/10.1093/hr/uhae329
Funding information
This work was supported by the National Natural Science Foundation of China (31972434), the China Agriculture Research System of MOF and MORA (CARS-25), the Hubei Provincial Key Research and Development Program (2023BBB033), the Fundamental Research Funds for the Central Universities (2662024JC004), the Young Scientist Forstering Funds for the National Key Laboratory for Germplasm Innovation and Utilization of Horticultural Crops (11909920008), and the Huazhong Agricultural University-Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences Cooperation Fund (SZYJY2021005), Shannan Science and Technology Plan Project(SNSBJKJJHXM2023004).
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.