The oil yield of Brassica napus is determined by both seed yield and oil content. A previous study showed that strong RNA interference, designed to simultaneously knock down all four BnAP2 paralogs, reduced seed yield due to abnormal floral development in B. napus. However, the function of individual, specific BnAP2 paralogs in seed oil production and the underlying mechanisms remain unclear. Here, we found that all four BnAP2 paralogs present in the genome of the B. napus ‘K407’ inbred line were highly expressed in flowers and developing seeds; however, BnaA01.AP2 was minimally expressed in floral tissues. Interestingly, BnaA01.AP2 knockout mediated by CRISPR/Cas9 resulted in significant increases in both seed yield and oil content, thereby increasing overall seed oil yield without detectable negative effects on the other examined agronomic traits. Furthermore, we demonstrated that BnaA01.AP2 repressed oil accumulation by directly downregulating BnaA09.WRI1, BnaA03.BCCP1, BnaA09.L1L, and BnaC09.L1L and indirectly regulating a series of key genes involved in glycolysis, fatty acid biosynthesis, and triacylglycerol assembly during B. napus seed development. Our research not only provides insights into the regulatory mechanisms of seed oil accumulation but also provides promising genetic resources and germplasm for breeding cultivars with higher seed oil yield in B. napus.

Rapeseed (Brassica napus L.) is a global oil crop. Salinity stress impedes the growth of rapeseed, especially during seed germination. The key genes mediating salinity stress response during seed germination in B. napus remain largely unknown. Here, we found that all six paralogs of C2H2 zinc finger transcription factor WIP DOMAIN PROTEIN 2 (BnaWIP2) showed increased expression during the initial 12 hours of germination, and expression was further enhanced by salinity stress. Under NaCl treatment, knocking out all six BnaWIP2 paralogs in B. napus led to significantly reduced germination, while overexpression of BnaC06.WIP2 promoted germination. Transcriptomic analysis revealed that BnaC06.WIP2 downregulated a series of genes related to abscisic acid (ABA) biosynthesis and signaling, among which BnaA05.NCED3, BnaC04.ABI5-2, BnaA03.EM6, and BnaA05.EM6 were directly repressed by BnaC06.WIP2. Further analysis showed that in germinating seeds, BnaC06.WIP2 was induced by ABA and in turn restrained ABA production, indicating that BnaC06.WIP2 forms a negative feedback loop with ABA to promote seed germination under salinity stress in B. napus. Collectively, these results enhance our understanding of the novel function of BnaWIP2 and provide valuable genetic resources for breeding salinity-tolerant rapeseed varieties.

Flower development and plant architecture determine the efficiency of mechanized harvest and seed yield in Brassica napus. Although TERMINAL FLOWER 1 (AtTFL1) is a regulator of flower development in Arabidopsis thaliana, the function and regulatory mechanism of TFL1 orthologs in B. napus remains unclear. Six BnTFL1 paralogs in the genome of the B. napus inbred line ‘K407’ showed steadily increasing expression during vernalization. CRISPR/Cas-induced mutagenesis of up to four BnTFL1 paralogs resulted in early flowering and alteration of plant architecture, whereas seed yield was not altered in BnTFL1 single, double, or triple mutants. Six BnTFL1 paralogs, but not BnaA02.TFL1, showed an additive and conserved effect on regulating flowering time, total and terminal flower number, and plant architecture. BnaA10.TFL1 regulates flower development by interacting with BnaA08.FD through the protein BnaA05.GF14nu, resulting in the transcriptional repression of floral integrator and floral meristem identity genes. These findings about the regulatory network controlling flower development and plant architecture present a promising route to modifying these traits in B. napus.