scholarly journals Transposon-induced methylation of the RsMYB1 promoter disturbs anthocyanin accumulation in red-fleshed radish

2020 ◽  
Vol 71 (9) ◽  
pp. 2537-2550 ◽  
Author(s):  
Qingbiao Wang ◽  
Yanping Wang ◽  
Honghe Sun ◽  
Liang Sun ◽  
Li Zhang
Keyword(s):  
Dna Methylation ◽  
Raphanus Sativus ◽  
Promoter Region ◽  
Sequence Variation ◽  
Anthocyanin Biosynthesis ◽  
Underlying Mechanism ◽  
Anthocyanin Accumulation ◽  
Virus Induced Gene Silencing ◽  
Genome Resequencing ◽  
Raphanus Sativus L

Abstract Red-fleshed radish (Raphanus sativus L.) is a unique cultivar whose taproot is rich in anthocyanins beneficial to human health. However, the frequent occurrence of white-fleshed mutants affects the purity of commercially produced radish and the underlying mechanism has puzzled breeders for many years. In this study, we combined quantitative trait location by genome resequencing and transcriptome analyses to identify a candidate gene (RsMYB1) responsible for anthocyanin accumulation in red-fleshed radish. However, no sequence variation was found in the coding and regulatory regions of the RsMYB1 genes of red-fleshed (MTH01) and white-fleshed (JC01) lines, and a 7372 bp CACTA transposon in the RsMYB1 promoter region occurred in both lines. A subsequent analysis suggested that the white-fleshed mutant was the result of altered DNA methylation in the RsMYB1 promoter. This heritable epigenetic change was due to the hypermethylated CACTA transposon, which induced the spreading of DNA methylation to the promoter region of RsMYB1. Thus, RsMYB1 expression was considerably down-regulated, which inhibited anthocyanin biosynthesis in the white-fleshed mutant. An examination of transgenic radish calli and the results of a virus-induced gene silencing experiment confirmed that RsMYB1 is responsible for anthocyanin accumulation. Moreover, the mutant phenotype was partially eliminated by treatment with a demethylating agent. This study explains the molecular mechanism regulating the appearance of white-fleshed mutants of red-fleshed radish.

MYB_SH[AL]QKY[RF] transcription factors MdLUX and MdPCL-like promote anthocyanin accumulation through DNA hypomethylation and MdF3H activation in apple

2020 ◽  
Author(s):  
Wen-Fang Li ◽  
Gai-Xing Ning ◽  
Cun-Wu Zuo ◽  
Ming-Yu Chu ◽  
Shi-Jin Yang ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Transcription Factors ◽  
Promoter Region ◽  
Anthocyanin Biosynthesis ◽  
Apple Fruit ◽  
Anthocyanin Synthesis ◽  
Mrna Levels ◽  
Anthocyanin Accumulation ◽  
Dna Hypomethylation ◽  
Fruit Skin

Abstract Heritable DNA methylation is a highly conserved epigenetic mark that is important for many biological processes. In a previous transcriptomic study on the fruit skin pigmentation of apple (Malus domestica Borkh.) cv. ‘Red Delicious’ (G0) and its four continuous-generation bud sport mutants including ‘Starking Red’ (G1), ‘Starkrimson’ (G2), ‘Campbell Redchief’ (G3) and ‘Vallee spur’ (G4), we identified MYB transcription factors (TFs) MdLUX and MdPCL-like involved in regulating anthocyanin synthesis. However, how these TFs ultimately determine the fruit skin colour traits remain elusive. Here, bioinformatics analysis revealed that MdLUX and MdPCL-like contained a well-conserved motif SH[AL]QKY[RF] in their C-terminal region and were located in the nucleus of onion epidermal cells. Overexpression of MdLUX and MdPCL-like in ‘Golden Delicious’ fruits, ‘Gala’ calli and Arabidopsis thaliana promoted the accumulation of anthocyanin, whereas MdLUX and MdPCL-like suppression inhibited anthocyanin accumulation in ‘Red Fuji’ apple fruit skin. Yeast one-hybrid assays revealed that MdLUX and MdPCL-like may bind to the promoter region of the anthocyanin biosynthesis gene MdF3H. Dual-luciferase assays indicated that MdLUX and MdPCL-like activated MdF3H. The whole-genome DNA methylation study revealed that the methylation levels of the mCG context at the upstream (i.e., promoter region) of MdLUX and MdPCL-like were inversely correlated with their mRNA levels and anthocyanin accumulation. Hence, the data suggest that MYB_SH[AL]QKY[RF] TFs MdLUX and MdPCL-like promote anthocyanin biosynthesis in apple fruit skins through the DNA hypomethylation of their promoter regions and the activation of the structural flavonoid gene MdF3H.

