A Stereoselective Strigolactone Biosynthesis Catalyzed by a 2-Oxoglutarate-Dependent Dioxygenase in Sorghum.

Seeds of root parasitic plants, Striga, Orobanche and Phelipanche spp., are induced to germinate by strigolactones (SLs) exudated from host roots. In Striga-resistant cultivars of Sorghum bicolor, the loss-of-function of the Low Germination Stimulant 1 (LGS1) gene changes the major SL from 5-deoxystrigol (5DS) to orobanchol, which has an opposite C-ring stereochemistry. The biosynthetic pathway of 5DS catalyzed by LGS1 has not been fully elucidated. Since other unknown regulators, in addition to LGS1 encoding a sulfotransferase, appear to be necessary for the stereoselective biosynthesis of 5DS, we examined Sobic.005G213500 (Sb3500), encoding a 2-oxoglutarate-dependent dioxygenase, as a candidate regulator, which is co-expressed with LGS1 and located 5'-upstream of LGS1 in the sorghum genome. When LGS1 was expressed with known SL biosynthetic enzyme genes including the cytochrome P450 SbMAX1a in Nicotiana benthamiana leaves, 5DS and its diastereomer 4-deoxyorobanchol (4DO) were produced in approximately equal amounts, while the production of 5DS was significantly larger than that of 4DO when Sb3500 was also co-expressed. We also confirmed the stereoselective 5DS production in an in vitro feeding experiment using synthetic chemicals with recombinant proteins expressed in Escherichia coli and yeast. This finding demonstrates that Sb3500 is a stereoselective regulator in the conversion of the SL precursor carlactone to 5DS, catalyzed by LGS1 and SbMAX1a, providing a detailed understanding of how different SLs are produced to combat parasitic weed infestations.

Structural analysis of a hormone-bound Striga strigolactone receptor.

Strigolactones (SLs) regulate many aspects of plant development, but ambiguities remain about how this hormone is perceived because SL-complexed receptor structures do not exist. We find that when SL binds the Striga receptor, ShHTL5, a series of conformational changes relative to the unbound state occur, but these events are not sufficient for signalling. Ligand-complexed receptors, however, form internal tunnels that posit an explanation for how SL exits its receptor after hydrolysis.

Late-Stage, Stereoretentive, and Site-Selective Modification of Synthetic Peptides by Using Photoredox Catalysis.

Late-stage functionalisation is an attractive method to generate peptide analogues containing non-natural residues. It is shown that cysteine residues can be activated as Crich-type thioethers, either by alkylation of a synthetic cysteine-continuing peptide or by incorporation of a modified cysteine unit into solid phase or solution phase peptide synthesis. Photoredox catalysed reaction of the thioether generates an alanyl radical intermediate in a stereoretentive and site-selective manner, even in the presence of free cysteine residues. The radical can react with non-activated alkenes to form non-natural residues bearing aliphatic, hydrophobic units. A method to avoid unwanted alkylation of amine residues was identified and the process was applied to the functionalization of both linear and cyclic synthetic peptides.

Generation and reaction of alanyl radicals in open flasks.

The generation and Giese-type reaction of alanyl radicals under metal-free reaction conditions is described. The procedure is operationally simple, occurring at ambient temperature in an open reaction vessel, and requiring short reaction times (≤5 min). The reaction occurs without epimerization and provides ready access to non-proteinogenic amino acids and peptides. Importantly, the process is tolerant of light absorbing groups including commonly used fluorescent tags.

A novel strigolactone receptor antagonist provides insights into the structural inhibition, conditioning, and germination of the crop parasite Striga.

