Chromosome-scale pearl millet genomes reveal CLAMT1b as key determinant of strigolactone pattern and Striga susceptibility.

The yield of pearl millet, a resilient cereal crop crucial for African food security, is severely impacted by the root parasitic weed Striga hermonthica, which requires host-released hormones, called strigolactones (SLs), for seed germination. Herein, we identify four SLs present in the Striga-susceptible line SOSAT-C88-P10 (P10) but absent in the resistant 29Aw (Aw). We generate chromosome-scale genome assemblies, including four gapless chromosomes for each line. The Striga-resistant Aw lacks a 0.7 Mb genome segment containing two putative CARLACTONOIC ACID METHYLTRANSFERASE1 (CLAMT1) genes, which may contribute to SL biosynthesis. Functional assays show that P10CLAMT1b produces the SL-biosynthesis intermediate methyl carlactonoate (MeCLA) and that MeCLA is the precursor of P10-specific SLs. Screening a diverse pearl millet panel confirms the pivotal role of the CLAMT1 section for SL diversity and Striga susceptibility. Our results reveal a reason for Striga susceptibility in pearl millet and pave the way for generating resistant lines through marker-assisted breeding or direct genetic modification.

Genetic diversity analysis of tropical and sub-tropical maize germplasm for Striga resistance and agronomic traits with SNP markers.

Striga hermonthica (Sh) and S. asiatica (Sa) are major parasitic weeds limiting cereal crop production and productivity in sub-Saharan Africa (SSA). Under severe infestation, Striga causes yield losses of up to 100%. Breeding for Striga-resistant maize varieties is the most effective and economical approach to controlling the parasite. Well-characterized and genetically differentiated maize germplasm is vital to developing inbred lines, hybrids, and synthetic varieties with Striga resistance and desirable product profiles. The objective of this study was to determine the genetic diversity of 130 tropical and sub-tropical maize inbred lines, hybrids, and open-pollinated varieties germplasm using phenotypic traits and single nucleotide polymorphism (SNP) markers to select Striga-resistant and complementary genotypes for breeding. The test genotypes were phenotyped with Sh and Sa infestations using a 13x10 alpha lattice design with two replications. Agro-morphological traits and Striga-resistance damage parameters were recorded under a controlled environment. Further, high-density Diversity Array Technology Sequencing-derived SNP markers were used to profile the test genotypes. Significant phenotypic differences (P<0.001) were detected among the assessed genotypes for the assessed traits. The SNP markers revealed mean gene diversity and polymorphic information content of 0.34 and 0.44, respectively, supporting the phenotypic variation of the test genotypes. Higher significant variation was recorded within populations (85%) than between populations using the analysis of molecular variance. The Structure analysis allocated the test genotypes into eight major clusters (K = 8) in concordance with the principal coordinate analysis (PCoA). The following genetically distant inbred lines were selected, displaying good agronomic performance and Sa and Sh resistance: CML540, TZISTR25, TZISTR1248, CLHP0303, TZISTR1174, TZSTRI113, TZDEEI50, TZSTRI115, CML539, TZISTR1015, CZL99017, CML451, CML566, CLHP0343 and CML440. Genetically diverse and complementary lines were selected among the tropical and sub-tropical maize populations that will facilitate the breeding of maize varieties with Striga resistance and market-preferred traits.

Comparative secretome analysis of Striga and Cuscuta species identifies candidate virulence factors for two evolutionarily independent parasitic plant lineages.