scholarly journals The Purple Leaf (pl6) Mutation Regulates Leaf Color by Altering the Anthocyanin and Chlorophyll Contents in Rice

Plants ◽  
2020 ◽  
Vol 9 (11) ◽  
pp. 1477
Author(s):  
Asadullah Khan ◽  
Sanaullah Jalil ◽  
Huan Cao ◽  
Yohannes Tsago ◽  
Mustapha Sunusi ◽  
...  
Keyword(s):  
Gamma Ray ◽  
Anthocyanin Biosynthesis ◽  
Light Attenuation ◽  
Transcript Level ◽  
Underlying Mechanism ◽  
Anthocyanin Accumulation ◽  
Morphological Marker ◽  
Chlorophyll Contents ◽  
Rice Mutant ◽  
Purple Coloration

The anthocyanin biosynthesis attracts strong interest due to the potential antioxidant value and as an important morphological marker. However, the underlying mechanism of anthocyanin accumulation in plant tissues is not clearly understood. Here, a rice mutant with a purple color in the leaf blade, named pl6, was developed from wild type (WT), Zhenong 41, with gamma ray treatment. By map-based cloning, the OsPL6 gene was located on the short arm of chromosome 6. The multiple mutations, such as single nucleotide polymorphism (SNP) at −702, −598, −450, an insertion at −119 in the promoter, three SNPs and one 6-bp deletion in the 5′-UTR region, were identified, which could upregulate the expression of OsPL6 to accumulate anthocyanin. Subsequently, the transcript level of structural genes in the anthocyanin biosynthesis pathway, including OsCHS, OsPAL, OsF3H and OsF3′H, was elevated significantly. Histological analysis revealed that the light attenuation feature of anthocyanin has degraded the grana and stroma thylakoids, which resulted in poor photosynthetic efficiency of purple leaves. Despite this, the photoabatement and antioxidative activity of anthocyanin have better equipped the pl6 mutant to minimize the oxidative damage. Moreover, the contents of abscisic acid (ABA) and cytokanin (CK) were elevated along with anthocyanin accumulation in the pl6 mutant. In conclusion, our results demonstrate that activation of OsPL6 could be responsible for the purple coloration in leaves by accumulating excessive anthocyanin and further reveal that anthocyanin acts as a strong antioxidant to scavenge reactive oxygen species (ROS) and thus play an important role in tissue maintenance.

scholarly journals SlBBX20 interacts with the COP9 signalosome subunit SlCSN5-2 to regulate anthocyanin biosynthesis by activating SlDFR expression in tomato

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Dan Luo ◽  
Cheng Xiong ◽  
Aihua Lin ◽  
Chunli Zhang ◽  
Wenhui Sun ◽  
...  
Keyword(s):  
Growth Regulation ◽  
Anthocyanin Biosynthesis ◽  
Plant Stress ◽  
Underlying Mechanism ◽  
Bimolecular Fluorescence Complementation ◽  
Cop9 Signalosome ◽  
Anthocyanin Accumulation ◽  
Related Transcription Factor ◽  
Hybrid Screening

