Sequential regiospecific gem-diprenylation of tetrahydroxyxanthone by prenyltransferases from Hypericum sp.

Polyprenylated acylphloroglucinol derivatives, such as xanthones, are natural plant products with interesting pharmacological properties. They are difficult to synthesize chemically. Biotechnological production is desirable but it requires an understanding of the biosynthetic pathways. cDNAs encoding membrane-bound aromatic prenyltransferase (aPT) enzymes from Hypericum sampsonii seedlings (HsPT8px and HsPTpat) and Hypericum calycinum cell cultures (HcPT8px and HcPTpat) were cloned and expressed in Saccharomyces cerevisiae and Nicotiana benthamiana, respectively. Microsomes and chloroplasts were used for functional analysis. The enzymes catalyzed the prenylation of 1,3,6,7-tetrahydroxyxanthone (1367THX) and/or 1,3,6,7-tetrahydroxy-8-prenylxanthone (8PX) and discriminated nine additionally tested acylphloroglucinol derivatives. The transient expression of the two aPT genes preceded the accumulation of the products in elicitor-treated H. calycinum cell cultures. C-terminal yellow fluorescent protein fusions of the two enzymes were localized to the envelope of chloroplasts in N. benthamiana leaves. Based on the kinetic properties of HsPT8px and HsPTpat, the enzymes catalyze sequential rather than parallel addition of two prenyl groups to the carbon atom 8 of 1367THX, yielding gem-diprenylated patulone under loss of aromaticity of the gem-dialkylated ring. Coexpression in yeast significantly increased product formation. The patulone biosynthetic pathway involves multiple subcellular compartments. The aPTs studied here and related enzymes may be promising tools for plant/microbe metabolic pathway engineering.

Biotechnological production of hyperforin for pharmaceutical formulation.

Hyperforin is a major active constituent of Hypericum perforatum (St. John's wort). It has amazing pharmacological activities, such as antidepressant properties, but it is labile and difficult to synthesize. Its sensitivity and lipophilicity are challenges for processing and formulation. Its chemical complexity provokes approaches of biotechnological production and modification. Dedifferentiated H. perforatum cell cultures lack appropriate storage sites and hence appreciable hyperforin levels. Shoot cultures are capable of forming hyperforin but less suitable for biomass up-scaling in bioreactors. Roots commonly lack hyperforin but a recently established adventitious root line has been demonstrated to produce hyperforin and derivatives at promising levels. The roots also contained lupulones, the typical constituents of hop (Humulus lupulus). Although shear-sensitive, these root cultures provide a potential production platform for both individual compounds and extracts with novel combinations of constituents and pharmacological activities. Besides in vitro cultivation techniques, the reconstruction of hyperforin biosynthesis in microorganisms is a promising alternative for biotechnological production. The biosynthetic pathway is under study, with omics-technologies being increasingly implemented. These biotechnological approaches may not only yield hyperforin at reasonable productivity but also allow for modifications of its chemical structure and pharmacological profile.

An ABC Transporter Is Involved in the Silicon-Induced Formation of Casparian Bands in the Exodermis of Rice.

Silicon (Si) promotes the formation of Casparian bands (CB) in rice and reduces radial oxygen loss (ROL). Further transcriptomic approaches revealed several candidate genes involved in the Si-induced formation of CB such as ATP binding cassette (ABC) transporter, Class III peroxidases, ligases and transferases. Investigation of these genes by means of overexpression (OE) and knockout (KO) mutants revealed the contribution of the ABC transporter (OsABCG25) to CB formation in the exodermis, which was also reflected in the expression of other OsABCG25 in the Si-promoted formation of CB genes related to the phenylpropanoid pathway, such as phenylalanine-ammonia-lyase, diacylglycerol O-acyltransferase and 4-coumarate-CoA ligase. Differential CB development in mutants and Si supply also affected the barrier function of the exodermis. OE of the ABC transporter and Si supply reduced the ROL from roots and Fe uptake. No effect on ROL and Fe uptake could be observed for the KO mutant. The presented research confirms the impact of the OsABCG25 in the Si-promoted formation of CB and its barrier functions.