Crop parasites of the Striga genera are a major biological deterrent to food security in Africa and are one of the largest obstacles to poverty alleviation on the continent. Striga seeds germinate by sensing small-molecule hormones, strigolactones (SLs), that emanate from host roots. Although SL receptors (Striga hermonthica HYPOSENSITIVE TO LIGHT [ShHTL]) have been identified, discerning their function has been difficult because these parasites cannot be easily grown under laboratory conditions. Moreover, many Striga species are obligate outcrossers that are not transformable, hence not amenable to genetic analysis. By combining phenotypic screening with ShHTL structural information and hybrid drug discovery methods, we discovered a potent SL perception inhibitor for Striga, dormirazine (DOZ). Structural analysis of this piperazine-based antagonist reveals a novel binding mechanism, distinct from that of known SLs, blocking access of the hormone to its receptor. Furthermore, DOZ reduces the flexibility of protein-protein interaction domains important for receptor signaling to downstream partners. In planta, we show, via temporal additions of DOZ, that SL receptors are required at a specific time during seed conditioning. This conditioning is essential to prime seed germination at the right time; thus, this SL-sensitive stage appears to be critical for adequate receptor signaling. Aside from uncovering a function for ShHTL during seed conditioning, these results suggest that future Ag-Biotech Solutions to Striga infestations will need to carefully time the application of antagonists to exploit receptor availability and outcompete natural SLs, critical elements for successful parasitic plant invasions.

Three mutations repurpose a plant karrikin receptor to a strigolactone receptor.

Uncovering the basis of small-molecule hormone receptors' evolution is paramount to a complete understanding of how protein structure drives function. In plants, hormone receptors for strigolactones are well suited to evolutionary inquiries because closely related homologs have different ligand preferences. More importantly, because of facile plant transgenic systems, receptors can be swapped and quickly assessed functionally in vivo. Here, we show that only three mutations are required to turn the nonstrigolactone receptor, KAI2, into a receptor that recognizes the plant hormone strigolactone. This modified receptor still retains its native function to perceive KAI2 ligands. Our directed evolution studies indicate that only a few keystone mutations are required to increase receptor promiscuity of KAI2, which may have implications for strigolactone receptor evolution in parasitic plants.

Generation and reaction of alkyl radicals in open reaction vessels.

An operationally simple process to transform alkyl iodides into reactive alkyl radicals is described. Aryl diazonium salts react with Hantzsch esters and molecular oxygen to give aryl radicals, which participate in halogen atom transfers to give alkyl radicals. These intermediates react with a variety of acceptors. The reaction cascade occurs at room temperature, in open reaction vessels, with short reaction times.

SMAX1-dependent seed germination bypasses GA signalling in Arabidopsis and Striga.

Parasitic plant infestations dramatically reduce the yield of many major food crops of sub-Saharan Africa and pose a serious threat to food security on that continent1. The first committed step of a successful infestation is the germination of parasite seeds primarily in response to a group of related small-molecule hormones called strigolactones (SLs), which are emitted by host roots2. Despite the important role of SLs, it is not clear how host-derived SLs germinate parasitic plants. In contrast, gibberellins (GA) acts as the dominant hormone for stimulation of germination in non-parasitic plant species by inhibiting a set of DELLA repressors3. Here, we show that expression of SL receptors from the parasitic plant Striga hermonthica in the presence of SLs circumvents the GA requirement for germination of Arabidopsis thaliana seed. Striga receptors co-opt and enhance signalling through the HYPOSENSITIVE TO LIGHT/KARRIKIN INSENSITIVE 2 (AtHTL/KAI2) pathway, which normally plays a rudimentary role in Arabidopsis seed germination4,5. AtHTL/KAI2 negatively controls the SUPPRESSOR OF MAX2 1 (SMAX1) protein5, and loss of SMAX1 function allows germination in the presence of DELLA repressors. Our data suggest that ligand-dependent inactivation of SMAX1 in Striga and Arabidopsis can bypass GA-dependent germination in these species.

σ-Bond initiated generation of aryl radicals from aryl diazonium salts.

σ-Bond nucleophiles and molecular oxygen transform aryl diazonium salts into aryl radicals. Experimental and computational studies show that Hantzsch esters transfer hydride to aryl diazonium species, and that oxygen initiates radical fragmentation of the diazene intermediate to produce aryl radicals. The operational simplicity of this addition-fragmentation process for the generation of aryl radicals, by a polar-radical crossover mechanism, has been illustrated in a variety of bond-forming reactions.

Hyperconjomer stereocontrol of cationic polyene cyclisations.

Polyene cyclisations are a powerful method for the direct generation of molecular complexity. This paper describes the use of computational methods to investigate the stereoselectivity of cationic polyene cyclisations of geranylbenzene derivatives. The outcomes highlight the different reactivity of hyperconjomers during the key Friedel-Crafts alkylation step, and informed a successful strategy for the synthesis of (±)-taiwaniaquinone G with improved levels of stereoselectivity.

