Comparison of MdMYB1 sequences and expression of anthocyanin biosynthetic and regulatory genes between Malus domestica Borkh. cultivar ‘Ralls’ and its blushed sport

Abstract

A mutant tree with blushed coloring pattern in fruit skin was found and identified as a bud sport of Malus domestica Borkh. cultivar 'Ralls'. This study was aimed at elucidating the genetic basis of this pattern by a comparison of 'Ralls' (striped red) with this bud sport. These two varieties were grown either under normal sunlight (control group) or the fruits were bagged during development (bagged group) and the bags were removed 1 month before harvesting. Anthocyanin content increased during fruit ripening and was higher in the blushed sport than in 'Ralls', by 57% in the control group and by 73% in the bagged group after bag removal. Correspondingly, in both the varieties transcription of six anthocyanin biosynthetic genes was coordinately up-regulated, concomitant with an increase in levels of transcripts of the regulatory gene, MdMYB1, and in general these levels were higher in the blushed sport than in ''Ralls''. However, there were no significant differences between the two varieties in the nucleotide sequences of the genomic region or coding sequence of MdMYB1; nor in the sequences of cDNAs of the other known transcription factors MdbHLH3 and MdbHLH33. Interestingly, DNA methylation in the promoter region of MdMYB1 was inversely correlated with transcript levels of MdMYB1 and was significantly lower in the blushed sport. Thus, we conclude that DNA methylation plays a role in regulating MdMYB1 expression, which in turn regulates anthocyanin biosynthesis and thereby influences the color pattern of the apples.

Recent advances on the development and regulation of flower color in ornamental plants

“…Other examples include the bleaching of gentian flowers due to mutations in GtMYB3 ( Nakatsuka et al, 2008 ) and the positive regulation of anthocyanin biosynthesis and its effects on organ and tissue-specific anthocyanin accumulation in lily via LhMYB6 and LhMYB12 ( Yamagishi et al, 2010 ). Further studies discovered that sequence variations and methylation levels in MYB transcription factor genes also affected anthocyanin accumulation, but this observation was only reported in studies with fruit trees ( Espley et al, 2009 ; Xu et al, 2012 ) and maize ( Das and Messing, 1994 ; Cocciolone et al, 2001 ; Robbins et al, 2009 ).…”

Malus sieversii: the origin, flavonoid synthesis mechanism, and breeding of red-skinned and red-fleshed apples

“…The increased anthocyanin synthesis in the red bud of “Ralls” was due to higher enzyme activities of CHI and DFR 103 . Additional research on the red bud of “Ralls” showed that the structural genes in anthocyanin biosynthesis were upregulated because of the increased expression of MdMYB1 , and the low methylation level of the MdMYB1 promoter was associated with its increased expression 104 . El-Sharkawy et al suggested that differential methylation levels in the MR3 and MR7 promoter regions of MdMYB10 were the epigenetic factors causing the color mutation 105 .…”

Transcriptome Profiling Reveals Transcriptional Regulation by DNA Methyltransferase Inhibitor 5-Aza-2′-Deoxycytidine Enhancing Red Pigmentation in Bagged “Granny Smith” Apples (Malus domestica)

“…Anthocyanin biosynthesis is usually regulated by various TFs, especially the MBW complex [ 29 ]. Previously, MYB was found to be a crucial regulator of anthocyanin accumulation and fruit pigmentation in apples [ 30 , 31 ]. In the present study, 28 R2R3-MYB TFs were identified in skin samples of bagged “Granny Smith” apples following 5-aza-dC treatment, seven of which were significantly upregulated in transcription in the T2 group, corresponding to structural genes related to anthocyanin biosynthesis ( Figure 4 ).…”

“…The disruption of anthocyanin accumulation has been linked to changes in DNA methylation or demethylation in the promoter of the master anthocyanin regulator MYB [ 15 , 16 , 31 ]. Compared with “Honeycrisp” and “Royal Gala” [ 16 ], differences in methylation levels between control and 5-aza-dC-treated fruits were more obvious in “Granny Smith” [ 17 ], which reflects the striking contrast in pigmentation between the two fruits (i.e., an almost complete loss of anthocyanins in control fruits vs. red pigmentation in 5-aza-dC-treated fruits).…”