02539nas a2200277 4500008004100000022001400041245011000055210006900165260007900234300001200313490000800325520151400333653001401847653001801861653002401879653002301903653005501926653002301981100001802004700002802022700002602050700002702076700000902103700002702112856012202139 2013 eng d a0273-228900aFunctional characterization of a flavonoid glycosyltransferase gene from withania somnifera (Ashwagandha)0 aFunctional characterization of a flavonoid glycosyltransferase g a999 RIVERVIEW DRIVE SUITE 208, TOTOWA, NJ 07512 USAbHUMANA PRESS INCcJUN a729-7410 v1703 a

Glycosylation of flavonoids is mediated by family 1 uridine diphosphate (UDP)-dependent glycosyltransferases (UGTs). Until date, there are few reports on functionally characterized flavonoid glycosyltransferases from Withania somnifera. In this study, we cloned the glycosyltransferase gene from W. somnifera (UGT73A16) showing 85-92 % homology with UGTs from other plants. UGT73A16 was expressed as a His(6)-tagged fusion protein in Escherichia coli. Several compounds, including flavonoids, were screened as potential substrates for UGT73A16. HPLC analysis and hypsochromic shift indicated that UGT73A16 transfers a glucose molecule to several different flavonoids. Based on kinetic parameters, UGT73A16 shows more catalytic efficiency towards naringenin. Here, we explored UGT73A16 of W. somnifera as whole cell catalyst in E. coli. We used flavonoids (genistein, apigenin, kaempferol, naringenin, biochanin A, and daidzein) as substrates for this study. More than 95 % of the glucoside products were released into the medium, facilitating their isolation. Glycosylation of substrates occurred on the 7- and 3-hydroxyl group of the aglycone. UGT73A16 also displayed regiospecific glucosyl transfer activity towards 3-hydroxy flavone compound, which is the backbone of all flavonols and also for a chemically synthesized compound, not found naturally. The present study generates essential knowledge and molecular as well as biochemical tools that allow the verification of UGT73A16 in glycosylation.

10aFlavonoid10aglycosylation10aGlycosyltransferase10aHypsochromic shift10aPlant secondary product glycosyltransferase (PSPG)10aWithania somnifera1 aSingh, Somesh1 aVishwakarma, Rishi., K.1 aKumar, R., J. Santosh1 aSonawane, Prashant, D.1 aRuby1 aKhan, Bashir, Mohammad uhttp://library.ncl.res.in/content/functional-characterization-flavonoid-glycosyltransferase-gene-withania-somnifera-002293nas a2200253 4500008004100000022001400041245011800055210006900173260005700242300001400299490000700313520138200320653001301702653001501715653002401730653002301754100002601777700000901803700001801812700002701830700002401857700002701881856013101908 2013 eng d a0735-964000aFunctional characterization of a glucosyltransferase specific to flavonoid 7-O-glucosides from withania somnifera0 aFunctional characterization of a glucosyltransferase specific to a233 SPRING ST, NEW YORK, NY 10013 USAbSPRINGERcOCT a1100-11080 v313 a

Flavonoids are a large class of phenylpropanoid-derived secondary metabolites, which are usually glycosylated by UDP-glycosyltransferases with one or more sugar groups. Here, we report the cloning and biochemical characterization of a flavonoid glycosyltransferase gene from Withania somnifera (WsGT), which is an important medicinal plant used in Ayurvedic formulations. Using PCR primers, designed for a highly conserved region of previously reported glycosyltransferases, we were able to isolate the corresponding fragment of the WsGT gene. Rapid amplification of cDNA ends (RACE) was then employed to isolate full-length cDNA, which had an open reading frame of 1,371 bp that encode for 456 amino acids. Phylogenetic analysis indicated that WsGT was similar to that of family 1 GT-B glycosyltransferase. Biochemical analysis revealed that WsGT interacts with UDP-glucose and was capable of regiospecifically glycosylating flavonoid-7-ols, such as apigenin, naringenin, luteolin, diadzein and genistein. Expression profiling studies showed that WsGT was highly expressed in young and mature leaves of W. somnifera. Furthermore, exposure to salicylic acid enhanced the expression of WsGT in the leaves and heat shock treatment resulted in decreased expression of WsGT after an initial increase. This may suggest the role of WsGT in response to abiotic/biotic stresses.

