Comprehensive identification of bHLH transcription factors in Litsea cubeba reveals candidate gene involved in the monoterpene biosynthesis pathway

Litsea cubeba (Lour.) Person, an economically important aromatic plant producing essential oils, has lemon-like fragrance and 96.44-98.44% monoterpene contents. bHLH transcription factor plays an important role in plant secondary metabolism and terpene biosynthesis. In this study, we used bioinformatics to identify bHLH transcription factors in L. cubeba, 173 bHLH genes were identified from L. cubeba and divided these into 26 subfamilies based on phylogenetic analysis. The majority of bHLHs in each subfamily shared comparable structures and motifs. While LcbHLHs were unevenly distributed across 12 chromosomes, 10 tandem repeats were discovered. Expression profiles of bHLH genes in different tissues demonstrated that LcbHLH78 is a potential candidate gene for regulating monoterpene biosynthesis. LcbHLH78 and the terpene synthase LcTPS42 showed comparable expression patterns in various tissues and fruit development stages of L. cubeba. Subcellular localization analysis revealed that LcbHLH78 protein localizes to the nucleus, consistent with a transcription factor function. Importantly, transient overexpression of LcbHLH78 increased geraniol and linalol contents. Our research demonstrates that LcbHLH78 enhances terpenoid biosynthesis. This finding will be beneficial for improving the quality of L. cubeba and provides helpful insights for further research into the control mechanism of LcbHLH genes over terpenoid biosynthesis.