BnA10.RCO, a homeobox gene, positively regulates leaf lobe formation in Brassica napus L.

KEY MESSAGE: BnA10.RCO positively regulates the development of leaf lobes in Brassica napus, and cis-regulatory divergences cause the different allele effects. The functional importance of lobed leaves in rapeseed (Brassica napus L.) has been identified with potential advantages for high-density planting and hybrid production. Our previous studies indicated that the tandemly duplicated LMI1-like genes BnA10. RCO and BnA10.LMI1 are candidate genes of an incompletely dominant locus, which is responsible for the lobed-leaf shape in rapeseed. We provided strong evidence that BnA10.LMI1 positively regulates leaf lobe formation. Here, we show that BnA10.RCO is a nucleus-specific protein, encoding an HD-ZIP I transcription factor, which is responsible for the lobed-leaf shape in rapeseed. Sequence analysis of parental alleles revealed that no vital sequence variation was detected in the coding sequence of BnA10.RCO, whereas abundant variations were identified in the regulatory regions. Consistent with this finding, the expression level of BnRCO was substantially elevated in the lobed-leaved parent HY compared with its near-isogenic line. Moreover, the altered expression of BnA10.RCO in transgenic lines showed a positive connection with leaf complexity without a substantial change in BnLMI1 transcript level. Furthermore, CRISPR/Cas9-induced null mutations of BnA10.RCO in the lobed-leaved parent HY were sufficient to produce an unlobed leaf without alteration in BnLMI1 transcript level. Our results indicate that BnA10.RCO functions together with BnA10.LMI1 to positively determine the lobed-leaf development, providing a fundamental basis for crop improvement by targeting leaf shape in rapeseed.