Striga hermonthica Suicidal Germination Activity of Potent Strigolactone Analogs: Evaluation from Laboratory Bioassays to Field Trials.

The obligate hemiparasite Striga hermonthica is one of the major global biotic threats to agriculture in sub-Saharan Africa, causing severe yield losses of cereals. The germination of Striga seeds relies on host-released signaling molecules, mainly strigolactones (SLs). This dependency opens up the possibility of deploying SL analogs as "suicidal germination agents" to reduce the accumulated seed bank of Striga in infested soils. Although several synthetic SL analogs have been developed for this purpose, the utility of these compounds in realizing the suicidal germination strategy for combating Striga is still largely unknown. Here, we evaluated the efficacy of three potent SL analogs (MP3, MP16, and Nijmegen-1) under laboratory, greenhouse, and farmer's field conditions. All investigated analogs showed around a 50% Striga germination rate, equivalent to a 50% reduction in infestation, which was comparable to the standard SL analog GR24. Importantly, MP16 had the maximum reduction of Striga emergence (97%) in the greenhouse experiment, while Nijmegen-1 appeared to be a promising candidate under field conditions, with a 43% and 60% reduction of Striga emergence in pearl millet and sorghum fields, respectively. These findings confirm that the selected SL analogs appear to make promising candidates as simple suicidal agents both under laboratory and real African field conditions, which may support us to improve suicidal germination technology to deplete the Striga seed bank in African agriculture.

A New Formulation for Strigolactone Suicidal Germination Agents, towards Successful Striga Management.

Striga hermonthica, a member of the Orobanchaceae family, is an obligate root parasite of staple cereal crops, which poses a tremendous threat to food security, contributing to malnutrition and poverty in many African countries. Depleting Striga seed reservoirs from infested soils is one of the crucial approaches to minimize subterranean damage to crops. The dependency of Striga germination on the host-released strigolactones (SLs) has prompted the development of the "Suicidal Germination" strategy to reduce the accumulated seed bank of Striga. The success of aforementioned strategy depends not only on the activity of the applied SL analogs, but also requires suitable application protocol with simple, efficient, and handy formulation for rain-fed African agriculture. Here, we developed a new formulation "Emulsifiable Concentration (EC)" for the two previously field-assessed SL analogs Methyl phenlactonoate 3 (MP3) and Nijmegen-1. The new EC formulation was evaluated for biological activities under lab, greenhouse, mini-field, and field conditions in comparison to the previously used Atlas G-1086 formulation. The EC formulation of SL analogs showed better activities on Striga germination with lower EC50 and high stability under Lab conditions. Moreover, EC formulated SL analogs at 1.0 µM concentrations reduced 89-99% Striga emergence in greenhouse. The two EC formulated SL analogs showed also a considerable reduction in Striga emergence in mini-field and field experiments. In conclusion, we have successfully developed a desired formulation for applying SL analogs as suicidal agents for large-scale field application. The encouraging results presented in this study pave the way for integrating the suicidal germination approach in sustainable Striga management strategies for African agriculture.

Synthesis of Analogs of Strigolactones and Evaluation of Their Stability in Solution.

Strigolactones (SLs) are new plant hormones that play an important role in the control development of plants. They are germination stimulants for seed of parasitic weeds, are the branching factor of arbuscular mycorrhizal fungi and inhibitors for bud outgrowth and shoot branching. Natural SLs contain an annulated system of three rings (ABC scaffold) connected to a furanone (the D-ring) by an enol ether unit. The natural distribution of strigolactones is low, and their synthesis is long and difficult. Therefore, SL analogs are designed to have the same bioactiphore as natural SLs and an appreciable bioactivity. For the design a model is used based on the natural bioactiphore. Typical SL analogs are GR24, Nijmegen-1, and EM1 (derived from ethyl 2-phenylacetate). The synthesis of these SL analogs is reported together with their stability in aqueous solution.

Hybrid-type strigolactone analogues derived from auxins.

