A Cluster of MYB Transcription Factors Regulates Anthocyanin Biosynthesis in Carrot (Daucus carota L.) Root and Petiole

Abstract

Purple carrots can accumulate large quantities of anthocyanins in their roots and –in some genetic backgrounds- petioles, and therefore they represent an excellent dietary source of antioxidant phytonutrients. In a previous study, using linkage analysis in a carrot F2 mapping population segregating for root and petiole anthocyanin pigmentation, we identified a region in chromosome 3 with co-localized QTL for all anthocyanin pigments of the carrot root, whereas petiole pigmentation segregated as a single dominant gene and mapped to one of these “root pigmentation” regions conditioning anthocyanin biosynthesis. In the present study, we performed fine mapping combined with gene expression analyses (RNA-Seq and RT-qPCR) to identify candidate genes controlling anthocyanin pigmentation in the carrot root and petiole. Fine mapping was performed in four carrot populations with different genetic backgrounds and patterns of pigmentation. The regions controlling root and petiole pigmentation in chromosome 3 were delimited to 541 and 535 kb, respectively. Genome wide prediction of transcription factor families known to regulate the anthocyanin biosynthetic pathway coupled with orthologous and phylogenetic analyses enabled the identification of a cluster of six MYB transcription factors, denominated DcMYB6 to DcMYB11, associated with the regulation of anthocyanin biosynthesis. No anthocyanin biosynthetic genes were present in this region. Comparative transcriptome analysis indicated that upregulation of DcMYB7 was always associated with anthocyanin pigmentation in both root and petiole tissues, whereas DcMYB11 was only upregulated with pigmentation in petioles. In the petiole, the level of expression of DcMYB11 was higher than DcMYB7. DcMYB6, a gene previously suggested as a key regulator of carrot anthocyanin biosynthesis, was not consistently associated with pigmentation in either tissue. These results strongly suggest that DcMYB7 is a candidate gene for root anthocyanin pigmentation in all the genetic backgrounds included in this study. DcMYB11 is a candidate gene for petiole pigmentation in all the purple carrot sources in this study. Since DcMYB7 is co-expressed with DcMYB11 in purple petioles, the latter gene may act also as a co-regulator of anthocyanin pigmentation in the petioles. This study provides linkage-mapping and functional evidence for the candidacy of these genes for the regulation of carrot anthocyanin biosynthesis.

DcMYB113, a root‐specific R2R3‐MYB, conditions anthocyanin biosynthesis and modification in carrot

“…Three QTLs associated with variations in Cy3XSGG in carrots have been proposed previously (Cavagnaro et al , ). Q1 is located in the same region as DcMYB7 , a gene related to anthocyanin biosynthesis in carrots with a purple root and petiole (Iorizzo et al , ). A UCGXT and a SAT gene , namely DcUCGXT1 and DcSAT1 , with the best BLAST match to UGT79B1 and SAT (At2g23000) from Arabidopsis (Montefiori et al , ), was found within the Q3 and Q2 locus, respectively (Xu et al , ).…”

“…In our previous report, we identified DcMYB6 , an R2R3‐MYB gene involved in carrot root anthocyanin biosynthesis within the P 3 region (Xu et al , ). Iorizzo et al identified an R2R3‐MYB gene, DcMYB7 , within the P 3 region as a candidate for the gene controlling anthocyanin biosynthesis in carrot with a purple root and petiole (Iorizzo et al , ). Recently, we confirmed that DcMYB7 is P 3 (Xu et al , ).…”

Changing Carrot Color: Insertions in DcMYB7 Alter the Regulation of Anthocyanin Biosynthesis and Modification

“…In addition, purple carrot root pigmentation extensively varies across different carrot genotypes, ranging from the purple peridermal carrot type (purple periderm but nonpurple phloem and xylem) to the solid purple carrot type (purple periderm, phloem, and xylem). The genetic control of anthocyanin pigmentation in purple carrots has been investigated (Simon, 1996;Yildiz et al, 2013;Cavagnaro et al, 2014;Xu et al, 2014Xu et al, , 2016Xu et al, , 2017Kodama et al, 2018;Iorizzo et al, 2019). Two genes that condition the anthocyanin pigmentation of carrot roots from different genetic backgrounds, P 1 and P 3 , have been identified and genetically mapped within 28.2-and 12-centimorgan regions, respectively, on chromosome 3, supporting the theory of two independent mutation and human selection events during the domestication of purple carrots (Cavagnaro et al, 2014).…”

“…DcMYB6, corresponding to DCAR_000385 in the carrot genome, encodes an R2R3-MYB that can induce anthocyanin biosynthesis in Arabidopsis (Arabidopsis thaliana; Xu et al, 2017). Very recently, DcMYB6 was anchored to the P 3 region (Iorizzo et al, 2019). Using phylogenetic analyses, these authors also identified five additional MYB transcription factors of the anthocyanin biosynthesis-related subgroup of MYBs, DcMYB7 (DCAR_010745), which was designated DcMYB113-like in the carrot genome (Iorizzo et al, 2016), DcMYB8 (DCAR_010746), DcMYB9 (DCAR_010747), DcMYB10 (DCAR_010749), and DcMYB11 (DCAR_010751), within the P 3 genomic region (Iorizzo et al, 2019).…”

“…Very recently, DcMYB6 was anchored to the P 3 region (Iorizzo et al, 2019). Using phylogenetic analyses, these authors also identified five additional MYB transcription factors of the anthocyanin biosynthesis-related subgroup of MYBs, DcMYB7 (DCAR_010745), which was designated DcMYB113-like in the carrot genome (Iorizzo et al, 2016), DcMYB8 (DCAR_010746), DcMYB9 (DCAR_010747), DcMYB10 (DCAR_010749), and DcMYB11 (DCAR_010751), within the P 3 genomic region (Iorizzo et al, 2019). Among these R2R3-MYBs, the expression of DcMYB7 was consistently associated with purple root pigmentation in all the purple carrots tested, while the expression of DcMYB6 was only associated with anthocyanin pigmentation in the solid purple carrots.…”