10aDiadzein10aFlavonoids10aGlycosyltransferase10aWithania somnifera1 aKumar, R., J. Santosh1 aRuby1 aSingh, Somesh1 aSonawane, Prashant, D.1 aVishwakarma, R., K.1 aKhan, Bashir, Mohammad uhttp://library.ncl.res.in/content/functional-characterization-glucosyltransferase-specific-flavonoid-7-o-glucosides-withania-002706nas a2200277 4500008004100000022001400041245015800055210006900213260005700282300001200339490000700351520166600358653003502024653002002059653002302079653002502102100002702127700000902154700001802163700002702181700002302208700001602231700002602247700002702273856012802300 2013 eng d a0735-964000aMolecular cloning, biochemical characterization, and differential expression of an acetyl-CoA C-acetyltransferase gene (AACT) of brahmi (Bacopa monniera)0 aMolecular cloning biochemical characterization and differential  a233 SPRING ST, NEW YORK, NY 10013 USAbSPRINGERcJUN a547-5570 v313 a

Bacopa monniera (Brahmi) is an important Indian medicinal herb found in wet damp and marshy places. It produces medicinally important compounds known as bacosides along with alkaloids like brahmine and herpestine. Bacosides are triterpenoid saponins and their biosynthesis takes place via the isoprenoid pathway starting with acetyl-CoA. Acetyl-CoA C-acetyltransferase (AACT; EC 2.3.1.9), also known as acetoacetyl-CoA thiolase (Thiolase II), catalyzes the condensation of two acetyl-CoA to form 4-C compound acetoacetyl-CoA. Acetoacetyl-CoA is an important starting molecule for biosynthesis of various metabolites. Here, we report the cDNA cloning and characterization of acetyl-CoA C-acetyltransferase gene from B. monniera. The full-length gene was isolated using a RACE PCR protocol. The cDNA encoding AACT was designated as BmAACT (FJ947159) revealed an ORF of 1,218 bp and 405 amino acids, and shares 80 % similarity with other plant AACTs. Phylogenetic analysis showed that BmAACT is related closely to other dicot plants AACTs. The BmAACT gene was over-expressed in Escherichia coli as a 6X His-tag fusion protein and purified to homogeneity by Ni-NTA and gel filtration chromatography. Activity of recombinant protein was confirmed by thiolytic cleavage of acetoacetyl-CoA in the presence of 5 mM Mg2+, showing K (m) and V (max) of 20.67 mu M and 96.21 mu mol/min, respectively, with high catalytic efficiency (k (cat) = 2.30 x 10(5) min(-1)). Quantitative real-time PCR analysis showed that the expression of BmAACT is tissue-specific, and accumulation of transcripts is greater in roots and petals, followed by sepals, stem, leaf and pedicel.

10aAcetyl-CoA C-acetyltransferase10aBacopa monniera10aIsoprenoid pathway10aTriterpenoid saponin1 aVishwakarma, Rishi, K.1 aRuby1 aSingh, Somesh1 aSonawane, Prashant, D.1 aSrivastava, Sameer1 aKumari, Uma1 aKumar, R., J. Santosh1 aKhan, Bashir, Mohammad uhttp://library.ncl.res.in/content/molecular-cloning-biochemical-characterization-and-differential-expression-acetyl-coa-c-002078nas a2200229 4500008004100000022001400041245008700055210006900142260007300211300001000284490000700294520124300301653004001544653001701584653001901601653002001620653001401640100002701654700002701681700002501708856011501733 2014 eng d a0141-813000aConformational transitions of cinnamoyl CoA reductase 1 from leucaena leucocephala0 aConformational transitions of cinnamoyl CoA reductase 1 from leu aPO BOX 211, 1000 AE AMSTERDAM, NETHERLANDSbELSEVIER SCIENCE BVcMAR a30-350 v643 a

