Telomere-to-telomere genome assembly of Plukenetia volubilis reveals a positive regulatory role of PvMYB14 on linolenic acid biosynthesis

Abstract

Alpha-linolenic acid (ALA) is an essential polyunsaturated fatty acid for the human body, possessing a broad spectrum of pharmacological effects. Plukenetia volubilis has garnered widespread cultivation interest for its exceptionally high ALA content. However, the lack of a high-quality reference genome has constrained in-depth genetic and functional research on this species. Here, we present the first telomere-to-telomere gap-free reference genome for P. volubilis. The assembled 665.29 Mb genome spans 29 chromosomes and demonstrates exceptional continuity and completeness (contig N50 = 22.88 Mb). The results of evolutionary analyses showed that the species divergence time of P. volubilis are significantly later than those of other oil crops. Moreover, comparative genomics revealed that P. volubilis underwent a lineage-specific whole-genome triplication around 39.83–48.39 million years ago, coinciding with its divergence from Ricinus communis. To elucidate the genetic basis of its high ALA content, we integrated comparative lipidomic, transcriptomic and genomic analyses, identifying the transcription factor PvMYB14 as a key regulator. The PvMYB14 localizes to the nucleus and directly activates the expression of PvFAD3, by binding to a specific CNGTTA-box motif. We used electrophoretic mobility shift assay to identify conserved nucleotide sites of the CNGTTA-box, with MT4 and MT6 mutations abolishing binding to PvMYB14. Heterologous expression demonstrated that PvMYB14 simultaneously increases total seed oil yield and the proportional content of ALA. This high-quality genome assembly and the elucidated PvMYB14-PvFAD3 regulatory module provide a foundational resource and potential target for the metabolic engineering of high-quality oil.