Functional analysis of MYB alleles from Solanum chilense and Solanum lycopersicum in controlling anthocyanin levels in heterologous tobacco plants

Flavonoids are natural pigments occurring in plants and are present in fruits, leaves, stems, roots, and flowers. Tobacco plants transformed with an MYB regulatory gene from either Solanum chilense (Sc) or S. lycopersicum (Sl) demonstrate that ScANT1 induces a higher level of anthocyanin accumulation in comparison to SlANT1 and that this gene is sufficient to promote increased anthocyanin levels. We compared the aptitude of ScANT1 protein to induce anthocyanin accumulation to that of SlANT1 protein in tobacco plants. We also tested the effect of amino acid substitutions in ScANT1 and SlANT1. We examined these synthetic alleles' effect following the over-expression of additional anthocyanin synthesis regulators, such as the tomato bHLH (SlJAF13) protein. Our results show that the amino acid changes that differentiate ScANT1 from SlANT1 are the main contributors to the advantage that ScANT1 has over SlANT1 in anthocyanin accumulation per transcript unit. We further demonstrated that altering the amino acid composition of SlANT1 could increase anthocyanin accumulation, while reciprocally modifying ScANT1 lowers the anthocyanin level. These results confirm the increased anthocyanin level in tobacco is attributed to the amino acid differences between ScANT1 and SlANT1. We also show that the co-expression of SlJAF13 with SlANT1 in tobacco plants represses the anthocyanin production.