CcMYB6-1 and CcbHLH1, two novel transcription factors synergistically involved in regulating anthocyanin biosynthesis in cornflower
CcMYB6-1 and CcbHLH1, two novel transcription factors synergistically involved in regulating anthocyanin biosynthesis in cornflower
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CcMYB6-1和CcbHLH1，两种新型转录因子协同调控矢车菊花青素生物合成
DOI：
10.1016/j.plaphy.2020.03.024
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发表时间：
2020-06-01
影响因子：
6.5
通讯作者：
Dai, Silan
中科院分区：
文献类型：
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作者：
Deng, Chengyan;Wang, Jiaying;Dai, Silan
矢车菊花色苷的合成受一系列基因的调控，但其转录调控机制尚不清楚。本研究以矢车菊为材料，研究了同一花瓣从白色到紫色再到蓝色的发育过程。色素分析表明，花青苷积累显着增加与花瓣颜色的发展。随后，从上述三个颜色区域构建九个文库用于RNA-seq，并通过从头组装获得105，506个unigenes。3个颜色区域的差异表达基因在苯丙素类生物合成和类黄酮生物合成途径中显著富集，共挖掘和分析了46个参与该过程的生物合成基因。通过与拟南芥同源物的系统发育分析，筛选出4个R2 R3-CcMYBs和1个CcbHLH 1，分别聚为亚群4和亚群6，聚为IIIf亚群。进一步分离黄烷酮3-羟化酶（CcF 3 H）和二氢黄酮醇4-还原酶（CcDFR）的启动子，以研究其上游调控机制。双荧光素酶检测和瞬时表达结果表明，CcMYB 6 -1显著上调了上述两个启动子的活性，促进了花青素的积累，与CcbHLH 1共渗后，其活性明显增强。此外，酵母双杂交和双分子荧光互补试验均表明这两种激活剂之间存在蛋白质-蛋白质相互作用。基于这些获得的结果，它揭示了CcMYB 6 -1和CcbHLH 1是两个新的转录因子协同参与调控花青素的生物合成。本研究为深入了解矢车菊花色素苷积累的调控机制提供了理论依据。
Anthocyanins in cornflower (Centaurea cyanus) is catalysed by a set of biosynthesis genes, however, the potential mechanism of transcriptional regulation remains unclear. In the present study, we traced the dynamic changes of petal colour development from white to violet and finally to blue on the same petal in cornflower. Pigment analysis showed that anthocyanin accumulation dramatically increased with petal colour development. Subsequently, nine libraries from above three colour regions were constructed for RNA-seq and 105,506 unigenes were obtained by de novo assembling. The differentially expressed genes among three colour regions were significantly enriched in the phenylpropanoid biosynthesis and flavonoid biosynthesis pathways, leading to the excavation and analysis of 46 biosynthesis genes involved in this process. Furthermore, four R2R3-CcMYBs clustered into subgroup 4 or subgroup 6 and one CcbHLH1 clustered into IIIf subgroup were screened out by phylogenetic analysis with Arabidopsis homologues. The promoters of flavanone 3-hydroxylase (CcF3H) and dihydroflavonol 4-reductase (CcDFR) were further isolated to investigate upstream regulation mechanism. CcMYB6-1 significantly upregulated the activity of above two promoters and stimulated anthocyanin accumulation by dual luciferase assay and transient expression in tobacco leaves, and its activity was obviously enhanced when co-infiltrated with CcbHLH1. Moreover, both yeast two-hybrid and bimolecular fluorescence complementation assays indicated the protein-protein interaction between these two activators. Based on these obtained results, it reveals that CcMYB6-1 and CcbHLH1 are two novel transcription factors synergistically involved in regulating anthocyanin biosynthesis. This study provides insights into the regulatory mechanism of anthocyanin accumulation in cornflower.