AUTHOR=Dong Wen , Tang Linfei , Peng Yali , Qin Yuzhi , Lin Yuan , Xiong Xingyao , Hu Xinxi 

TITLE=Comparative transcriptome analysis of purple-fleshed sweet potato and its yellow-fleshed mutant provides insight into the transcription factors involved in anthocyanin biosynthesis in tuberous root

JOURNAL=Frontiers in Plant Science

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/plant-science/articles/10.3389/fpls.2022.924379

DOI=10.3389/fpls.2022.924379

ISSN=1664-462X

ABSTRACT=<p>In various plant species, many transcription factors (TFs), such as MYB, bHLH, and WD40, have been identified as regulators of anthocyanin biosynthesis in underground organs. However, the regulatory elements of anthocyanin biosynthesis in the tuberous roots of sweet potato have not been elucidated yet. Here, we selected the purple-fleshed sweet potato cultivar “Zhezi1” (ZZ<italic><sup>P</sup></italic>) and its spontaneous yellow-fleshed mutant “Xinli” (XL<italic><sup>Y</sup></italic>) to investigate the regulatory mechanism of the anthocyanin biosynthesis in the tuberous roots of sweet potato. By analyzing the <italic>IbMYB1</italic> genotype in ZZ<italic><sup>P</sup></italic> and XL<italic><sup>Y</sup></italic>, we found that the <italic>IbMYB1-2</italic>, a MYB TF involved in anthocyanin biosynthesis, was missing in the XL<italic><sup>Y</sup></italic> genome, which might lead to an extreme decrease in anthocyanins in XL<italic><sup>Y</sup></italic>. A comparative transcriptome analysis of ZZ<italic><sup>P</sup></italic> and XL<italic><sup>Y</sup></italic> was conducted to find the TFs involved in anthocyanin biosynthesis in ZZ<italic><sup>P</sup></italic> and XL<italic><sup>Y</sup></italic>. The anthocyanin structural genes were significantly enriched among the differentially expressed genes. Moreover, one MYB activator (<italic>IbMYB1</italic>), one bHLH (<italic>IbbHLH2</italic>), three WRKY activator candidates (<italic>IbWRKY21</italic>, <italic>IbWRKY24</italic>, and <italic>IbWRKY44</italic>), and two MYB repressors (<italic>IbMYB27</italic> and <italic>IbMYBx-ZZ</italic>) were highly expressed in ZZ<italic><sup>P</sup></italic> accompanied with anthocyanin structural genes. We also tested the expression of these TFs in six purple- and two orange-fleshed sweet potato cultivars. Interestingly, most of these TFs were significantly positively correlated with anthocyanin contents in these cultivars. The function of the anthocyanin biosynthesis repression of <italic>IbMYB27</italic> and <italic>IbMYBx-ZZ</italic> was verified through transient co-transformation with <italic>IbMYB1</italic> into tobacco leaves. Further functional verification of the above TFs was conducted by Y2H, BiFC, and dual-luciferase assays. These tests showed that the MYB-bHLH-WD40/MYB-bHLH-WD40-WRKY complex activated the promoter of anthocyanin structural gene <italic>IbDFR</italic> and promoters for <italic>IbWRKY44</italic>, <italic>IbMYB27</italic>, and <italic>IbMYBx-ZZ</italic>, indicating reinforcement and feedback regulation to maintain the level of anthocyanin accumulation in the tuberous roots of purple-fleshed sweet potato. These results may provide new insights into the regulatory mechanism of anthocyanin biosynthesis and accumulation in underground organs of sweet potatoes.</p>