Auxin-Induced Transcriptional Complex Regulates Garlic Somatic Embryogenesis

Garlic (Allium sativum L.) is usually propagated through bulbs rather than seeds, a pattern that can lead to variety degeneration, low multiplication rates, and the accumulation of viral diseases. Somatic embryogenesis offers a powerful way to regenerate plants from somatic cells and supports germplasm conservation, rapid propagation, virus elimination, and molecular breeding. Although auxin is known to be central to plant regeneration, the RNA-based regulatory networks that translate auxin signals into embryogenic development in garlic have remained unclear. Based on these challenges, in-depth research is needed into the hormone-responsive molecular mechanisms that regulate garlic somatic embryogenesis.

A research team from the Sanya Research Institute of Nanjing Agricultural University and the College of Horticulture, Nanjing Agricultural University, with collaborators from the University of Georgia and Aarhus University, published (DOI: 10.1093/hr/uhag016) the study in Horticulture Research on 20 January 2026. The study reports that an auxin-induced AsARF16-centered transcriptional complex controls a competing endogenous RNA network involving lncRNA125175, AsmiR393h, and AsTIR1, thereby regulating garlic somatic embryogenesis.

The researchers first confirmed that lncRNA125175 is a true long noncoding RNA (lncRNA), rather than a protein-coding transcript. They then examined how it interacts with AsmiR393h, a microRNA (miRNA), and AsTIR1, a gene encoding an auxin receptor linked to hormone perception. Using transient expression assays in tobacco leaves and garlic protoplasts, they showed that AsmiR393h directly cleaves AsTIR1 transcripts and suppresses their expression. However, lncRNA125175 functions as an endogenous target mimic (eTM), binding AsmiR393h and reducing its ability to repress AsTIR1. This forms a competing endogenous RNA (ceRNA) module that fine-tunes auxin signaling during regeneration.

The team further tested the response of this module to 2,4-Dichlorophenoxyacetic acid (2,4-D), a synthetic auxin widely used to induce plant somatic embryogenesis, and 1-N-naphthylphthalamic acid (NPA), an auxin transport inhibitor. Their results showed that 2,4-D promoted callus formation, while NPA disrupted this process. Promoter analysis found auxin-responsive elements in lncRNA125175, AsmiR393h, and AsTIR1, and reporter assays showed that auxin enhanced the promoter activity of lncRNA125175 and AsTIR1. Weighted gene co-expression network analysis then identified AsARF16 as a key upstream regulator. Yeast one-hybrid assays showed that AsARF16 binds directly to the promoter of lncRNA125175. Protein interaction assays further demonstrated that AsARF16 interacts with AsWRKY31 and AsIAA33, forming a transcriptional activation complex that links auxin perception to RNA-mediated regulation.

The authors said the study explains how garlic cells convert an external hormone cue into a precise developmental program. They said the pathway is important because it connects three regulatory layers that are often studied separately: auxin signaling, transcription factor control, and post-transcriptional RNA regulation. In this model, AsARF16 serves as the central DNA-binding regulator, AsIAA33 connects the system to canonical auxin signaling, and AsWRKY31 may strengthen developmental specificity. Together, they activate lncRNA125175, allowing the downstream ceRNA network to support the embryogenic transition.

The findings offer practical value for garlic breeding and biotechnology. By identifying lncRNA125175, AsmiR393h, AsTIR1, and the AsARF16–AsWRKY31–AsIAA33 complex as key regulatory components, the study provides candidate molecular targets for improving garlic regeneration efficiency. Stronger somatic embryogenesis systems could support virus-free seedling production, germplasm preservation, and more reliable genetic transformation or gene editing platforms. More broadly, the work highlights how long noncoding RNAs can shape plant hormone responses, offering insights that may also benefit other asexually propagated crops facing similar breeding and regeneration bottlenecks.