Integrated transcriptome and chromatin accessibility dynamics in developing walnut kernels reveal regulators of oil accumulation

Walnuts are valued for their high lipid content, including polyunsaturated fatty acids. However, the molecular mechanisms driving oil biosynthesis during kernel development are not fully understood. To explore this, we constructed a comprehensive full-length transcriptome of walnut kernels across five developmental stages, identifying 221 candidate genes associated with oil biosynthesis, including 120 in fatty acid biosynthesis, 86 in triacylglycerol biosynthesis, and 15 in oil body formation. Integrating chromatin accessibility data with gene expression profiles, we found ten oil biosynthesis-related genes that showed coordinated changes in promoter accessibility and expression. Transcriptional regulatory network analysis further identified 23 hub transcription factors (TFs), including 14 putative negative regulators and 9 positive regulators of oil accumulation. Among these, JrERF93 as a member of AP2 family bound the promoter-accessible region of JrACC/CTα-5 and activated its expression in both yeast one-hybrid and dual-luciferase assay. Moreover, overexpression of JrERF93 in Arabidopsis thaliana significantly increased oil content and altered fatty acid composition in seeds. Collectively, these findings enhance our understanding the regulation of walnut oil biosynthesis and provide insights for strategies for genetic improvement. • Oil biosynthesis pathway was recharacterized using full-length walnut kernel transcriptome. • Regulatory dynamics during walnut oil accumulation were elucidated by chromatin accessibility profiling. • Twenty-three hub TFs involved in oil biosynthesis were revealed by walnut regulatory networks. • Enhancement of seed oil content by JrERF93 was demonstrated through Arabidopsis validation.