Frick Karen Michiko, Lorensen Marcus Daniel Brandbjerg Bohn, Micic Nikola, Esteban Eddi, Pasha Asher, Schulz Alexander, Provart Nicholas James, Nour-Eldin Hussam Hassan, Bjarnholt Nanna, Janfelt Christian, Geu-Flores Fernando
Section for Plant Biochemistry and Copenhagen Plant Science Centre, Department of Plant and Environmental Sciences, University of Copenhagen, 1871, Frederiksberg, Denmark.
Department of Pharmacy, University of Copenhagen, 2100, Copenhagen, Denmark.
New Phytol. 2025 Mar;245(5):2052-2068. doi: 10.1111/nph.20384. Epub 2025 Jan 14.
Lupins are promising protein crops that accumulate toxic quinolizidine alkaloids (QAs) in the seeds, complicating their end-use. QAs are synthesized in green organs (leaves, stems, and pods) and a subset of them is transported to the seeds during fruit development. The exact sites of biosynthesis and accumulation remain unknown; however, mesophyll cells have been proposed as sources, and epidermal cells as sinks. We investigated the exact sites of QA biosynthesis and accumulation in biosynthetic organs of narrow-leafed lupin (Lupinus angustifolius) using mass spectrometry-based imaging (MSI), laser-capture microdissection coupled to RNA-Seq, and precursor feeding studies coupled to LC-MS and MSI. We found that the QAs that accumulate in seeds ('core' QAs) were evenly distributed across tissues; however, their esterified versions accumulated primarily in the epidermis. Surprisingly, RNA-Seq revealed strong biosynthetic gene expression in the epidermis, which was confirmed in leaves by quantitative real-time polymerase chain reaction. Finally, feeding studies using a stably labeled precursor showed that the lower leaf epidermis is highly biosynthetic. Our results indicate that the epidermis is a major site of QA biosynthesis in narrow-leafed lupin, challenging the current assumptions. Our work has direct implications for the elucidation of the QA biosynthesis pathway and the long-distance transport network from source to seed.
羽扇豆是很有前景的蛋白质作物，但它们的种子中会积累有毒的喹嗪生物碱（QAs），这使得其最终用途变得复杂。QAs在绿色器官（叶片、茎和豆荚）中合成，其中一部分在果实发育过程中被运输到种子中。生物合成和积累的确切部位尚不清楚；然而，有人提出叶肉细胞是来源，表皮细胞是储存部位。我们使用基于质谱成像（MSI）、激光捕获显微切割结合RNA测序，以及前体饲喂研究结合液相色谱-质谱联用和MSI，研究了窄叶羽扇豆（Lupinus angustifolius）生物合成器官中QA生物合成和积累的确切部位。我们发现，在种子中积累的QAs（“核心”QAs）在各组织中分布均匀；然而，它们的酯化形式主要积累在表皮中。令人惊讶的是，RNA测序显示表皮中有很强的生物合成基因表达，这在叶片中通过定量实时聚合酶链反应得到了证实。最后，使用稳定标记前体的饲喂研究表明，下部叶片表皮具有高度的生物合成能力。我们的结果表明，表皮是窄叶羽扇豆中QA生物合成的主要部位，这对当前的假设提出了挑战。我们的工作对阐明QA生物合成途径以及从来源到种子的长距离运输网络具有直接意义。