Conformational transitions of cinnamoyl CoA reductase, a key regulatory enzyme in lignin biosynthesis, from Leucaena leucocephala (L1-CCRH1) were studied using fluorescence and circular dichroism spectroscopy. The native protein possesses four trp residues exposed on the surface and 66% of helical structure, undergoes rapid structural transitions at and above 45 C and starts forming aggregates at 55 C. LI-CCRH1 was transformed into acid induced (pH 2.0) molten globule like structure, exhibiting altered secondary structure, diminished tertiary structure and exposed hydrophobic residues. The molten globule like structure was examined for the thermal and chemical stability. The altered secondary structure of Ll -CCRH1 at pH 2.0 was stable up to 90 C. Also, in presence of 0.25 M guanidine hydrochloride (GdnHCI), it got transformed into different structure which was stable in the vicinity of 2 M GdnHCI (as compared to drastic loss of native structure in 2 M GdnHC1) as seen in far UV-CD spectra. The structural transition of LI-CCRH1 at pH 2.0 followed another transition after readjusting the pH to 8.0, forming a structure with hardly any similarity to that of native protein. (C) 2013 Elsevier B.V. All rights reserved.

10aAggregation Cinnamoyl CoA reductase10aConformation10aMolten globule10aThermostability10aUnfolding1 aSonawane, Prashant, D.1 aKhan, Bashir, Mohammad1 aGaikwad, Sushama, M. uhttp://library.ncl.res.in/content/conformational-transitions-cinnamoyl-coa-reductase-1-leucaena-leucocephala-002263nas a2200301 4500008004100000022001400041245011700055210006900172260009000241300002600331490000600357520118200363100002401545700002701569700001901596700002701615700002001642700002001662700001401682700001601696700002001712700002101732700002001753700002001773700001801793700001901811856013101830 2016 eng d a2041-172300aGAME9 regulates the biosynthesis of steroidal alkaloids and upstream isoprenoids in the plant mevalonate pathway0 aGAME9 regulates the biosynthesis of steroidal alkaloids and upst aMACMILLAN BUILDING, 4 CRINAN ST, LONDON N1 9XW, ENGLANDbNATURE PUBLISHING GROUPcFEB aArticle Number: 106540 v73 a

Steroidal glycoalkaloids (SGAs) are cholesterol-derived molecules produced by solanaceous species. They contribute to pathogen defence but are toxic to humans and considered as anti-nutritional compounds. Here we show that GLYCOALKALOID METABOLISM 9 (GAME9), an APETALA2/Ethylene Response Factor, related to regulators of alkaloid production in tobacco and Catharanthus roseus, controls SGA biosynthesis. GAME9 knockdown and overexpression in tomato and potato alters expression of SGAs and upstream mevalonate pathway genes including the cholesterol biosynthesis gene STEROL SIDE CHAIN REDUCTASE 2 (SSR2). Levels of SGAs, C24-alkylsterols and the upstream mevalonate and cholesterol pathways intermediates are modified in these plants. Delta(7)-STEROL-C5(6)-DESATURASE (C5-SD) in the hitherto unresolved cholesterol pathway is a direct target of GAME9. Transactivation and promoter-binding assays show that GAME9 exerts its activity either directly or cooperatively with the SlMYC2 transcription factor as in the case of the C5-SD gene promoter. Our findings provide insight into the regulation of SGA biosynthesis and means for manipulating these metabolites in crops.