Total Synthesis and Stereochemical Confirmation of Heliolactone.

Until now, the relative stereochemistry of the noncanonical strigolactone, heliolactone, has remained ambiguous. The total synthesis of heliolactone is described, with the key bond-forming event being a Stille cross-coupling that relied upon a reversal of the nucleophile-electrophile coupling partners. Spectroscopic analysis of synthetic heliolactone (and other stereoisomers) and comparisons with the isolated material enabled the absolute and relative stereochemistry of heliolactone to be secured.

Visible-Light-Driven MADIX Polymerisation via a Reusable, Low-Cost, and Non-Toxic Bismuth Oxide Photocatalyst.

The continuous amalgamation of photocatalysis into existing reversible deactivation radical polymerisation (RDRP) processes has initiated a rapidly propagating area of polymer research in recent years. We introduce bismuth oxide (Bi2 O3 ) as a heterogeneous photocatalyst for polymerisations, operating at room temperature with visible light. We demonstrate formidable control over degenerative chain-transfer polymerisations, such as macromolecular design by interchange of xanthate (MADIX) and reversible addition-fragmentation chain-transfer (RAFT) polymerisation. We achieved narrow molecular weight distributions and attribute the excellent temporal control of a photo-induced electron transfer (PET) process. This methodology was employed to synthesise diblock copolymers combining differently activated monomers. The Bi2 O3 catalyst system has the additional benefits of low toxicity, reusability, low-cost, and ease of removal from the reaction mixture.

Which are the major players, canonical or non-canonical strigolactones?

Strigolactones (SLs) can be classified into two structurally distinct groups: canonical and non-canonical SLs. Canonical SLs contain the ABCD ring system, and non-canonical SLs lack the A, B, or C ring but have the enol ether-D ring moiety, which is essential for biological activities. The simplest non-canonical SL is the SL biosynthetic intermediate carlactone. In plants, carlactone and its oxidized metabolites, such as carlactonoic acid and methyl carlactonoate, are present in root and shoot tissues. In some plant species, including black oat (Avena strigosa), sunflower (Helianthus annuus), and maize (Zea mays), non-canonical SLs in the root exudates are major germination stimulants. Various plant species, such as tomato (Solanum lycopersicum), Arabidopsis, and poplar (Populus spp.), release carlactonoic acid into the rhizosphere. These observations suggest that both canonical and non-canonical SLs act as host-recognition signals in the rhizosphere. In contrast, the limited distribution of canonical SLs in the plant kingdom, and the structure-specific and stereospecific transportation of canonical SLs from roots to shoots, suggest that plant hormones inhibiting shoot branching are not canonical SLs but, rather, are non-canonical SLs.

Methyl zealactonoate, a novel germination stimulant for root parasitic weeds produced by maize.

One of the germination stimulants for root parasitic weeds produced by maize (Zea mays) was isolated and named methyl zealactonoate (1). Its structure was determined to be methyl (2E,3E)-4-((RS)-3,3-dimethyl-2-(3-methylbut-2-en-2-yl)-5-oxotetrahydrofuran-2-yl)-2-((((R)-4-methyl-5-oxo-2,5-dihydrofran-2-yl)oxy)methylene)but-3-enoate using by 1D and 2D NMR spectroscopy and ESI and EI-MS spectrometry. Feeding experiments with 13C-carlactone (CL), a biosynthetic intermediate for strigolactones, confirmed that 1 is produced from CL in maize. Methyl zealactonoate strongly elicits Striga hermonthica and Phelipanche ramosa seed germination, while Orobanche minor seeds are 100-fold less sensitive to this stimulant.

Total Synthesis of (+)-Panacene.

The first total synthesis of the naturally occurring enantiomer of the marine bromoallene (+)-panacene is described. Central to this concise enantioselective synthesis was the use of a Noyori transfer hydrogenation for a Dynamic Kinetic Resolution (DKR) that set the desired absolute stereochemistry. A highly stereoselective Julia coupling was then used to install a Z-configured enyne, which enabled the biomimetic construction of the axially chiral bromoallene.

Expression of MAX2 under SCARECROW promoter enhances the strigolactone/MAX2 dependent response of Arabidopsis roots to low-phosphate conditions.

