The LibHLH22 and LibHLH63 from Lilium ‘Siberia’ Can Positively Regulate Volatile Terpenoid Biosynthesis

Abstract

Basic helix-loop-helix (bHLH) transcription factors (TFs) play irreplaceable roles in plant growth and development, especially in plant secondary metabolism. However, the functions of most bHLH TFs in Lilium ‘Siberia’ are still unknown, especially their roles in regulating floral fragrance. In this study, two bHLH TFs in lily, i.e., LibHLH22 and LibHLH63, were identified and functionally characterized. A bioinformatics analysis revealed that LibHLH22 and LibHLH63 were unstable proteins. Subcellular localization demonstrated that LibHLH22 and LibHLH63 proteins were in the cell nucleus. Quantitative real-time PCR showed that the highest expression level of LibHLH22 was at the initial flowering stage and in the stigma, and the highest expression level of LibHLH63 was at the budding stage and in the filaments. The results of transient overexpression and virus-induced gene silencing (VIGS) of LibHLH22 and LibHLH63 in lily petals showed that these two transcription factors significantly promoted the expression of LiDXR and LiTPS2, and thus, markedly enhanced the release of floral fragrance. Our results indicated that LibHLH22 and LibHLH63 could effectively regulate the fragrance of Lilium ‘Siberia’, laying the foundation for fragrance breeding and improving the terpenoid transcriptional regulatory pathway.