BACKGROUND: Strigolactones (SLs) have a vast number of ecological implications because of the broad spectrum of their biological activities. Unfortunately, the limited availability of SLs restricts their applicability for the benefit of humanity and renders synthesis the only option for their production. However, the structural complexity of SLs impedes their economical synthesis, which is unfeasible on a large scale. Synthesis of SL analogues and mimics with a simpler structure, but with retention of bioactivity, is the solution to this problem. RESULTS: Here, we present eight new hybrid-type SL analogues derived from auxin, synthesized via coupling of auxin ester [ethyl 2-(1H-indol-3-yl)acetate] and of ethyl 2-phenylacetate with four D-rings (mono-, two di- and trimethylated). The new hybrid-type SL analogues were bioassayed to assess the germination activity of seeds of the parasitic weeds Striga hermonthica, Orobanche minor and Phelipanche ramosa using the classical method of counting germinated seeds and a colorimetric method. The bioassays revealed that analogues with a natural monomethylated D-ring had appreciable to good activity towards the three species and were the most active derivatives. By contrast, derivatives with the trimethylated D-ring showed no activity. The dimethylated derivatives (2,4-dimethyl and 3,4-dimethyl) were slightly active, especially towards P. ramosa. CONCLUSIONS: New hybrid-type analogues derived from auxins have been prepared. These analogues may be attractive as potential suicidal germination agents for parasitic weed control because of their ease of preparation and relevant bioactivity. © 2019 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

New hybrid type strigolactone mimics derived from plant growth regulator auxin.

Strigolactones (SLs) constitute a new class of plant hormones of increasing importance in plant science. The structure of natural SLs is too complex for ready access by synthesis. Therefore, much attention is being given to design of SL analogues and mimics with a simpler structure but with retention of bioactivity. Here new hybrid type SL mimics have been designed derived from auxins, the common plant growth regulators. Auxins were simply coupled with the butenolide D-ring using bromo (or chloro) butenolide. D-rings having an extra methyl group at the vicinal C-3' carbon atom, or at the C-2' carbon atom, or at both have also been studied. The new hybrid type SL mimics were bioassayed for germination activity of seeds of the parasitic weeds S. hermonthica, O. minor and P. ramosa using the classical method of counting germinated seeds and a colorimetric method. For comparison SL mimics derived from phenyl acetic acid were also investigated. The bioassays revealed that mimics with a normal D-ring had appreciable to good activity, those with an extra methyl group at C-2' were also appreciably active, whereas those with a methyl group in the vicinal C-3' position were inactive (S. hermonthica) or only slightly active. The new hybrid type mimics may be attractive as potential suicidal germination agents in agronomic applications.

Strigolactones: new plant hormones in the spotlight.

The development and growth of plants are regulated by interplay of a plethora of complex chemical reactions in which plant hormones play a pivotal role. In recent years, a group of new plant hormones, namely strigolactones (SLs), was discovered and identified. The first SL, strigol, was isolated in 1966, but it took almost 20 years before the details of its structure were fully elucidated. At present, two families of SLs are known, one having the stereochemistry of (+)-strigol and the other that of (-)-orobanchol, the most abundant naturally occurring SL. The most well-known bioproperty of SLs is the germination of seeds of the parasitic weeds Striga and Orobanche. It is evident that SLs are going to play a prominent role in modern molecular botany. In this review, relevant molecular and bioproperties of SLs are discussed. Items of importance are the effect of stereochemistry, structure-activity relationships, design and synthesis of analogues with a simple structure, but with retention of bioactivity, introduction of fluorescent labels into SLs, biosynthetic origin of SLs, mode of action in plants, application in agriculture for the control of parasitic weeds, stimulation of the branching of arbuscular mycorrhizal (AM) fungi, and the control of plant architecture. The future potential of SLs in molecular botany is highlighted.

Stability of strigolactone analog GR24 toward nucleophiles.

