Cooperation between transcription factors BBX15 and bHLH87 enhances lily thermotolerance by activating a heat shock factor gene

BBX (B-box) proteins are critical regulators in plant growth and development, playing pivotal roles in processes such as photomorphogenesis and floral transition. Additionally, they are involved in plant responses to various abiotic stresses, including salt, drought, and cold. However, their specific roles in thermotolerance remain largely unexplored. In this study, we identified a heat-inducible BBX gene, LlBBX15, which belongs to the class III BBX subfamily of lily (Lilium longiflorum). LlBBX15 localized to the nucleus and exhibited transcriptional activation activity. Stable overexpression of LlBBX15 resulted in increased thermotolerance in both Arabidopsis (Arabidopsis thaliana) and lily, whereas silencing LlBBX15 in lily led to a reduction in thermotolerance. Furthermore, LlBBX15 interacted with LlbHLH87 (BASIC HELIX-LOOP-HELIX FACTOR 87) and directly bound to the promoter of LlHSFA2 (HEAT STRESS TRANSCRIPTION FACTOR A2), thereby activating its expression. Subsequent analyses revealed that the heterologous interaction between LlBBX15 and LlbHLH87 facilitated their respective homologous interactions. A complex of LlBBX15 and LlbHLH87 enhanced their DNA-binding capacity and cooperatively promoted the expression of LlHSFA2. Moreover, both LlHSFA1 and LlHSFA2 were identified as direct regulators of LlBBX15, with evidence suggesting a synergetic activation effect on its expression. This interaction indicates the existence of a feedback loop between the HSFs and LlBBX15. Collectively, these findings establish LlBBX15 as a positive regulator that collaborates with LlbHLH87 within the HSF signaling pathway to facilitate thermotolerance in plants.