The AhDREB transgene expression activates NtP5CS and NtSUSY, promoting osmotic adjustment in transgenic tobacco under salt stress

Soil salinity isamajor environmental constraint that limits the growth and productivity ofpeanut (Arachis hypogaea L.), alegume adapted tomildly acidic soils but highly sensitive tosaline–alkaline conditions. Dehydration-Responsive Element Binding (DREB) transcription factors are key regulators ofplant responses toabiotic stresses. Inthis study, the AhDREB gene from peanut was introduced into tobacco (Nicotiana tabacum) toexamine its functional role under salt stress. The transgenic lines (L32.2 and L37.2) exhibited strong induction ofAhDREB expression upon exposure to150 and 250 mM NaCl, with transcript levels increasing up to2.34-fold compared with untreated controls (P<0.001). Quantitative RT-PCR analysis revealed that AhDREB enhanced the transcription oftwo osmolyte-related genes, NtP5CS and NtSUSY. Under saline conditions, the expression ofthese genes was 1.20–1.89-fold higher intransgenic lines than inwild-type (WT) plants and 4.74–7.66-fold higher than innon-stress conditions (P<0.001). Consistently, both lines accumulated greater amounts ofproline and soluble sugars, showing 2.09–2.30-fold and 2.40–4.70-fold increases, respectively, compared with the WT. Relative tonon-stress conditions, proline and sugar contents increased by3.59–5.47 fold and 3.75–7.65-fold, respectively. Line L37.2 accumulated higher proline levels, whereas L32.2 exhibited greater sugar content, indicating distinct osmolyte regulation patterns. Overall, the AhDREB gene enhances salt tolerance in tobacco by transcriptionally activating osmolyte biosynthetic pathways and improving cellular osmotic adjustment, providing molecular evidence for its potential application in developing stress-tolerant peanut cultivars.