AbstractAnthocyanins play vital roles in plant stress tolerance and growth regulation. Previously, we reported that the photomorphogenesis-related transcription factor SlBBX20 regulates anthocyanin accumulation in tomato. However, the underlying mechanism remains unclear. Here, we showed that SlBBX20 promotes anthocyanin biosynthesis by binding the promoter of the anthocyanin biosynthesis gene SlDFR, suggesting that SlBBX20 directly activates anthocyanin biosynthesis genes. Furthermore, we found by yeast two-hybrid screening that SlBBX20 interacts with the COP9 signalosome subunit SlCSN5-2, and the interaction was confirmed by bimolecular fluorescence complementation and coimmunoprecipitation assays. SlCSN5 gene silencing led to anthocyanin hyperaccumulation in the transgenic tomato calli and shoots, and SlCSN5-2 overexpression decreased anthocyanin accumulation, suggesting thSlCSN5-2 enhanced the ubiquitination of SlBBX20 and promoted the degradation of SlBBX20 in vivo. Consistently, silencing the SlCSN5-2 homolog in tobacco significantly increased the accumulation of the SlBBX20 protein. Since SlBBX20 is a vital regulator of photomorphogenesis, the SlBBX20-SlCSN5-2 module may represent a novel regulatory pathway in light-induced anthocyanin biosynthesis.

scholarly journals Loss of the R2R3 MYB Transcription Factor RsMYB1 Shapes Anthocyanin Biosynthesis and Accumulation in Raphanus sativus

2021 ◽  
Vol 22 (20) ◽  
pp. 10927
Author(s):  
Da-Hye Kim ◽  
Jundae Lee ◽  
JuHee Rhee ◽  
Jong-Yeol Lee ◽  
Sun-Hyung Lim
Keyword(s):  
Transcription Factor ◽  
Raphanus Sativus ◽  
Anthocyanin Biosynthesis ◽  
Antioxidant Properties ◽  
Myb Transcription Factor ◽  
Anthocyanin Accumulation ◽  
Truncated Protein ◽  
Frame Shift ◽  
Frame Shift Mutation ◽  
R2r3 Myb

The red or purple color of radish (Raphanus sativus L.) taproots is due to anthocyanins, which have nutritional and aesthetic value, as well as antioxidant properties. Moreover, the varied patterns and levels of anthocyanin accumulation in radish roots make them an interesting system for studying the transcriptional regulation of anthocyanin biosynthesis. The R2R3 MYB transcription factor RsMYB1 is a key positive regulator of anthocyanin biosynthesis in radish. Here, we isolated an allele of RsMYB1, named RsMYB1Short, in radish cultivars with white taproots. The RsMYB1Short allele carried a 4 bp insertion in the first exon causing a frame-shift mutation of RsMYB1, generating a truncated protein with only a partial R2 domain at the N-terminus. Unlike RsMYB1Full, RsMYB1Short was localized to the nucleus and the cytoplasm and failed to interact with their cognate partner RsTT8. Transient expression of genomic or cDNA sequences for RsMYB1Short in radish cotyledons failed to induce anthocyanin accumulation, but that for RsMYB1Full activated it. Additionally, RsMYB1Short showed the lost ability to induce pigment accumulation and to enhance the transcript level of anthocyanin biosynthetic genes, while RsMYB1Full promoted both processes when co-expressed with RsTT8 in tobacco leaves. As the result of the transient assay, co-expressing RsTT8 and RsMYB1Full, but not RsMYB1Short, also enhanced the promoter activity of RsCHS and RsDFR. We designed a molecular marker for RsMYB1 genotyping, and revealed that the RsMYB1Short allele is common in white radish cultivars, underscoring the importance of variation at the RsMYB1 locus in anthocyanin biosynthesis in the radish taproot. Together, these results indicate that the nonsense mutation of RsMYB1 generated the truncated protein, RsMYB1Short, that had the loss of ability to regulate anthocyanin biosynthesis. Our findings highlight that the frame shift mutation of RsMYB1 plays a key role in anthocyanin biosynthesis in the radish taproot.