1 aCardenas, Pablo, D.1 aSonawane, Prashant, D.1 aPollier, Jacob1 aBossche, Robin, Vanden1 aDewangan, Veena1 aWeithorn, Efrat1 aTal, Lior1 aMeir, Sagit1 aRogachev, Ilana1 aMalitsky, Sergey1 aGiri, Ashok, P.1 aGoossens, Alain1 aBurdman, Saul1 aAharoni, Asaph uhttp://library.ncl.res.in/content/game9-regulates-biosynthesis-steroidal-alkaloids-and-upstream-isoprenoids-plant-mevalonate-102231nas a2200181 4500008004100000022001500041245010100056210006900157260000800226490000700234520156800241100001801809700002201827700002701849700002701876700002101903856012501924 2018 eng d a 0959-399300a Enhanced activity of Withania somnifera family-1 glycosyltransferase (UGT73A16) via mutagenesis0 aEnhanced activity of Withania somnifera family1 glycosyltransfer cOCT0 v343 aThis work used an approach of enzyme engineering towards the improved production of baicalin as well as alteration of acceptor and donor substrate preferences in UGT73A16. The 3D model of Withania somnifera family-1 glycosyltransferase (UGT73A16) was constructed based on the known crystal structures of plant UGTs. Structural and functional properties of UGT73A16 were investigated using docking and mutagenesis. The docking studies were performed to understand the key residues involved in substrate recognition. In the molecular model of UGT73A16, substrates binding pockets are located between N- and C-terminal domains. Modeled UGT73A16 was docked with UDP-glucose, UDP-glucuronic acid (UDPGA), kaempferol, isorhamnetin, 3-hydroxy flavones, naringenin, genistein and baicalein. The protein-ligand interactions showed that His 16, Asp 246, Lys 255, Ala 337, Gln 339, Val 340, Asn 358 and Glu 362 amino acid residues may be important for catalytic activity. The kinetic parameters indicated that mutants A337C and Q339A exhibited 2-3 fold and 6-7 fold more catalytic efficiency, respectively than wild type, and shifted the sugar donor specificity from UDP-glucose to UDPGA. The mutant Q379H displayed large loss of activity with UDP-glucose and UDPGA strongly suggested that last amino acid residue of PSPG box is important for glucuronosylation and glucosylation and highly specific to sugar binding sites. The information obtained from docking and mutational studies could be beneficial in future to engineer this biocatalyst for development of better ones.1 aSingh, Somesh1 aPatel, Krunal, A.1 aSonawane, Prashant, D.1 aVishwakarma, Rishi, K.1 aKhan, Bashir, M. uhttp://library.ncl.res.in/content/enhanced-activity-withania-somnifera-family-1-glycosyltransferase-ugt73a16-mutagenesis02365nas a2200397 4500008004100000022001400041245013600055210006900191260000800260300000900268490000700277520113700284100002401421700002701445700001601472700001801488700002001506700001901526700002101545700002201566700001701588700001601605700001601621700002201637700001601659700001801675700001701693700001801710700001601728700001601744700001701760700002301777700002001800700001901820856012801839 2019 eng d a2041-172300aPathways to defense metabolites and evading fruit bitterness in genus Solanum evolved through 2-oxoglutarate-dependent dioxygenases0 aPathways to defense metabolites and evading fruit bitterness in  cNOV a51690 v103 a

The genus Solanum comprises three food crops (potato, tomato, and eggplant), which are consumed on daily basis worldwide and also producers of notorious anti-nutritional steroidal glycoalkaloids (SGAs). Hydroxylated SGAs (i.e. leptinines) serve as precursors for leptines that act as defenses against Colorado Potato Beetle (Leptinotarsa decemlineata Say), an important pest of potato worldwide. However, SGA hydroxylating enzymes remain unknown. Here, we discover that 2-OXOGLUTARATE-DEPENDENT-DIOXYGENASE (2-ODD) enzymes catalyze SGA-hydroxylation across various Solanum species. In contrast to cultivated potato, Solanum chacoense, a widespread wild potato species, has evolved a 2-ODD enzyme leading to the formation of leptinines. Furthermore, we find a related 2-ODD in tomato that catalyzes the hydroxylation of the bitter a-tomatine to hydroxytomatine, the first committed step in the chemical shift towards downstream ripening-associated non-bitter SGAs (e.g. esculeoside A). This 2-ODD enzyme prevents bitterness in ripe tomato fruit consumed today which otherwise would remain unpleasant in taste and more toxic.