MAIN CONCLUSION: MAX2/strigolactone signaling in the endodermis and/or quiescent center of the root is partially sufficient to exert changes in F-actin density and cellular trafficking in the root epidermis, and alter gene expression during plant response to low Pi conditions. Strigolactones (SLs) are a new group of plant hormones that regulate different developmental processes in the plant via MAX2, an F-box protein that interacts with their receptor. SLs and MAX2 are necessary for the marked increase in root-hair (RH) density in seedlings under conditions of phosphate (Pi) deprivation. This marked elevation was associated with an active reduction in actin-filament density and endosomal movement in root epidermal cells. Also, expression of MAX2 under the SCARECROW (SCR) promoter was sufficient to confer SL sensitivity in roots, suggesting that SL signaling pathways act through a root-specific, yet non-cell-autonomous regulatory mode of action. Here we show evidence for a non-cell autonomous signaling of SL/MAX2, originating from the root endodermis, and necessary for seedling response to conditions of Pi deprivation. SCR-derived expression of MAX2 in max2-1 mutant background promoted the root low Pi response, whereas supplementation of the synthetic SL GR24 to these SCR:MAX2 expressing lines further enhanced this response. Moreover, the SCR:MAX2 expression led to changes in actin density and endosome movement in epidermal cells and in TIR1 and PHO2 gene expression. These results demonstrate that MAX2 signaling in the endodermis and/or quiescent center is partially sufficient to exert changes in F-actin density and cellular trafficking in the epidermis, and alter gene expression under low Pi conditions.

Arabidopsis response to low-phosphate conditions includes active changes in actin filaments and PIN2 polarization and is dependent on strigolactone signalling.

Strigolactones (SLs) are plant hormones that regulate the plant response to phosphate (Pi) growth conditions. At least part of SL-signalling execution in roots involves MAX2-dependent effects on PIN2 polar localization in the plasma membrane (PM) and actin bundling and dynamics. We examined PIN2 expression, PIN2 PM localization, endosome trafficking, and actin bundling under low-Pi conditions: a MAX2-dependent reduction in PIN2 trafficking and polarization in the PM, reduced endosome trafficking, and increased actin-filament bundling were detected in root cells. The intracellular protein trafficking that is related to PIN proteins but unassociated with AUX1 PM localization was selectively inhibited. Exogenous supplementation of the synthetic SL GR24 to a SL-deficient mutant (max4) led to depletion of PIN2 from the PM under low-Pi conditions. Accordingly, roots of mutants in MAX2, MAX4, PIN2, TIR3, and ACTIN2 showed a reduced low-Pi response compared with the wild type, which could be restored by auxin (for all mutants) or GR24 (for all mutants except max2-1). Changes in PIN2 polarity, actin bundling, and vesicle trafficking may be involved in the response to low Pi in roots, dependent on SL/MAX2 signalling.

Avenaol, a germination stimulant for root parasitic plants from Avena strigosa.

Root exudates from the allelopathic plant, black oat (Avena strigosa Schreb.), were found to contain at least six different germination stimulants for root parasitic plants, but no known strigolactones (SLs). One of these germination stimulants was purified and named avenaol. Its HR-ESI-TOFMS analysis indicated that the molecular formula of avenaol is C20H24O7, and thus it contains an additional carbon compared with known C19-SLs. Its structure was determined as 5-((E)-(5-(3-hydroxy-1,5,5-trimethyl-2-oxobicyclo[4.1.0]heptan-7-yl)-2-oxodihydrofuran-3(2H)-ylidene)methoxy)-3-methylfuran-2(5H)-one, by 1D and 2D NMR spectroscopy, and ESI- and EI-MS spectrometry. Although avenaol contains the C-D moiety, the common structural feature for all known SLs, it lacks the B ring and has an additional carbon atom between the A and C rings. Avenaol is a potent germination stimulant of Phelipanche ramosa seeds, but only a weak stimulant for seeds of Striga hermonthica and Orobanche minor.

Enantioselective synthesis of the strigolactone mimic (+)-GR24.

A short and cost-effective synthesis of the important strigolactone analogue (+)-GR24 is described. Central to this new approach is the concise, enantioselective synthesis of the A-C ring system.