Cloning and functional analysis of transcription factor <i>AaCMB1</i> on carotenoid accumulation in spathe of <i>Anthurium andraeanum</i>

<i>Anthurium andraeanum</i> Linden, a flowering foliage plant belonging to the Araceae family, exhibits colorful spathes but lacks deep yellow cultivars. The carotenoid metabolic pathway is crucial for the accumulation of yellow pigments (e.g. lutein) in plant cells, with transcriptional regulation playing a key role in this process. Among these regulators, the MADS-box family represents one of the major transcription factor families involved in this regulatory network. To investigate the regulatory role of MADS-box family transcription factors in carotenoid metabolism in <i>Anthurium</i> spathes, a MADS-box gene from the light yellow spathe cultivar ‘Vanilla’ was cloned and designated as <i>AaCMB1</i>. Subcellular localization analysis revealed that AaCMB1 is localized in the nucleus. Transient overexpression of <i>AaCMB1</i> increased carotenoid accumulation in <i>Anthurium</i> spathes, and quantitative real-time polymerase chain reaction (qRT-PCR) analysis showed that overexpression of <i>AaCMB1</i> upregulated the expression of key structural genes in the carotenoid biosynthetic pathway. Yeast two-hybrid and bimolecular fluorescence complementation (BiFC) assays demonstrated that AaCMB1 interacts with AaNAC1574. Yeast one-hybrid and dual-luciferase reporter assays further indicated that AaCMB1 can activate the <i>AaLCYB</i> promoter. These findings suggest that AaCMB1 and AaNAC1574 may form a heterodimer to regulate the expression of <i>AaLCYB</i>, thereby modulating carotenoid accumulation in <i>Anthurium</i> spathes.

花烛（Anthurium andraeanum）Linden是隶属于天南星科的观花观叶植物，其佛焰苞色彩丰富，但目前尚无深黄色栽培品种。类胡萝卜素代谢通路对植物细胞内黄色色素（如叶黄素）的积累至关重要，而转录调控在该过程中发挥关键作用。在这类调控因子中，MADS-box家族是参与该调控网络的主要转录因子家族之一。为探究MADS-box家族转录因子在花烛佛焰苞类胡萝卜素代谢中的调控作用，研究人员从浅黄色佛焰苞品种‘Vanilla’中克隆得到一个MADS-box基因，并将其命名为AaCMB1。亚细胞定位分析显示，AaCMB1定位于细胞核中。对AaCMB1进行瞬时过表达可提升花烛佛焰苞的类胡萝卜素积累量，实时荧光定量聚合酶链式反应（quantitative real-time polymerase chain reaction，qRT-PCR）分析表明，AaCMB1过表达会上调类胡萝卜素生物合成通路中的关键结构基因的表达。酵母双杂交和双分子荧光互补（bimolecular fluorescence complementation，BiFC）实验证实，AaCMB1可与AaNAC1574发生相互作用。酵母单杂交和双荧光素酶报告基因实验进一步表明，AaCMB1能够激活AaLCYB启动子。上述研究结果表明，AaCMB1与AaNAC1574可能形成异源二聚体，通过调控AaLCYB的表达，进而调节花烛佛焰苞中的类胡萝卜素积累。