1 aCardenas, Pablo, D.1 aSonawane, Prashant, D.1 aHeinig, Uwe1 aJozwiak, Adam1 aPanda, Sayantan1 aAbebie, Bekele1 aKazachkova, Yana1 aPliner, Margarita1 aUnger, Tamar1 aWolf, Dalia1 aOfner, Itai1 aVilaprinyo, Ester1 aMeir, Sagit1 aDavydov, Olga1 aGal-On, Amit1 aBurdman, Saul1 aGiri, Ashok1 aZamir, Dani1 aScherf, Tali1 aSzymanski, Jedrzej1 aRogachev, Ilana1 aAharoni, Asaph uhttp://library.ncl.res.in/content/pathways-defense-metabolites-and-evading-fruit-bitterness-genus-solanum-evolved-through-202883nas a2200409 4500008004100000022001400041245012200055210006900177260000800246300001400254490000800268520160500276653001201881653002701893653003601920653002501956653001101981100002701992700001902019700002002038700002002058700001902078700002102097700002302118700001602141700001702157700001602174700001602190700002002206700002602226700002202252700002102274700002002295700001702315700001902332856012202351 2022 eng d a0028-646X00a2-oxoglutarate-dependent dioxygenases drive expansion of steroidal alkaloid structural diversity in the genus Solanum0 a2oxoglutaratedependent dioxygenases drive expansion of steroidal cMAY a1394-14100 v2343 a

Solanum steroidal glycoalkaloids (SGAs) are renowned defence metabolites exhibiting spectacular structural diversity. Genes and enzymes generating the SGA precursor pathway, SGA scaffold and glycosylated forms have been largely identified. Yet, the majority of downstream metabolic steps creating the vast repertoire of SGAs remain untapped. Here, we discovered that members of the 2-OXOGLUTARATE-DEPENDENT DIOXYGENASE (2-ODD) family play a prominent role in SGA metabolism, carrying out three distinct backbone-modifying oxidative steps in addition to the three formerly reported pathway reactions. The GLYCOALKALOID METABOLISM34 (GAME34) enzyme catalyses the conversion of core SGAs to habrochaitosides in wild tomato S. habrochaites. Cultivated tomato plants overexpressing GAME34 ectopically accumulate habrochaitosides. These habrochaitoside enriched plants extracts potently inhibit Puccinia spp. spore germination, a significant Solanaceae crops fungal pathogen. Another 2-ODD enzyme, GAME33, acts as a desaturase (via hydroxylation and E/F ring rearrangement) forming unique, yet unreported SGAs. Conversion of bitter alpha-tomatine to ripe fruit, nonbitter SGAs (e.g. esculeoside A) requires two hydroxylations; while the known GAME31 2-ODD enzyme catalyses hydroxytomatine formation, we find that GAME40 catalyses the penultimate step in the pathway and generates acetoxy-hydroxytomatine towards esculeosides accumulation. Our results highlight the significant contribution of 2-ODD enzymes to the remarkable structural diversity found in plant steroidal specialized metabolism.

