Identification of PrMYB96 as a positive regulator of PrPDAT2 in oil biosynthesis of Paeonia rockii

The seeds of Paeonia rockii are characterized by exceptionally elevated levels of polyunsaturated fatty acids (PUFAs, >90 %), with α-linolenic acid (ALA) comprising over 40 % of the total fatty acids (FAs). ALA is critical for lipid metabolism, neural development, as well as in the production of biofuels and anti-aging applications. Our previous study identified diacylglycerol acyltransferase2 (PrPDAT2), a phospholipid: specifically transferring ALA to promote lipid deposition. However, the transcriptional mechanisms of seed oil biosynthesis remain largely unknown. Here, we identified PrMYB96, an R2R3-MYB factor was shown to act as a transcriptional activator of PrPDAT2 . Seed-preferential expression of PrMYB96 , coupled with its nuclear localization, suggests a promotive role in oil deposition. Overexpression of PrMYB96 in transgenic Nicotiana tabacum L. seeds upregulated oil biosynthesis and changed in FA composition, while its suppression in P. rockii seeds and embryos led to reduced total FA and ALA content. Furthermore, PrMYB96 is transcriptionally activated by nuclear factor Y, subunit C2 ( PrNF-YC2) . Overall, the study highlights the pivotal role of PrMYB96 in seed oil biosynthesis, providing new insights into its regulatory network and potential for genetic improvement in oil-producing species. • Revealing the regulatory role of PrMYB96 in seed oil biosynthesis by activating PrPDAT2 in Paeonia rockii .. • Providing a genetic basis for enhancing α-linolenic acid (ALA) accumulation via PrMYB96 in Paeonia rockii . • Demonstrating the role of PrMYB96 in oil deposition using the TTAES embryo-derived seedling system. • Uncovering that PrNF-YC2 transcriptionally activates PrMYB96 to regulate seed oil biosynthesis.