CtCYP71A1 promotes drought stress tolerance and lignin accumulation in safflower and Arabidopsis

Lignin is a complex phenolic polymer that forms interwoven networks in the cell wall and is crucial to plant development and stress responses. The root tissues of safflower contain a high concentration of lignin, which may contribute to drought regulation mechanism in safflower. Cytochrome P450 is the largest oxidase family, which is often dubbed with a significant role in plant metabolism. However, little is known about the molecular mechanism of CYP450-mediated regulation of drought stress tolerance in safflower roots. In this study, a putative CtCYP71A1 gene from safflower was identified by combining genome-wide and transcriptomic analysis. In safflower, a total of 16 CtCYP71A enzyme coding genes were identified. The conserved structural topology of CtCYP71As reveals several signature motifs that are crucial to plant growth and stress response. The expression of CtCYP71A1 was shown to be diversely expressed in different tissues, however, it was significantly up-regulated in roots under normal condition. Similarly, in response to PEG stress, the expression of CtCYP71A1 in roots gradually increased, whereas it was upregulated initially and subsequently downregulated with ABA, GA3, and SA stress. In addition, the phenotypic analysis and root morphology of CtCYP71A1 overexpressed Arabidopsis lines showed enhanced drought tolerance with improved lignin content than that of wild-type and antisense plants, which was further confirmed by expression analysis, and various physiological indexes. Similarly, overexpression of CtCYP71A1 resulted in the positive regulation of several other genes involved in lignin biosynthesis compared to WT and antisense plants. Furthermore, drought sensitivity was observed in safflower roots with CtCYP71A1 virus-induced gene silencing assay. Together, these findings revealed important insights into the crucial role of CtCYP71A1 in drought tolerance via lignin accumulation in safflower.