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      Anthocyanin Accumulation in the Leaves of the Purple Sweet Potato ( Ipomoea batatas L.) Cultivars

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          Abstract

          Sweet potato anthocyanins are water-soluble pigments with many physiological functions. Previous research on anthocyanin accumulation in sweet potato has focused on the roots, but the accumulation progress in the leaves is still unclear. Two purple sweet potato cultivars (Fushu No. 23 and Fushu No. 317) with large quantities of anthocyanin in the leaves were investigated. Anthocyanin composition and content were assessed with ultra-performance liquid chromatography diode-array detection (UPLC-DAD) and ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS), and the expressions of genes were detected by qRT-PCR. The two cultivars contained nine cyanidin anthocyanins and nine peonidin anthocyanins with an acylation modification. The acylation modification of anthocyanins in sweet potato leaves primarily included caffeoyl, p-coumaryl, feruloyl, and p-hydroxy benzoyl. We identified three anthocyanin compounds in sweet potato leaves for the first time: cyanidin 3- p-coumarylsophoroside-5-glucoside, peonidin 3- p-coumarylsophoroside-5-glucoside, and cyanidin 3-caffeoyl- p-coumarylsophoroside-5-glucoside. The anthocyanidin biosynthesis downstream structural genes DFR4, F3H1, anthocyanin synthase ( ANS), and UDP-glucose flavonoid 3-O-glucosyltransferase ( UFGT3), as well as the transcription factor MYB1, were found to be vital regulatory genes during the accumulation of anthocyanins in sweet potato leaves. The composition of anthocyanins (nine cyanidin-based anthocyanins and nine peonidin-based anthocyanins) in all sweet potato leaves were the same, but the quantity of anthocyanins in leaves of sweet potato varied by cultivar and differed from anthocyanin levels in the roots of sweet potatoes. The anthocyanidin biosynthesis structural genes and transcription factor together regulated and controlled the anthocyandin biosynthesis in sweet potato leaves.

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          TT8 controls its own expression in a feedback regulation involving TTG1 and homologous MYB and bHLH factors, allowing a strong and cell-specific accumulation of flavonoids in Arabidopsis thaliana.

          The control of TT8 expression was investigated in this study, and it was demonstrated that it constitutes a major regulatory step in the specific activation of the expression of flavonoid structural genes. First, the GUS activity generated in planta from a TT8::uidA construct revealed cell-specific activation of the TT8 promoter consistent with the known involvement of the TT8 bHLH factor in proanthocyanidin, anthocyanin and mucilage biosynthesis. Moreover, the activity of this reporter construct was strongly affected in ttg1, TT2 overexpressers (OE), and PAP1-OE, suggesting interplay between TT2, PAP1, TTG1 and the activation of the TT8 promoter in planta. To further investigate the mechanisms involved, we used 35S::TT2-GR and 35S::TTG1-GR transgenic plants (expressing fusion proteins with the glucocorticoid receptor), as well as one-hybrid experiments, to determine the direct effect of these factors on TT8 expression. Interestingly, in vivo binding of TT2 and PAP1 to the TT8 promoter was dependent on the simultaneous expression of TT8 or the homologous bHLH factors GL3 and EGL3. Consistent with these results, the activity of the TT8::uidA reporter was strongly affected in the seed endothelium of a tt8 mutant. Similarly, a strong decrease in the level of TT8 mRNA was detected in the siliques of a gl3 x egl3 mutant and in plants that express a dominant negative form of the PAP1 protein, suggesting that TT8 expression is controlled by different combinations of MYB and bHLH factors in planta. The importance of this positive feedback mechanism in the strong and specific induction of proanthocyanidin biosynthesis in the seed coat of Arabidopsis thaliana is discussed.
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            Isolation of a regulatory gene of anthocyanin biosynthesis in tuberous roots of purple-fleshed sweet potato.

            Many transcriptional factors harboring the R2R3-MYB domain, basic helix-loop-helix domain, or WD40 repeats have been identified in various plant species as regulators of flavonoid biosynthesis in flowers, seeds, and fruits. However, the regulatory elements of flavonoid biosynthesis in underground organs have not yet been elucidated. We isolated the novel MYB genes IbMYB1 and IbMYB2s from purple-fleshed sweet potato (Ipomoea batatas L. Lam. cv Ayamurasaki). IbMYB1 was predominantly expressed in the purple flesh of tuberous roots but was not detected (or only scarcely) in other anthocyanin-containing tissues such as nontuberous roots, stems, leaves, or flowers. IbMYB1 was also expressed in the tuberous roots of other purple-fleshed cultivars but not in those of orange-, yellow-, or white-fleshed cultivars. Although the orange- or yellow-fleshed cultivars contained anthocyanins in the skins of their tuberous roots, we could not detect IbMYB1 transcripts in these tissues. These results suggest that IbMYB1 controls anthocyanin biosynthesis specifically in the flesh of tuberous roots. The results of transient and stable transformation experiments indicated that expression of IbMYB1 alone was sufficient for induction of all structural anthocyanin genes and anthocyanin accumulation in the flesh of tuberous roots, as well as in heterologous tissues or heterologous plant species.
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              Convergent evolution in the BAHD family of acyl transferases: identification and characterization of anthocyanin acyl transferases from Arabidopsis thaliana.

              Members of the BAHD family of plant acyl transferases are very versatile catalytically, and are thought to be able to evolve new substrate specificities rapidly. Acylation of anthocyanins occurs in many plant species and affects anthocyanin stability and light absorption in solution. The versatility of BAHD acyl transferases makes it difficult to identify genes encoding enzymes with defined substrate specificities on the basis of structural homology to genes of known catalytic function alone. Consequently, we have used a modification to standard functional genomics strategies, incorporating co-expression profiling with anthocyanin accumulation, to identify genes encoding three anthocyanin acyl transferases from Arabidopsis thaliana. We show that the activities of these enzymes influence the stability of anthocyanins at neutral pH, and some acylations also affect the anthocyanin absorption maxima. These properties make the BAHD acyl transferases suitable tools for engineering anthocyanins for an improved range of biotechnological applications.
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                Author and article information

                Journal
                Molecules
                Molecules
                molecules
                Molecules
                MDPI
                1420-3049
                17 October 2019
                October 2019
                : 24
                : 20
                : 3743
                Affiliations
                Institute of Crop Sciences, Fujian Academy of Agricultural Sciences, Scientific Observing and Experimental Station of Tuber and Root Crops in South China, Ministry of Agriculture. Fuzhou, Fujian 350013, China; uslgl@ 123456126.com (G.L.); linzhaomiao@ 123456foxmail.com (Z.L.); teeteeking@ 123456163.com (H.Z.); lhl8620@ 123456163.com (Z.L.); qingqing0722@ 123456126.com (Y.X.); xgc_faas@ 123456163.com (G.X.); fjpotato@ 123456126.com (H.L.); jrc1976@ 123456163.com (R.J.); lwb9630@ 123456163.com (W.L.); qyxlm@ 123456sohu.com (Y.Q.); tanghao9403@ 123456163.com (H.T.)
                Author notes
                [* ]Correspondence: qiusixin@ 123456faas.cn ; Tel.: +86-0591-87572407
                Author information
                https://orcid.org/0000-0003-4221-2899
                Article
                molecules-24-03743
                10.3390/molecules24203743
                6832942
                31627373
                9458f8ee-21ea-452f-9172-2264a14c3156
                © 2019 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                : 20 September 2019
                : 15 October 2019
                Categories
                Article

                sweet potato,anthocyanin compositions,biosynthesis structural genes,transcription factor

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