Functional Annotation, Genome Organization and Phylogeny of the Grapevine (<it>Vitis vinifera</it>) Terpene Synthase Gene Family Based on Genome Assembly, FLcDNA Cloning, and Enzyme Assays

<p>Abstract</p> <p>Background</p> <p>Terpenoids are among the most important constituents of grape flavour and wine bouquet, and serve as useful metabolite markers in viticulture and enology. Based on the initial 8-fold sequencing of a nearly homozygous Pinot noir inbred line, 89 putative terpenoid synthase genes (<it>VvTPS</it>) were predicted by <it>in silico </it>analysis of the grapevine (<it>Vitis vinifera</it>) genome assembly <abbrgrp><abbr bid="B1">1</abbr></abbrgrp>. The finding of this very large <it>VvTPS </it>family, combined with the importance of terpenoid metabolism for the organoleptic properties of grapevine berries and finished wines, prompted a detailed examination of this gene family at the genomic level as well as an investigation into <it>VvTPS </it>biochemical functions.</p> <p>Results</p> <p>We present findings from the analysis of the up-dated 12-fold sequencing and assembly of the grapevine genome that place the number of predicted <it>VvTPS </it>genes at 69 putatively functional <it>VvTPS</it>, 20 partial <it>VvTPS</it>, and 63 <it>VvTPS </it>probable pseudogenes. Gene discovery and annotation included information about gene architecture and chromosomal location. A dense cluster of 45 <it>VvTPS </it>is localized on chromosome 18. Extensive FLcDNA cloning, gene synthesis, and protein expression enabled functional characterization of 39 <it>VvTPS</it>; this is the largest number of functionally characterized <it>TPS </it>for any species reported to date. Of these enzymes, 23 have unique functions and/or phylogenetic locations within the plant <it>TPS </it>gene family. Phylogenetic analyses of the <it>TPS </it>gene family showed that while most <it>VvTPS </it>form species-specific gene clusters, there are several examples of gene orthology with <it>TPS </it>of other plant species, representing perhaps more ancient <it>VvTPS</it>, which have maintained functions independent of speciation.</p> <p>Conclusions</p> <p>The highly expanded <it>VvTPS </it>gene family underpins the prominence of terpenoid metabolism in grapevine. We provide a detailed experimental functional annotation of 39 members of this important gene family in grapevine and comprehensive information about gene structure and phylogeny for the entire currently known <it>VvTPS </it>gene family.</p>