10aSolanum10aspecialized metabolism10asteroidal glycoalkaloids (SGAs)10astructural diversity10aTomato1 aSonawane, Prashant, D.1 aJozawiak, Adam1 aBarbole, Ranjit1 aPanda, Sayantan1 aAbebie, Bekele1 aKazachkova, Yana1 aGharat, Sachin, A.1 aRamot, Ofir1 aUnger, Tamar1 aWizler, Guy1 aMeir, Sagit1 aRogachev, Ilana1 aDoron-Faigenboim, Adi1 aPetreikov, Marina1 aSchaffer, Arthur1 aGiri, Ashok, P.1 aScherf, Tali1 aAharoni, Asaph uhttp://library.ncl.res.in/content/2-oxoglutarate-dependent-dioxygenases-drive-expansion-steroidal-alkaloid-structural02801nas a2200445 4500008004100000022001400041245013400055210006900189260000800258300001400266490000800280520142600288653001901714653001201733653002101745653001101766653001901777653003601796653001101832100002001843700001801863700002701881700002301908700002101931700001901952700003101971700002602002700002002028700002002048700001802068700001602086700001602102700002002118700002202138700001702160700001802177700002002195700001902215856012102234 2022 eng d a0028-646X00aSteroidal alkaloids defence metabolism and plant growth are modulated by the joint action of gibberellin and jasmonate signalling0 aSteroidal alkaloids defence metabolism and plant growth are modu cFEB a1220-12370 v2333 aSteroidal glycoalkaloids (SGAs) are protective metabolites constitutively produced by Solanaceae species. Genes and enzymes generating the vast structural diversity of SGAs have been largely identified. Yet, mechanisms of hormone pathways coordinating defence (jasmonate; JA) and growth (gibberellin; GA) controlling SGAs metabolism remain unclear. We used tomato to decipher the hormonal regulation of SGAs metabolism during growth vs defence tradeoff. This was performed by genetic and biochemical characterisation of different JA and GA pathways components, coupled with in vitro experiments to elucidate the crosstalk between these hormone pathways mediating SGAs metabolism. We discovered that reduced active JA results in decreased SGA production, while low levels of GA or its receptor led to elevated SGA accumulation. We showed that MYC1 and MYC2 transcription factors mediate the JA/GA crosstalk by transcriptional activation of SGA biosynthesis and GA catabolism genes. Furthermore, MYC1 and MYC2 transcriptionally regulate the GA signalling suppressor DELLA that by itself interferes in JA-mediated SGA control by modulating MYC activity through protein-protein interaction. Chemical and fungal pathogen treatments reinforced the concept of JA/GA crosstalk during SGA metabolism. These findings revealed the mechanism of JA/GA interplay in SGA biosynthesis to balance the cost of chemical defence with growth.10aalpha-tomatine10adefence10agibberellin (GA)10agrowth10ajasmonate (JA)10asteroidal glycoalkaloids (SGAs)10aTomato1 aPanda, Sayantan1 aJozwiak, Adam1 aSonawane, Prashant, D.1 aSzymanski, Jedrzej1 aKazachkova, Yana1 aVainer, Andrii1 aKilambi, Himabindu, Vasuki1 aAlmekias-Siegl, Efrat1 aDikaya, Varvara1 aBocobza, Samuel1 aShohat, Hagai1 aMeir, Sagit1 aWizler, Guy1 aGiri, Ashok, P.1 aSchuurink, Robert1 aWeiss, David1 aYasuor, Hagai1 aKamble, Avinash1 aAharoni, Asaph uhttp://library.ncl.res.in/content/steroidal-alkaloids-defence-metabolism-and-plant-growth-are-modulated-joint-action02357nas a2200253 4500008004100000245011500041210006900156260000800225300000900233490000700242520149500249100002701744700002301771700001801794700002001812700002001832700002601852700001601878700002001894700002401914700002001938700001901958856012601977 2023 eng d00aBAHD-type acyltransferase concludes the biosynthetic pathway of non-bitter glycoalkaloids in ripe tomato fruit0 aBAHDtype acyltransferase concludes the biosynthetic pathway of n cJUL a45400 v143 a