BACKGROUND: Strigolactones (SLs) are plant hormones that play various roles in plant development. The chemical stability of SLs depends on the solvent, the pH, and the presence of nucleophiles. Hydrolysis leads to detachment of the butenolide ring, and plays a crucial role in the initial stages of the signal-transduction process occurring between the receptor and the SL signaling molecule. RESULTS: To date, two different mechanisms have been proposed for SL hydrolysis. Results obtained from kinetic, thermodynamic, and mass spectral data for the reaction between the widely used synthetic SL analog GR24 and seven different nucleophiles demonstrate that the reaction proceeds via the Michael addition-elimination mechanism. CONCLUSION: This study provides valuable information on the chemical stability of GR24 in different plant growth media and buffers. Such information is valuable for scientists using GR24 treatments to study SL-regulated processes in plants. © 2017 Society of Chemical Industry.

Strigolactone Analogues with a D-Ring Modified at C-2.

Strigolactones (SLs) are important new plant hormones that receive much attention in current plant science. SLs are produced by many plants and are exuded by the root system. SLs are, amongst others, germination stimulants for seed of parasitic weeds. Naturally occurring SLs invariably contain three annelated rings, the ABC-scaffold, connected to a butenolide (the D-ring) via an enol ether unit. The synthesis of natural SLs requires many steps, therefore there is a continuous search for SL analogues with a simpler structure but with retention of bioactivity. In this study modified D-ring variants are investigated, especially analogues having a methyl group at C-2 instead of a hydrogen. For these analogues the ABC-scaffolds of GR24 and Nijmegen-1 were used. The coupling reaction proceeds profoundly better with chlorobutenolides than with the corresponding bromides. Bioassays reveal that the introduction an extra methyl at C-2 does not influence the germination activity, which is relevant for gaining insight in the mode of action of SLs.

Strigolactones: new plant hormones in action.

MAIN CONCLUSION: The key step in the mode of action of strigolactones is the enzymatic detachment of the D-ring. The thus formed hydroxy butenolide induces conformational changes of the receptor pocket which trigger a cascade of reactions in the signal transduction. Strigolactones (SLs) constitute a new class of plant hormones which are of increasing importance in plant science. For the last 60 years, they have been known as germination stimulants for parasitic plants. Recently, several new bio-properties of SLs have been discovered such as the branching factor for arbuscular mycorrhizal fungi, regulation of plant architecture (inhibition of bud outgrowth and of shoot branching) and the response to abiotic factors, etc. To broaden horizons and encourage new ideas for identifying and synthesising new and structurally simple SLs, this review is focused on molecular aspects of this new class of plant hormones. Special attention has been given to structural features, the mode of action of these phytohormones in various biological actions, the design of SL analogs and their applications.

Suicidal germination for parasitic weed control.

Parasitic weeds of the genera Striga and Orobanche spp. cause severe yield losses in agriculture, especially in developing countries and the Mediterranean. Seeds of these weeds germinate by a chemical signal exuded by the roots of host plants. The radicle thus produced attaches to the root of the host plant, which can then supply nutrients to the parasite. There is an urgent need to control these weeds to ensure better agricultural production. The naturally occurring chemical signals are strigolactones (SLs), e.g. strigol and orobanchol. One option to control these weeds involves the use of SLs as suicidal germination agents, where germination takes place in the absence of a host. Owing to the lack of nutrients, the germinated seeds will die. The structure of natural SLs is too complex to allow multigram synthesis. Therefore, SL analogues are developed for this purpose. Examples are GR24 and Nijmegen-1. In this paper, the SL analogues Nijmegen-1 and Nijmegen-1 Me were applied in the field as suicidal germination agents. Both SL analogues were formulated using an appropriate EC-approved emulsifier (polyoxyethylene sorbitol hexaoleate) and applied to tobacco (Nicotiana tabacum L.) fields infested by Orobanche ramosa L. (hemp broomrape), following a strict protocol. Four out of 12 trials showed a reduction in broomrape of ≥95%, two trials were negative, two showed a moderate result, one was unclear and in three cases there was no Orobanche problem in the year of the trials. The trial plots were ca 2000 m2 ; half of that area was treated with stimulant emulsion, the other half was not treated. The optimal amount of stimulant was 6.25 g ha-1 . A preconditioning prior to the treatment was a prerequisite for a successful trial. In conclusion, the suicidal germination approach to reducing O. ramosa in tobacco fields using formulated SL analogues was successful. Two other options for weed control are discussed: deactivation of stimulants prior to action and biocontrol by Fusarium oxysporum. © 2016 Society of Chemical Industry.

