A Novel R2R3-MYB Transcription Factor Gene AaMYB6 Involved in Anthocyanin Biosynthesis in Anthurium andraeanum

doi:10.16420/j.issn.0513-353x.2021-0476

Acta Horticulturae Sinica ›› 2021, Vol. 48 ›› Issue (10): 1859-1872.doi: 10.16420/j.issn.0513-353x.2021-0476

• Research Papers • Previous Articles Next Articles

A Novel R2R3-MYB Transcription Factor Gene AaMYB6 Involved in Anthocyanin Biosynthesis in Anthurium andraeanum

LI Chonghui¹^,³, YANG Guangsui¹^,³, ZHANG Zhiqun¹^,³, YIN Junmei²^,³^,^*()

¹Tropical Crops Genetic Resources Institute,Chinese Academy of Tropical Agricultural Sciences;Key Laboratory of Crop Gene Resources and Germplasm Enhancement in Southern China,Ministry of Agriculture,Haikou 571101,China
²Haikou Experimental Station,Chinese Academy of Tropical Agricultural Sciences,Haikou 571101,China
³The Engineering Technology Research Center of Tropical Ornamental Plant Germplasm Innovation and Utilization,Danzhou,Hainan 571737,China

Received:2021-07-19 Revised:2021-09-02 Online:2021-10-25 Published:2021-11-01
Contact: YIN Junmei E-mail:yinjunmei2004@163.com

Abstract

Abstract:

A new R2R3-MYB gene AaMYB6 with GenBank accession number MZ171064 was isolated from the spadix of Anthurium andraeanum‘Pink Champion’based on the information from previous transcriptome data for identification of transcription factor genes associated with anthocyanin biosynthesis. The function of AaMYB6 was analyzed through the assays such as amino acid sequence alignment,phylogenetic tree analysis,subcellular localization,overexpression in tobacco,qRT-PCR,yeast two-hybrid assay（Y2H）,and bimolecular fluorescence complementation（BiFC）. The results suggested that AaMYB6 showed homology with previously identified AaMYB1 from anthurium and belong to C1 subgroup R2R3-MYBs,represented by Zea may C1,that regulate anthocyanin biosynthesis in monocots. AaMYB6 was proved to be nucleus-localized protein. Overexpression of AaMYB6 in tobacco markedly activated the expression of the enzyme genes NtF3′H,NtDFR,NtANS,NtUFGT,specifically involved in anthocyanin biosynthesis in filaments and promoted the accumulation of anthocyanin pigments. AaMYB6 was mainly expressed in the spadix of‘Rapido’. And the expression of AaMYB6 was closely linked to anthocyanin accumulation in the spadix of various cultivars. The interaction between AaMYB6 and the AabHLH1 protein,which was identified previously to regulate the biosynthesis of anthocyanin and proanthocyanidin in anthurium,was confirmed by Y2H and BiFC assays. The results indicated that AaMYB6 play a role in the regulation of anthocyanin biosynthesis in anthurium.

Key words: Anthurium andraeanum, MYB transcription factor, anthocyanin, heterologous overexpression, transcriptional regulation, gene function

CLC Number:

S682.14

LI Chonghui, YANG Guangsui, ZHANG Zhiqun, YIN Junmei. A Novel R2R3-MYB Transcription Factor Gene AaMYB6 Involved in Anthocyanin Biosynthesis in Anthurium andraeanum[J]. Acta Horticulturae Sinica, 2021, 48(10): 1859-1872.

Figures/Tables 7

Fig. 1 Sequences alignment of AaMYB6 from anthurium with known R2R3-MYBs involved in anthocyanin biosynthesis A:The red box indicates the bHLH interacting motif;the green box indicates the different conserved“DNEI”motif for monocots R2R3-MYB;The blue box indicates the C-terminal-conserved motif KAX[K/R]C[S/T] for anthocyanin-regulating MYBs of monocots. Aa:Anthurium andraeanum;Zm:Zea mays;Pe:Phalaenopsis equestris;Fh:Freesia hybrid.

Fig. 2 Phylogenic tree of AaMYB6 from anthurium with known R2R3-MYBs involved in anthocyanin biosynthesis * Both correlated with anthocyanin and proanthocyanidin.

Fig. 3 Subcellular location of AaMYB6 in protoplasts of Arabidopsis

Fig. 4 The expression of AaMYB6 and the anthocyanin content in anthurium P < 0.05.

Fig. 5 The phenotypes and the content of anthocyanin of AaMYB6 transgenic tobacco（A）and detection of gene expression（B,C） WT:Wild type tobacco plants;L1-L3:Three AaMYB6 overexpression T1 tobacco lines,and there were two plants of each line（-1,-2）.t-test [Comparison between transgenic lines and wild type plants（A,D）or between different tissue（B,C）],** α = 0.01.

Fig. 6 The relative expression of enzyme genes in petals and filaments of transgenic tobacco plants FC:Fold changes against the WT;L1-L3:Three AaMYB6 overexpression T1 tobacco lines,there were two plants of each line（-1,-2）.

Fig. 7 Interactions between AaMYB6 and AabHLH1 detected using the Y2H and BiFC assays A:Y2H Gold yeast cells containing plasmids were grown on double- and quadruple-selection SD media. The X-α-gal assay was performed to confirm the positive interactions;BD:pGBKT7 empty plasmid;AD:pGADT7 empty plasmid;AD-AaMYB6:pGAD-AaMYB6 recombinant plasmid;BD-AabHLH1:pGBK-AabHLH1 recombinant plasmid;B:Bimolecular fluorescence complementation visualization of the AaMYB6 and AabHLH1 interaction in the in protoplasts of Arabidopsis.

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A Novel R2R3-MYB Transcription Factor Gene AaMYB6 Involved in Anthocyanin Biosynthesis in Anthurium andraeanum

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