scholarly journals The RNA Directed DNA Methylation (RdDM) Pathway Regulates Anthocyanin Biosynthesis in Crabapple (Malus cv. spp.) Leaves by Methylating the McCOP1 Promoter

Plants ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 2466
Author(s):  
Yifan Xing ◽  
Ziyi Xie ◽  
Weilei Sun ◽  
Yuying Sun ◽  
Zhenyun Han ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Methylation Level ◽  
Bisulfite Sequencing ◽  
Anthocyanin Biosynthesis ◽  
Apple Fruit ◽  
Bimolecular Fluorescence Complementation ◽  
Rna Expression ◽  
Anthocyanin Accumulation ◽  
Transcript Levels ◽  
Direct Target

The synthesis of anthocyanin pigments in plants is known to be regulated by multiple mechanisms, including epigenetic regulation; however, the contribution of the RNA-directed DNA methylation (RdDM) pathway is not well understood. Here, we used bisulfite sequencing and Real Time (RT)-quantitative (q) PCR to analyze the methylation level of the promoter of constitutively photomorphogenic 1 (McCOP1) from Malus cv. spp, a gene involved in regulating anthocyanin biosynthesis. The CHH methylation level of the McCOP1 promoter was negatively correlated with McCOP1 RNA expression, and inhibiting DNA methylation caused decreased methylation of the McCOP1 promoter and asymmetric cytosine CHH methylation. We observed that the McCOP1 promoter was a direct target of the RdDM pathway argonaute RISC component 4 (McAGO4) protein, which bound to a McCOP1 promoter GGTTCGG site. Bimolecular fluorescence complementation (BIFC) analysis showed that RNA-directed DNA methylation (McRDM1) interacted with McAGO4 and another RdDM protein, domains rearranged methyltransferase 2 (McDRM2), to regulate the CHH methylation of the McCOP1 promoter. Detection of CHH methylation and COP1 gene expression in the Arabidopsis thalianaatago4, atdrm2 and atrdm1 mutants showed that RDM1 is the effector of the RdDM pathway. This was confirmed by silencing McRDM1 in crabapple leaves or apple fruit, which resulted in a decrease in McCOP1 CHH methylation and an increase in McCOP1 transcript levels, as well as in anthocyanin accumulation. In conclusion, these results show that the RdDM pathway is involved in regulating anthocyanin accumulation through CHH methylation of the McCOP1 promoter.

FvbHLH9 Functions as a Positive Regulator of Anthocyanin Biosynthesis by Forming a HY5–bHLH9 Transcription Complex in Strawberry Fruits

2020 ◽  
Vol 61 (4) ◽  
pp. 826-837 ◽  
Author(s):  
Yang Li ◽  
Pengbo Xu ◽  
Guanqun Chen ◽  
Jun Wu ◽  
Zhongchi Liu ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Promoter Region ◽  
Anthocyanin Biosynthesis ◽  
Anthocyanin Synthesis ◽  
Bhlh Transcription Factor ◽  
Anthocyanin Accumulation ◽  
Positive Regulator ◽  
Key Enzyme ◽  
Synergistic Regulation ◽  
Transient Overexpression

