Abstract
Gene families involved in specialised metabolism play a key role in a myriad of ecophysiological and biochemical functions. The Vitis vinifera sesquiterpene synthases represent the largest subfamily of grapevine terpene synthase (VviTPS) genes and are important volatile metabolites for wine flavour and aroma, as well as ecophysiological interactions. The functional characterisation of VviTPS genes is complicated by a reliance on a single reference genome that greatly underrepresents this large gene family, exacerbated by extensive duplications and paralogy. The recent release of multiple phased diploid grapevine genomes, as well as extensive whole-genome resequencing efforts, provide a wealth of new sequence information that can be utilised to overcome the limitations of the reference genome. A large cluster of sesquiterpene synthases, localised to chromosome 18, was explored by means of comparative sequence analyses using the publicly available grapevine reference genome, three PacBio phased diploid genomes and whole-genome resequencing data from multiple genotypes. Two genes, VviTPS04 and -10, were identified as putative paralogues and/or allelic variants. Subsequent gene isolation from multiple grapevine genotypes and characterisation by means of a heterologous in planta expression and volatile analysis resulted in the identification of genotype-specific structural variations and polymorphisms that impact the gene function. These results present novel insight into how grapevine domestication likely shaped the VviTPS landscape to result in genotype-specific functions.
Highlights
The Vitis vinifera reference genome has been an invaluable resource for the identification of genes involved in metabolic processes of agronomic interest [1]
The PN40024 genome showed that the forward strand of the VviTPS04 locus had a start codon 27 bp upstream of the start codon for the VviTPS10 locus [1,7]
The 28 phased diploid and two PN40024 genomic DNA (gDNA) models were subsequently used to map short reads from 11 different genotypes that were used in the whole-genome resequencing (WGRS) effort by reference [37]
Summary
The Vitis vinifera reference genome has been an invaluable resource for the identification of genes involved in metabolic processes of agronomic interest [1]. The near-homozygous reference genome PN40024 contains 152 V. vinifera terpene synthase (VviTPS)-like loci, of which 69 are predicted to be functional [1,7]. Thirty of these gene models were subsequently linked to a functional enzyme but were isolated from various genotypes [7]. The VviTPS family annotated on the PN40024 reference genome largely encodes for genes involved in mono- (C10) and sesquiterpene (C15) biosynthesis. These metabolites are biosynthesised via two compartmentalised pathways.
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