Genetic variation in a tandemly duplicated TPS gene cluster contributes to the diversity of aroma in lychee fruit

Fruits undergo a similar ripening process, yet they exhibit a range of differences in color, taste, and shape, both across different species and within the same species. How does this diversity arise? We uncovered a conserved fruit ripening process in lychee fruit in which a NAC transcription factor, LcNAC1, acts as a master regulator. LcNAC1 regulates the expression of two terpene synthase genes, LcTPSa1 and LcTPSa2, which belong to a gene cluster consisting of four TPS genes. LcTPSa1-LcTPSa3 are responsible for catalyzing the production of farnesol, which in turn dictates the aromatic diversity in fruit of different lychee varieties. Through comparative, transcriptomic, and genomic analyses across various lychee varieties, we found these four TPS genes exhibit distinct expression levels due to natural genetic variation. These include copy number variations, presence/absence variations, insertions and deletions, and single nucleotide polymorphisms, many of which affect the binding affinity of LcNAC1. A single nucleotide mutation in LcTPSa1 caused a premature translational termination, resulting in a truncated version of the TPS protein, which surprisingly remains functional. All these genomic changes in the LcNAC1-regulated TPS genes are likely to contribute to the great aromatic diversity observed in lychee fruit. This diversification of fruit aroma in lychee varieties offers a compelling example of how species- or variety-specific traits evolve - the phenotypic diversity is primarily derived from natural genetic variation accumulated in downstream structural genes within an evolutionarily conserved regulatory circuit.