Abstract Anthocyanin accumulation is transcriptionally regulated by the MYB–bHLH–WD40 complex. Light is indispensable for anthocyanin accumulation, and light-inducible MYB and HY5 were considered to promote anthocyanin accumulation in many fruits. Whether and how light-inducible bHLH transcription factor and HY5 regulate anthocyanin synthesis in strawberry is unknown. In this study, we identified a bHLH transcription factor, FvbHLH9, which was induced by light as well as FvHY5, and found that, similar to FvHY5, the transient overexpression and interference FvbHLH9 in strawberry fruits can promote and decrease anthocyanin accumulation, respectively, indicating FvbHLH9 functions as a positive regulator of anthocyanin biosynthesis. Furthermore, we confirmed that both FvHY5 and FvbHLH9 specifically bind to the promoter region of some key enzyme genes, including FvDFR, and the expression of FvDFR was activated through the heterodimer formation between FvHY5 and FvbHLH9. Finally, we confirmed that FvbHLH9-promoted anthocyanin accumulation is dependent on HY5–bHLH heterodimerisation in Arabidopsis. Our findings provide insights into a mechanism involving the synergistic regulation of light-dependent coloration and anthocyanin biosynthesis via a HY5–bHLH heterodimer formed by the interaction of FvHY5 and FvbHLH9 in strawberry fruits.

scholarly journals Transcriptomic dynamics changes in development of carmine radish (Raphanus sativus L.) fleshy roots using RNA-seq method

2019 ◽  
Author(s):  
Jian Gao ◽  
Mao Luo ◽  
Yi Liu ◽  
Fabo Chen ◽  
Hua Peng ◽  
...  
Keyword(s):  
Candidate Genes ◽  
Raphanus Sativus ◽  
Anthocyanin Biosynthesis ◽  
Profile Analysis ◽  
Development Stage ◽  
Seedling Stage ◽  
Anthocyanin Synthesis ◽  
Rna Seq ◽  
Vegetable Crop ◽  
Raphanus Sativus L

Abstract Radish ( Raphanus sativus L.), belonging to biennial root vegetable crop of Brassicaceae family, is an economically important vegetable crop with an edible taproot. Recently, most of differential expressed genes associating with anthocyanin biosynthesis have been identified in most of important fruit crops. However, transcriptome analysis of anthocyanin biosynthesis and expression of anthocyanin biosynthesis related genes in ‘Hongxin’ radish have not been fully investigated. Here, based on results from HPLC analysis, young fleshy roots obtained from the dynamics development stage of fleshy roots in carmine radish ‘Hongxin 1’ was used for RNA-Seq, including fleshy roots from seedling stage (SS), initial expansion (IE), full-expansion (FE), bolting stage (BS), initial flowering stage (IFS); full-bloom stage (FBS) and podding stage (PS). Subsequently, the putative candidate genes involved in the dynamics development stage of fleshy roots in carmine radish were identified. After that, DGE (differential gene expression) profile analysis was used to identify the pupative transcripts, compared with fleshy roots from seedling stage (SS). In addition, co-modulated DEGs (Common DEGs in the dynamic growing stages of fleshyroot in carmine radish) were also identified, from which most DGEs were more likely to participate in anthocyanin biosynthesis, including two transcription factors RsMYB and Rs RZFP . In addition, some related proteins e.g. RsCHS , RsDFR , RsANS , RsF’3H , RsF3GGT1 , Rs3AT1 , glutathione S-transferase F12, RsUFGT78D2-like and RsUDGT-75C1-like were significantly contributed to the regulatory mechanism during anthocyanin synthesis in the development stage of fleshy roots. Furthermore, GO terms comprised of “anthocyanin-containing compound biosynthetic process” and “anthocyanin-containing compound metabolic process” were commonly overrepresented in the other dynamics growing stages of fleshy roots after initial expansion of fleshy roots. Moreover, these results indicated that five significantly enrichment pathways of DEG were identified for the dynamics growing stages of fleshy roots in carmine radish, including Flavonoid biosynthesis, Flavone and flavonol biosynthesis, Diterpenoid biosynthesis, Anthocyanin biosynthesis, as well as Benzoxazinoid biosynthesis. These results will expand our understanding of complex molecular mechanism of the putative candidate genes involved in the dynamics development stage of fleshyroot in carmine radish.

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