Promoter variations in a homeobox gene, BnA10.LMI1, determine lobed leaves in rapeseed (Brassica napus L.)

Limin Hu, Hao Zhang, Qingyong Yang, Qingwei Meng, Shaoqing Han, Chinedu Charles Nwafor, Muhammad Hafeez Ullah Khan, Chuchuan Fan, Yongming Zhou

BnA10.LMI1 positively regulates the development of leaf lobes in Brassica napus, and cis-regulatory divergences cause the different allele effects. Leaf shape is an important agronomic trait, and large variations in this trait exist within the Brassica germplasm. The lobed leaf is a unique morphological characteristic for Brassica improvement. Nevertheless, the molecular basis of leaf lobing in Brassica is poorly understood. Here, we show that an incompletely dominant locus, BnLLA10, is responsible for the lobed-leaf shape in rapeseed. A LATE MERISTEM IDENTITY1 (LMI1)-like gene (BnA10.LMI1) encoding an HD-Zip I transcription factor is the causal gene underlying the BnLLA10 locus. Sequence analysis of parental alleles revealed no sequence variations in the coding sequences, whereas abundant variations were identified in the regulatory region. Consistent with this finding, the expression levels of BnLMI1 were substantially elevated in the lobed-leaf parent compared with its near-isogenic line. The knockout mutations of BnA10.LMI1 gene were induced using the CRISPR/Cas9 system in both HY (the lobed-leaf parent) and J9707 (serrated leaf) genetic backgrounds. BnA10.LMI1 null mutations in the HY background were sufficient to produce unlobed leaves, whereas null mutations in the J9707 background showed no obvious changes in leaf shape compared with the control. Collectively, our results indicate that BnA10.LMI1 positively regulates the development of leaf lobes in B. napus, with cis-regulatory divergences causing the different allelic effects, providing new insights into the molecular mechanism of leaf lobe formation in Brassica crops.