Synthesis of strigolactones, a strategic account.

Strigolactones (SLs) constitute a new class of plant hormones that have received growing interest in recent years. They firstly became known as signalling molecules for host recognition by parasitic plants, and for symbiosis of plants with arbuscular mycorrhizal fungi. Furthermore, they are involved in numerous physiological processes in plants, such as the regulation of plant architecture and the response to abiotic factors. SLs are produced by plants in extremely low quantities, and they may be unstable during the purification process. Therefore, their total synthesis is highly relevant for confirming the structures assigned on the basis of spectroscopic and other physical data. A second important theme in SL research is the design and synthesis of SL analogues that have a simplified structure while still featuring the essential bioproperties. This review summarises the strategy and synthesis of naturally occurring SLs, and the design and synthesis of SL analogues with appreciable bioactivity.

Rice cytochrome P450 MAX1 homologs catalyze distinct steps in strigolactone biosynthesis.

Strigolactones (SLs) are a class of phytohormones and rhizosphere signaling compounds with high structural diversity. Three enzymes, carotenoid isomerase DWARF27 and carotenoid cleavage dioxygenases CCD7 and CCD8, were previously shown to convert all-trans-ß-carotene to carlactone (CL), the SL precursor. However, how CL is metabolized to SLs has remained elusive. Here, by reconstituting the SL biosynthetic pathway in Nicotiana benthamiana, we show that a rice homolog of Arabidopsis More Axillary Growth 1 (MAX1), encodes a cytochrome P450 CYP711 subfamily member that acts as a CL oxidase to stereoselectively convert CL into ent-2'-epi-5-deoxystrigol (B-C lactone ring formation), the presumed precursor of rice SLs. A protein encoded by a second rice MAX1 homolog then catalyzes the conversion of ent-2'-epi-5-deoxystrigol to orobanchol. We therefore report that two members of CYP711 enzymes can catalyze two distinct steps in SL biosynthesis, identifying the first enzymes involved in B-C ring closure and a subsequent structural diversification step of SLs.

New strigolactone mimics: structure-activity relationship and mode of action as germinating stimulants for parasitic weeds.

Strigolactones (SLs) are new plant hormones with varies important bio-functions. This Letter deals with germination of seeds of parasitic weeds. Natural SLs have a too complex structure for synthesis. Therefore, there is an active search for SL analogues and mimics with a simpler structure with retention of activity. SL analogues all contain the D-ring connected with an enone moiety through an enol ether unit. A new mechanism for the hydrolysis SL analogues involving bidentate bound water and an α,ß-hydrolase with a Ser-His-Asp catalytic triad has been proposed. Newly discovered SL mimics only have the D-ring with an appropriate leaving group at C-5. A mode of action for SL mimics was proposed for which now supporting evidence is provided. As predicted an extra methyl group at C-4 of the D-ring blocks the germination of seeds of parasitic weeds.

Structure and activity of strigolactones: new plant hormones with a rich future.

