Comprehending and improving cannabis specialized metabolism in the systems biology era.

Cannabis sativa is a source of food, fiber and specialized metabolites such as cannabinoids, with psychoactive and pharmacological effects. Due to its expanding and increasingly-accepted use in medicine, cannabis cultivation is acquiring more importance and less social stigma. Humans initiated different domestication episodes whose later spread gave rise to a plethora of landrace cultivars. At present, breeders cross germplasms from different gene pools depending on their specific use. The fiber (hemp) and drug (marijuana) types of C. sativa differ in their cannabinoid chemical composition phenotype (chemotype) and also in the accumulation of terpenoid compounds that constitute a strain's particular flavor and scent. Cannabinoids are isoprenylated polyketides among which cannabidiolic acid (CBDA) and (-)-trans-Δ9-tetrahydrocannabinol acid (THCA) have been well-documented for their many effects on humans. Here, we review the most studied specialized metabolic pathways in C. sativa, showing how terpenes and cannabinoids share both part of the isoprenoid pathway and the same biosynthetic compartmentalization (i.e. glandular trichomes of leaves and flowers). We enlist the several studies that have deciphered these pathways in this species including physical and genetic maps, QTL analyses and localization and enzymatic studies of cannabinoid and terpene synthases. In addition, new comparative modeling of cannabinoid synthases and phylogenetic trees are presented. We describe the genome sequencing initiatives of several accessions with the concomitant generation of next-generation genome maps and transcriptomic data. Very recently, proteomic characterizations and systems biology approaches such as those applying network theory or the integration of multi-omics data have increased the knowledge on gene function, enzyme diversity and metabolite content in C. sativa. In this revision we drift through the history, present and future of cannabis research and on how second- and third-generation sequencing technologies are bringing light to the field of cannabis specialized metabolism. We also discuss different biotechnological approaches for producing cannabinoids in engineered microorganisms.

Isolation of WDR and bHLH genes related to flavonoid synthesis in grapevine (Vitis vinifera L.).

Anthocyanins and tannins are two of the most abundant flavonoids found in grapevine, and their synthesis is derived from the phenylpropanoid pathway. As described for model species such as Arabidopsis thaliana, maize and petunia, the end-point branches of this pathway are tightly regulated by the combinatorial interaction of three families of regulatory factors; MYB, bHLH (also known as MYC) and WDR proteins. Among these, only MYB genes have been previously identified in grapes. Here, we report the isolation of the first members from the WDR and bHLH families found in Vitis vinifera, named WDR1, WDR2 and MYCA1. WDR1 contributed positively to the accumulation of anthocyanins when it was overexpressed in A. thaliana, although it was not possible to determine the function of WDR2 by ectopic expression. The sub-cellular localizations of WDR1 and MYCA1 were observed by means of GFP-fusion proteins, indicating both cytoplasm and nuclear localization, in contrast to the localization of a MYB factor exclusively in the nucleus. The expression patterns of these genes were quantified in coloured reproductive organs throughout development, and correlated with anthocyanin accumulation and the expression profiles of the flavonoid-related MYBA1-2, UFGT, and ANR genes. In vitro grapevine plantlets grown under high salt concentrations showed a cultivar-dependent response for anthocyanin accumulation, which correlated with the expression of MYBA1-2, MYCA1 and WDR1 genes. These results suggest that MYCA1 may regulate ANR and UFGT and that this last control is easier to distinguish whenever MYBA genes are absent or in low abundance. Future studies should address the specific interactions of these proteins and their quantitative contribution to flavonoid synthesis in grape berries.