Function of R2R3-MYB Gene LcMYB113 on regulating anthocyanin biosynthesis in Loropetalum chinense var. rubrum postprint

Abstract: Anthocyanin plays one of the key factors in determining ornamental value of Loropetalum chinense var. rubrum. In order to investigate the molecular mechanisms of anthocyanin biosynthesis in L. chinense var. rubrum, an anthocyanin biosynthesis relative R2R3-MYB gene, named LcMYB113 with GenBank accession number OR344758, was cloned from L. chinense var. rubrum. The deduced amino acid sequence of LcMYB113 gene was analyzed by bioinformatics methods. The relative expression levels of LcMYB113 gene in leaves of L. chinense var. rubrum and L. chinense were tested by real-time quantitative RT-PCR method. The phenotypes of leaves and flowers of transgenic tobacco lines were recorded. The relative expression levels of anthocyanin biosynthetic structural genes in leaves of transgenic tobacco lines were examined. The results were as follows: (1) The open reading frame of LcMYB113 gene was 789 bp, encoding 262 amino acids. Multiple alignment analysis showed that the N-terminal of LcMYB113 contained a canonical R2R3 DNA binding domain and a bHLH transcript factor binding motif [D/E]Lx2[R/K]x3Lx6Lx3R. Two anthocyanin biosynthesis activator characteristic motifs ANDV and [K/R]P[Q/R]P[Q/R] were also found in the amino acid sequence of LcMYB113. Phylogenetic tree analysis indicated that LcMYB113 was closely related to anthocyanin specific R2R3-MYB subfamily transcription factors, including PsMYB58 of Paeonia suffruticosa and VvMYBA1 of Vitis vinifera. (2) The anthocyanin content and relative expression level of LcMYB113 gene in leaves of L. chinense var. rubrum were 7.4 and 101 times that of L. chinense respectively, which indicated that the relative expression levels of LcMYB113 gene was correlated with the anthocyanin content in leaves. (3) The LcMYB113 gene overexpression vector was constructed and transformed into tobacco variety WS38. Heterologous expression of LcMYB113 gene in tobacco induced anthocyanin accumulation in leaves and flowers of transgenic lines. Further more, compared with wide type tobacco line, transgenic lines had remarkably higher relative expression levels of anthocyanin biosynthetic structural genes, such as NtANS, NtDFR and NtCHS. In summary, this research results indicate that LcMYB113 can regulate the biosynthesis of anthocyanin, providing theoretical support for leaf color breeding of L. chinense var. rubrum.