Transcriptional Regulation of Anthocyanin Synthesis by MYB-bHLH-WDR Complexes in Kiwifruit (<i>Actinidia chinensis</i>)

Abstract

The anthocyanin synthetic pathway is regulated centrally by an MYB-bHLH-WD40 (MBW) complex. Anthocyanin pigmentation is an important fruit quality trait in red-fleshed kiwifruit; however, the underlying regulatory mechanisms involving the MBW complex are not well understood. In this study, one R2R3MYB (<i>AcMYBF110</i> expressed in fruit characteristically), one bHLH (<i>AcbHLH1</i>), two upstream regulators of <i>AcbHLH1</i> (<i>AcbHLH4</i> and <i>AcbHLH5</i>), and one WDR (<i>AcWDR1</i>) are characterized as being involved in the regulation of anthocyanin synthesis in kiwifruit. <i>AcMYBF110</i> plays an important role in the regulation of anthocyanin accumulation by specifically activating the promoters of several anthocyanin pathway genes including <i>AcCHS</i>, <i>AcF3</i>'<i>H</i>, <i>AcANS</i>, <i>AcUFGT3a</i>, <i>AcUFGT6b</i>, and <i>AcGST1</i>. Coexpression of <i>AcbHLH1</i>, <i>AcbHLH4</i>, or <i>AcbHLH5</i> together with <i>AcMYBF110</i> induces much greater anthocyanin accumulation in both tobacco leaves and in <i>Actinidia arguta</i> fruit compared with <i>AcMYBF110</i> alone. Moreover, this activation is further enhanced by adding <i>AcWDR1</i>. We found that both AcMYBF110 and AcWDR1 interact with all three AcbHLH factors, while AcMYBF110 also interacts with AcWDR1 to form three different MBW complexes that have different regulatory roles in anthocyanin accumulation of kiwifruit. The AcMYBF110-AcbHLH1-AcWDR1 complex directly targets the promoters of anthocyanin synthetic genes. Other features of the regulatory pathways identified include promotion of <i>AcMYBF110</i>, <i>AcbHLH1,</i>and <i>AcWDR1</i> activities by this MBW complex, providing for both reinforcement and feedback regulation, whereas the AcMYBF110-AcbHLH4/5-AcWDR1 complex is indirectly involved in the regulation of anthocyanin synthesis by activating the promoters of <i>AcbHLH1</i> and <i>AcWDR1</i> to amplify the regulation signals of the first MBW complex.