During tomato fruit ripening, bitter and toxic steroidal glycoalkaloids (SGAs) are converted to nonbitter and less toxic forms, but proposed acylating enzyme in pathway remain unknown. Here, authors report BAHD-type acyltransferase that catalyze acylation step in biosynthesis of non-bitter SGAs in tomato. Tomato is the highest value fruit and vegetable crop worldwide, yet produces & alpha;-tomatine, a renowned toxic and bitter-tasting anti-nutritional steroidal glycoalkaloid (SGA) involved in plant defense. A suite of modifications during tomato fruit maturation and ripening converts & alpha;-tomatine to the non-bitter and less toxic Esculeoside A. This important metabolic shift prevents bitterness and toxicity in ripe tomato fruit. While the enzymes catalyzing glycosylation and hydroxylation reactions in the Esculeoside A pathway have been resolved, the proposed acetylating step remains, to date, elusive. Here, we discovered that GAME36 (GLYCOALKALOID METABOLISM36), a BAHD-type acyltransferase catalyzes SGA-acetylation in cultivated and wild tomatoes. This finding completes the elucidation of the core Esculeoside A biosynthetic pathway in ripe tomato, allowing reconstitution of Esculeoside A production in heterologous microbial and plant hosts. The involvement of GAME36 in bitter SGA detoxification pathway points to a key role in the evolution of sweet-tasting tomato as well as in the domestication and breeding of modern cultivated tomato fruit.

1 aSonawane, Prashant, D.1 aGharat, Sachin, A.1 aJozwiak, Adam1 aBarbole, Ranjit1 aHeinicke, Sarah1 aAlmekias-Siegl, Efrat1 aMeir, Sagit1 aRogachev, Ilana1 aO'Connor, Sarah, E.1 aGiri, Ashok, P.1 aAharoni, Asaph uhttp://library.ncl.res.in/content/bahd-type-acyltransferase-concludes-biosynthetic-pathway-non-bitter-glycoalkaloids-ripe02390nas a2200349 4500008004100000022001400041245005100055210005100106260000800157300000900165490000600174520140100180100001801581700002901599700001801628700002001646700001601666700001901682700002301701700002001724700001701744700001701761700001801778700002601796700002001822700003401842700001601876700002001892700002701912700001901939856008201958 2023 eng d a2055-026X00aParallel evolution of cannabinoid biosynthesis0 aParallel evolution of cannabinoid biosynthesis cMAY a817+0 v93 a

This study reveals a cannabinoid biosynthetic pathway in Helichrysum umbraculigerum, a plant genetically distant from Cannabis, providing a potential source of new cannabinoids and metabolic engineering tools. Modulation of the endocannabinoid system is projected to have therapeutic potential in almost all human diseases. Accordingly, the high demand for novel cannabinoids stimulates the discovery of untapped sources and efficient manufacturing technologies. Here we explored Helichrysum umbraculigerum, an Asteraceae species unrelated to Cannabis sativa that produces Cannabis-type cannabinoids (for example, 4.3% cannabigerolic acid). In contrast to Cannabis, cannabinoids in H. umbraculigerum accumulate in leaves' glandular trichomes rather than in flowers. The integration of de novo whole-genome sequencing data with unambiguous chemical structure annotation, enzymatic assays and pathway reconstitution in Nicotiana benthamiana and in Saccharomyces cerevisiae has uncovered the molecular and chemical features of this plant. Apart from core biosynthetic enzymes, we reveal tailoring ones producing previously unknown cannabinoid metabolites. Orthology analyses demonstrate that cannabinoid synthesis evolved in parallel in H. umbraculigerum and Cannabis. Our discovery provides a currently unexploited source of cannabinoids and tools for engineering in heterologous hosts.

1 aBerman, Paula1 ade Haro, Luis, Alejandro1 aJozwiak, Adam1 aPanda, Sayantan1 aPinkas, Zoe1 aDong, Younghui1 aCveticanin, Jelena1 aBarbole, Ranjit1 aLivne, Rotem1 aScherf, Tali1 aShimoni, Eyal1 aLevin-Zaidman, Smadar1 aDezorella, Nili1 aPetrovich-Kopitman, Ekaterina1 aMeir, Sagit1 aRogachev, Ilana1 aSonawane, Prashant, D.1 aAharoni, Asaph uhttp://library.ncl.res.in/content/parallel-evolution-cannabinoid-biosynthesis