Mechanism of tobacco hairy root development and functional study of the HSF gene family based on comparative transcriptomics

Hairy roots induced by Agrobacterium rhizogenes exhibit unique phenotypic traits such as rapid proliferation, hormone-independent growth. To further elucidate the transcriptomic differences between hairy roots and the roots, stems, and leaves of normal plants, as well as their potential mechanisms of formation, this study conducted a comparative transcriptomic analysis. The results indicated that differentially expressed genes (DEGs) were significantly enriched in pathways related to energy metabolism, stress response, and secondary metabolism. Then weighted gene co-expression network analysis (WGCNA) analysis identified two key co-expression modules. Moreover, among the differentially expressed transcription factors (TFs), the HSF (heat shock transcription factor) family showed the most significant enrichment. Further analysis of 17 HSFs revealed that all of them were significantly correlated with plant hormones. Among these, two were specifically expressed in hairy roots, and nine exhibited both constitutive high expression and responsiveness to salicylic acid (SA), melatonin and drought treatments. These results establish an HSF-mediated regulatory network, offering valuable theoretical insights and potential targets for breeding stress-resistant germplasm with enhanced root systems, as well as for the synthetic biology-based engineering of hairy roots.