Strigolactones (SLs) constitute a new class of plant hormones which are active as germination stimulants for seeds of parasitic weeds of Striga, Orobanche, and Pelipanchi spp, in hyphal branching of arbuscular mycorrhizal (AM) fungi and as inhibitors of shoot branching. In this review, the focus is on molecular features of these SLs. The occurrence of SLs in root exudates of host plants is described. The naming protocol for SL according to the International Union of Pure and Applied Chemistry (IUPAC) rules and the 'at a glance' method is explained. The total synthesis of some natural SLs is described with details for all eight stereoisomers of strigol. The problems encountered with assigning the correct structure of natural SLs are analyzed for orobanchol, alectrol, and solanacol. The structure-activity relationship of SLs as germination stimulants leads to the identification of the bioactiphore of SLs. Together with a tentative mechanism for the mode of action, a model has been derived that can be used to design and prepare active SL analogs. This working model has been used for the preparation of a series of new SL analogs such as Nijmegen-1, and analogs derived from simple ketones, keto enols, and saccharine. The serendipitous finding of SL mimics which are derived from the D-ring in SLs (appropriately substituted butenolides) is reported. For SL mimics, a mode of action is proposed as well. Recent new results support this proposal. The stability of SLs and SL analogs towards hydrolysis is described and some details of the mechanism of hydrolysis are discussed as well. The attempted isolation of the protein receptor for germination and the current status concerning the biosynthesis of natural SLs are briefly discussed. Some non-SLs as germinating agents are mentioned. The structure-activity relationship for SLs in hyphal branching of AM fungi and in repression of shoot branching is also analyzed. For each of the principle functions, a working model for the design of new active SL analogs is described and its applicability and implications are discussed. It is shown that the three principal functions use a distinct perception system. The importance of stereochemistry for bioactivity has been described for the various functions.

Strigolactone analogues and mimics derived from phthalimide, saccharine, p-tolylmalondialdehyde, benzoic and salicylic acid as scaffolds.

A series of new strigolactone (SL) analogues is derived from simple and cheap starting materials. These SL analogues are designed using a working model. The first analogue is a modified Nijmegen-1, the second contains saccharin as substituent (bio-isosteric replacement of a carbonyl in Nijmegen-1 by a sulfonyl group) and the third one is derived from p-tolylmalondialdehyde. These new SL analogues are appreciably to highly active as germination stimulants of seeds of Striga hermonthica and Orobanche cernua. The SL analogue derived from saccharin is the most active one. A serendipitous and most rewarding finding is that the compound obtained by a direct coupling of saccharin with the chlorobutenolide exhibits a high germination activity especially towards O. cernua seeds. Two other SL mimics are obtained from benzoic and salicylic aid by a direct coupling reaction with chlorobutenolide, both of them are very active germinating agents. These SL mimics represent a new type of germination stimulants. A tentative molecular mechanism for the mode of action of these SL mimics has been proposed.

Single step synthesis of strigolactone analogues from cyclic keto enols, germination stimulants for seeds of parasitic weeds.

The single step synthesis of a newly designed series of strigolactones (SLs) from cyclic keto enols is described. The germinating activity of these SL analogues towards seeds of the parasitic weeds Striga and Orobanche spp. is reported. The first of these SL analogues are derived from the hydroxyl γ-pyrones kojic acid and maltol, the second type from hydroxyl α-pyrones, namely, 4-hydroxy-6-methyl-2H-pyran-2-one and 4-hydroxy-coumarin and the third type from 1,3-diketones, namely, 1,3-cyclohexane-dione (dimedone) and tricyclic 1,3-dione. All keto enols are coupled in a single step with the appropriate D-ring precursor in the presence of a base to give the desired SL analogues. All SL analogues are acceptably biologically active in inducing the germination of seeds of Striga hermonthica and Orobanchecernua. Most interesting are the analogues derived from 4-hydroxy coumarin and dimedone, as they have a remarkably high biological activity towards the seeds of parasitic weeds at relatively low concentrations, comparable with that of the general standard stimulant GR24.

Strigolactone analogs derived from ketones using a working model for germination stimulants as a blueprint.

