Isolation and characterization of R2R3-MYB and basic helix-loop-helix (bHLH) transcription factors involved in anthocyanin biosynthesis in tulip tepals

摘要

Anthocyanin is a kind of secondary metabolites that contribute to flower color. Transcription factors R2R3-MYB and bHLH (basic helix-loop-helix) are required to regulate the transcription of structural genes involved in anthocyanin biosynthesis. In this study, full-length cDNAs of four R2R3-MYB (TfMYB2, TfMYB3, TfMYB4, TfMYB5) and two bHLH (TfbHLH1, TfBHLH2) genes were first cloned from tepals of Tulipa fosteriana 'Albert heijn' by RACE method. Four MYB genes belong to AN2 subgroup possessing high homology with lily MYBs, and bHLH genes share high homology with lily bHLHs. TfMYB3, TfMYB4, TfMYB5, and TfbHLH1 expressed highly in tulip tepal, while TfMYB2 and TfbHLH2 highly in stem and leaf. The spatial expression patterns of above four TFs also coincided well with some structural genes involving in the anthocyanin biosynthesis. The transcripts of TfMYB3, TfMYB4, TfMYB5, and TfbHLH1 constantly increased along with flower development, which were consistent with the expression of structural genes and accompanied by the anthocyanin accumulation. Y2H analysis revealed that TfMYB3, TfMYB4, or TfMYB5 interacted with TfbHLH1. The transient expression assay suggested that TfMYB5 activated the promoter of TfCHS1, while TfMYB3 or TfMYB4 promoted the promoter activity of TfANS1. Meanwhile, no significant promoting effect of TfbHLH1 was observed; however, the enhancing effect of TfMYBs on the promoters was much promoted when interacted with TfbHLH1. Taken together, our findings strongly suggested that TfMYB3, TfMYB4, TfMYB5, and TfbHLH1 were definitely participated in the regulating of anthocyanin biosynthesis in tepals of tulip.