Multi‐Omics Dissection Reveals the Central Regulator NtHY5 Rewires Primary and Secondary Metabolism to Strengthen Biotic and Abiotic Stress Defences in Nicotiana Tabacum

ABSTRACT

ELONGATED HYPOCOTYL5 (HY5), a bZIP transcription factor, is a central regulator of light signalling and secondary metabolism, yet its role in coordinating primary metabolism with plant stress responses remains unclear. Here, we investigated HY5 function in Nicotiana tabacum using wild‐type, HY5‐overexpressing (NtHY5OX), and CRISPR/Cas9‐generated HY5 knockout ( NtHY5 ^CR ) lines. Integrated transcriptomic analyses of leaves and roots, combined with LC/MS‐ and GC/MS‐based metabolite profiling, revealed that HY5 overexpression promotes a broad metabolic reprogramming characterised by enhanced expression of genes associated with the Calvin cycle, tricarboxylic acid (TCA) cycle, flavonoid biosynthesis, and nicotine metabolism. These transcriptional changes were accompanied by increased accumulation of phenolic compounds and alkaloids, indicating a shift in metabolic reprogramming toward defence‐related specialised metabolism. In contrast, NtHY5 ^CR mutants accumulated higher levels of amino acids, lipids, and organic acids, consistent with prioritisation of growth‐associated primary metabolism at the expense of protective secondary metabolite production. Pathway enrichment analyses identified HY5 as a key regulatory node integrating central carbon metabolism with specialised metabolite biosynthesis. Functionally, this HY5‐dependent metabolic configuration enhanced resistance to Alternaria solani and improved tolerance to salt stress, demonstrating that HY5‐mediated metabolic plasticity underpins adaptive stress responses in tobacco. Together, these findings establish HY5 as a critical coordinator of metabolic and defence networks, providing mechanistic insight into how light‐responsive transcriptional regulation shapes plant stress resilience and offering targets for engineering stress‐tolerant crops.