biologia plantarum

Biologia plantarum 58:436-444, 2014 | DOI: 10.1007/s10535-014-0411-z

Molecular cloning, characterization, and expression analysis of LeMYB1 from Lithospermum erythrorhizon

H. Zhao^1,2, S. K. Baloch^1,3, L. R. Kong¹, W. J. Zhang¹, A. L. Zou¹, X. M. Wang¹, J. L. Qi^1,4,*, Y. H. Yang^1,*

¹ State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing, P.R. China

² Engineering Technology Research Center of Anti-Aging Chinese Herb, School of Life Sciences, Fuyang Normal College, Fuyang, P.R. China

³ Department of Biotechnology FCPD, Sindh Agriculture University, Tandojam, Pakistan

⁴ Huaian High-Tech Research Institute, Nanjing University, Huaian, P.R. China

MYB transcription factors (TFs) are known to have important functions in regulating the biosynthesis of secondary metabolites in plants. In this study, LeMYB1, a member of the MYB gene family of Lithospermum erythrorhizon, was cloned via the rapid amplification of cDNA ends. The alignment of the predicted translations of LeMYB1 with other MYB proteins revealed that LeMYB1 contained an N-terminal R2R3 repeat and a high degree of amino acid identity to NtMYBJS1 which is involved in jasmonic acid signalling and phenylpropanoid biosynthetic pathway regulation. To determine the expression pattern of LeMYB1, its promoter was cloned and the sequence analysis was performed. The results revealed a number of potential regulatory motifs related to tissue-specific gene expression and abiotic and biotic stress responses. Real-time PCR results suggest that LeMYB1 was induced transiently during the early stage when L. erythrorhizon cells were transferred from a B5 growth medium to a M9 production medium for shikonin formation. Exogenous methyl jasmonate (MeJA), an effective inducer of shikonin biosynthesis, induced the rapid LeMYB1 expression. In contrast, a treatment with ibuprofen (IBU), an inhibitor of jasmonate biosynthesis, significantly inhibited the LeMYB1 expression. Another inhibitor of shikonin formation, 2,4-dichlorophenoxyacetic acid (2,4-D), also markedly repressed the expression of LeMYB1. Tissue-specific expression analysis showed that LeMYB1 mRNA was predominantly accumulated in roots where shikonin was synthesized. Thus, the LeMYB1 gene may be a valuable member of the R2R3-MYB family in L. erythrorhizon and is possibly involved in the regulation of shikonin biosynthesis.

Keywords: 2,4-dichlorophenoxyacetic acid; ibuprofen; methyl jasmonate; R2R3 repeat; RACE; real-time PCR; shikonin

Subjects: Myb transcription factor; 2.4-dichlorophenoxyacetic acid; ibuprofen; methyl jasmonate; RACE; shikonin; phylogenetic tree

Received: May 8, 2013; Revised: November 28, 2013; Accepted: December 4, 2013; Published: September 1, 2014  Show citation