A rare glycoalkaloid biosynthetic pathway from wild potato unlocks genetic routes to Colorado potato beetle resistance.

Abstract

Plants in the family <i>Solanaceae</i> produce steroidal glycoalkaloids (SGAs), specialized metabolites that are toxic to consumers while providing protection against herbivores. The Colorado potato beetle (CPB) is a major foliar pest of potatoes, and the wild species <i>Solanum chacoense</i> PI458310 displays resistance through the accumulation of the specific SGAs leptine I/II, the C-23-acetoxylated derivatives of α-chaconine/α-solanine. Although these leptines and their presumed C-23-hydroxylated precursors, leptinine I/II, have been recognized for decades, their biosynthetic routes have remained unresolved. In this study, we show that leptinines and leptines do not originate from direct modificfation of α-chaconine/α-solanine, but instead arise from earlier transformations of spirosolane-type precursors. We identified <i>Sc23DOX</i>, a spirosolane C-23 hydroxylase, and <i>Sc23ACT</i>, a 23-<i>O</i>-acetyltransferase, and demonstrated in vitro that these enzymes convert α/β-solamarine into 23-hydroxy and 23-<i>O</i>-acetyl intermediates, which are subsequently processed into leptinines and leptines by the ring-rearrangement enzymes Dioxygenase for Potato Solanidane synthesis (DPS), Reductase for Potato Glycoalkaloid biosynthesis 1 (RPG1), and RPG2. Notably, these C-23 hydroxylation and O-acetylation reactions parallel the initial steps of α-tomatine detoxification during fruit ripening in tomato (<i>Solanum lycopersicum</i>). Moreover, when <i>Sc23DOX</i> and <i>Sc23ACT</i> were expressed heterologously in cultivated potato, the plants produced leptinines and leptines de novo. Genomic and transcriptomic analyses further revealed that many cultivated potatoes (<i>Solanum tuberosum</i>) retain apparently functional 23DOX and 23ACT orthologues, and that expression of 23DOX alone can restore leptine biosynthesis in certain cultivars. Collectively, these findings clarify the leptinine/leptine biosynthetic pathway, illustrate the co-option of a detoxification-related module into a defense system, and provide a genetic basis for breeding or engineering CPB-resistant potatoes.