High AN1 variability and interaction with basic helix‐loop‐helix co‐factors related to anthocyanin biosynthesis in potato leaves

Abstract

SUMMARYAN1 is a regulatory gene that promotes anthocyanin biosynthesis in potato tubers and encodes a R2R3 MYB transcription factor. However, no clear evidence implicates AN1 in anthocyanin production in leaves, where these pigments might enhance environmental stress tolerance. In our study we found that AN1 displays intraspecific sequence variability in both coding/non-coding regions and in the promoter, and that its expression is associated with high anthocyanin content in leaves of commercial potatoes. Expression analysis provided evidence that leaf pigmentation is associated to AN1 expression and that StJAF13 acts as putative AN1 co-regulator for anthocyanin gene expression in leaves of the red leaf variety 'Magenta Love,' while a concomitant expression of StbHLH1 may contribute to anthocyanin accumulation in leaves of 'Double Fun.' Yeast two-hybrid experiments confirmed that AN1 interacts with StbHLH1 and StJAF13 and the latter interaction was verified and localized in the cell nucleus by bimolecular fluorescence complementation assays. In addition, transgenic tobacco (Nicotiana tabacum) overexpressing a combination of either AN1 with StJAF13 or AN1 with StbHLH1 showed deeper purple pigmentation with respect to AN1 alone. This further confirmed AN1/StJAF13 and AN1/StbHLH1 interactions. Our findings demonstrate that the classical loci identified for potato leaf anthocyanin accumulation correspond to AN1 and may represent an important step to expand our knowledge on the molecular mechanisms underlying anthocyanin biosynthesis in different plant tissues.

ANTHOCYANINS SYNTHESIS IN POTATO (Solanum tuberosum L.): GENETIC MARKERS FOR SMART BREEDING (review)

“…Sucrose may enhance the synthesis of anthocyanins, but is not capable of causing changes in anthocyanin deficient mutants [65,66]. High intraspecific variability of the StAN1 gene is noted [59,61], with mutations detected both in the coding and non-coding regions (i.e. in the intron 2 and the exon 3 of the gene).…”

“…In another study, the presence of the SINE retrotransposone in the promoter was shown to be associated with anthocyanin pigmentation of leaves. In genotypes with green leaves (without anthocyanin pigmentation), this element does not appear in the StAN1 promoter [59]. The presence of mobile elements modifies the expression of genes encoding MYB [67][68][69][70][71], hence further study of the modification effect of SINE on the StAN1 promoter functions is necessary.…”

“…2, A) in controlling the expression of structural genes involved in the biosynthesis of anthocyanins and other phenylpropanoids, especially in the periderm and pulp of pigmented tubers [57,58,63]. Besides, StAN1 is one of the key genes responsible for differences in the biosynthesis of anthocyanins not only in tubers, but in potato leaves [59]. The StAN1 sequence resembles the petunia gene PhAN2, so this locus was originally named similarly, but was later renamed [57].…”

“…can be pigmented independently, it has been suggested that locus D contains two (or more) tandem duplication of MYB gene, at that the phenotype modifications are associated with changes in the sequences of different MYB genes [57,59]. Indeed, by the homology with StAN1, another gene was found, first designated as StAN3, then renamed to StAN2, and also having the synonym StMYBA1 [57][58][59][60][61]. The predicted amino acid sequence for StMYBA1 with high similarity corresponds to a protein of the R2R3 MYB class (66 % identity to StAN1) [58].…”

“…In recent years, much attention has been paid to molecular mechanisms and genes controlling the biosynthesis of anthocyanins or their accumulation in potato tubers [57][58][59][60][61][62], and all researches emphasized the role of gene StAN1-ANTHOCYANIN1 (GenBank accession number JQ418343) (Fig. 2, A) in controlling the expression of structural genes involved in the biosynthesis of anthocyanins and other phenylpropanoids, especially in the periderm and pulp of pigmented tubers [57,58,63].…”