NtMYC2a plays a "master switch" role in the regulation of JA-induced nicotine accumulation in tobacco.

• The biosynthesis and transportation of nicotine has been shown to be coordinately up-regulated by jasmonate (JA). MYC2, a member of basic helix-loop-helix (bHLH) transcription factor (TF) family, is well-documented as the core player in JA signaling pathway to regulate diverse plant developmental processes. Four MYC2 genes were found to be present in tobacco genome, NtMYC2a/2b and 1a/1b. In this study, we tested whether one of them: NtMYC2a acts as a "master switch" in the regulation of nicotine biosynthesis and transportation in tobacco plant. • We generated NtMYC2a knockout tobacco plants using CRISPR-Cas9 technique and analyzed the effect of NtMYC2a knockout on the expression of the nicotine biosynthetic genes (NtAO, NtQS, NtPMT1a, NtQPT2, NtODC2, NtMPO1, NtA622 and NtBBLa) and transport genes (NtMATE2 and NtJAT1), as well as the leaf accumulation of nicotine in the NtMYC2a knockout plants. • We found that all the nicotine biosynthetic and transport genes tested in this study were significantly down-regulated (more than 50% reduction compared with wild type control) in the NtMYC2a knockout plants. Meanwhile, the leaf nicotine content in the knockout plants was dramatically reduced by approximate 80% compared with the wild type control. • The results from this study clearly shows that NtMYC2a acts as a "master switch" to coordinate the JA-induced nicotine accumulation in tobacco plant and suggests that NtMYC2a may play an important role in tobacco plant's nicotine-mediated defense against herbivory.