Strigolactones are important signaling compounds in the plant kingdom. Here we focus on their germination stimulatory effect on seeds of the parasitic weeds Striga and Orobanche spp. and more particularly on the design and synthesis of new active strigolactone analogs derived from simple cyclic ketones. New analogs derived from 1-indanone, 1-tetralone, cyclopentanone, cyclohexanone and a series of substituted cyclohexanones (including carvone and pulegone) are prepared by formylation of the ketones with ethyl formate followed by coupling with a halo butenolide. Both enantiomers of the analog derived from 1-tetralone have been prepared by employing a homochiral synthon for the coupling reaction. For three other strigolactone analogs the antipodes have been obtained by chromatography on a chiral column. All analogs have an appreciable germinating activity towards seeds of Striga hermomonthica and Orobanche crenata and O. cernua. Stereoisomers having the same configuration at the D-ring as in naturally occurring strigol have a higher stimulatory effect than the corresponding antipodes. The analogs obtained from 1-indanone and 1-tetralone have an activity comparable with that of the well known stimulant GR 24. Analogs derived from 2-phenyl-cylohexanone, carvone and pulegone also have a good germinating response. The results show that the working model for designing new bioactive strigolactones is applicable.

Aromatic A-ring analogues of orobanchol, new germination stimulants for seeds of parasitic weeds.

Strigolactones are signaling compounds in plants of increasing importance. In this paper the focus is on their activity as germinating agents for seeds of parasitic weeds. The syntheses of aromatic A-ring analogues of the germination stimulant orobanchol have been described. Starting substrate is the ABC unit of the stimulant GR24. Oxidation at the C-4 position gives a 4-oxo derivative which on subsequent reduction produces two C-4 epimeric alcohols, syn and anti in a ratio of 82 : 3. For practical access of the C-4 anti alcohol, the predominant syn epimer is inverted by a Mitsunobu procedure. The anti C-4 alcohol is then coupled with the D-ring in a one-pot two-step process involving a formylation and a reaction with bromobutenolide to give a mixture of the diastereomeric aromatic A-ring analogues of orobanchol. In contrast, the syn C-4 alcohol cannot be coupled directly with the D-ring. Protection of the C-4 syn OH is a prequisite. The best protecting function is the SEM group as deprotection after coupling with the D-ring can then readily be achieved. The structures of these new analogues have been ascertained by X-ray analyses. Both diastereomers of the C-4 syn as well as the C-4 anti orobanchol analogues have been tested as germination agents of seeds of Striga hermonthica and Orobanche ramosa. In addition, the acetates of both epimeric C-4 alcohols have been prepared and tested. Both diastereomers of the 4-oxo derivative have been prepared and bioassayed as well. The bioassays reveal that the diastereomers having the natural relative configuration are most active. The data also suggest that hydrogen bonding is not an important factor in the binding of the stimulant molecules in the receptor.

Structure and function of natural and synthetic signalling molecules in parasitic weed germination.

The structures of naturally occurring germination stimulants for seeds of the parasitic weeds Striga spp. and Orobanche spp. are described. The bioactiphore in this strigolactone family of stimulants is deduced from a structure-activity relationship and shown to reside in the CD part of the stimulant molecule. A molecular mechanism for the initial stages of seed germination is proposed. The influence of stereochemistry on the stimulant activity is significant. Combining this molecular information leads to a model for the design of synthetic strigolactones. Nijmegen-1 is a typical example of a highly active, newly designed synthetic stimulant. The occurrence of natural stimulants not belonging to the strigolactone family, such as cotylenin and parthenolide, is briefly described. The biosynthesis of natural strigolactones from beta-carotene is analysed in terms of isolated and predicted stimulants. This scheme will be helpful in the search for new strigolactones from root exudates. Protein fishing experiments to isolate and characterise the receptor protein using biotin-labelled GR 24 are described. A receptor protein of 60 kD was identified by this method. Nijmegen-1 has been tested as a suicidal germination agent in field trials on tobacco infested by Orobanche ramosa L. The preliminary results are highly rewarding. Finally, some future challenges in synthesis are described. These include synthesising new natural and synthetic stimulants and establishing the molecular connection between strigolactones as germination stimulants, as the branching factor for arbuscular mycorrhizal fungi and as an inhibitor of shoot branching.