BACKGROUND: Many parasitic plants of the genera Striga and Cuscuta inflict huge agricultural damage worldwide. To form and maintain a connection with a host plant, parasitic plants deploy virulence factors (VFs) that interact with host biology. They possess a secretome that represents the complement of proteins secreted from cells and like other plant parasites such as fungi, bacteria or nematodes, some secreted proteins represent VFs crucial to successful host colonisation. Understanding the genome-wide complement of putative secreted proteins from parasitic plants, and their expression during host invasion, will advance understanding of virulence mechanisms used by parasitic plants to suppress/evade host immune responses and to establish and maintain a parasite-host interaction. RESULTS: We conducted a comparative analysis of the secretomes of root (Striga spp.) and shoot (Cuscuta spp.) parasitic plants, to enable prediction of candidate VFs. Using orthogroup clustering and protein domain analyses we identified gene families/functional annotations common to both Striga and Cuscuta species that were not present in their closest non-parasitic relatives (e.g. strictosidine synthase like enzymes), or specific to either the Striga or Cuscuta secretomes. For example, Striga secretomes were strongly associated with 'PAR1' protein domains. These were rare in the Cuscuta secretomes but an abundance of 'GMC oxidoreductase' domains were found, that were not present in the Striga secretomes. We then conducted transcriptional profiling of genes encoding putatively secreted proteins for the most agriculturally damaging root parasitic weed of cereals, S. hermonthica. A significant portion of the Striga-specific secretome set was differentially expressed during parasitism, which we probed further to identify genes following a 'wave-like' expression pattern peaking in the early penetration stage of infection. We identified 39 genes encoding putative VFs with functions such as cell wall modification, immune suppression, protease, kinase, or peroxidase activities, that are excellent candidates for future functional studies. CONCLUSIONS: Our study represents a comprehensive secretome analysis among parasitic plants and revealed both similarities and differences in candidate VFs between Striga and Cuscuta species. This knowledge is crucial for the development of new management strategies and delaying the evolution of virulence in parasitic weeds.

Genome-wide association analyses of agronomic traits and Striga hermonthica resistance in pearl millet.

Pearl millet (Pennisetum glaucum [L.] R. Br.) is a nutrient-dense, relatively drought-tolerant cereal crop cultivated in dry regions worldwide. The crop is under-researched, and its grain yield is low (< 0.8 tons ha-1) and stagnant in the major production regions, including Burkina Faso. The low productivity of pearl millet is mainly attributable to a lack of improved varieties, Striga hermonthica [Sh] infestation, downy mildew infection, and recurrent heat and drought stress. Developing high-yielding and Striga-resistant pearl millet varieties that satisfy the farmers' and market needs requires the identification of yield-promoting genes linked to economic traits to facilitate marker-assisted selection and gene pyramiding. The objective of this study was to undertake genome-wide association analyses of agronomic traits and Sh resistance among 150 pearl millet genotypes to identify genetic markers for marker-assisted breeding and trait introgression. The pearl millet genotypes were phenotyped in Sh hotspot fields and screen house conditions. Twenty-nine million single nucleotide polymorphisms (SNPs) initially generated from 345 pearl millet genotypes were filtered, and 256 K SNPs were selected and used in the present study. Phenotypic data were collected on days to flowering, plant height, number of tillers, panicle length, panicle weight, thousand-grain weight, grain weight, number of emerged Striga and area under the Striga number progress curve (ASNPC). Agronomic and Sh parameters were subjected to combined analysis of variance, while genome-wide association analysis was performed on phenotypic and SNPs data. Significant differences (P < 0.001) were detected among the assessed pearl millet genotypes for Sh parameters and agronomic traits. Further, there were significant genotype by Sh interaction for the number of Sh and ASNPC. Twenty-eight SNPs were significantly associated with a low number of emerged Sh located on chromosomes 1, 2, 3, 4, 6, and 7. Four SNPs were associated with days-to-50%-flowering on chromosomes 3, 5, 6, and 7, while five were associated with panicle length on chromosomes 2, 3, and 4. Seven SNPs were linked to thousand-grain weight on chromosomes 2, 3, and 6. The putative SNP markers associated with a low number of emerged Sh and agronomic traits in the assessed genotypes are valuable genomic resources for accelerated breeding and variety deployment of pearl millet with Sh resistance and farmer- and market-preferred agronomic traits.

Hydroxamic acids: New players in the multifactorial mechanisms of maize resistance to Striga hermonthica.

Striga hermonthica is the most widespread and destructive plant parasite infesting maize and other major crops in sub-Saharan Africa where it causes severe yield losses and threatens food security. Several tolerant maize lines supporting reduced S. hermonthica emergence have been deployed. However, the molecular bases of such resistance are yet poorly understood. Based on a time course comparative gene expression analysis between susceptible and resistant maize lines we have confirmed resistance mechanisms known to be activated upon plant parasite infestation and identified potential novel players worth further investigation e.g. iron homeostasis and mitochondrial respiration-related genes. Most intriguingly, we show a previously unknown strategy of maize post-attachment resistance based on DIMBOA accumulation in S. hermonthica-infested maize roots. S. hermonthica infestation triggers positive regulation of gene expression in the hydroxamic acid (HA) pathway culminating with an accumulation of benzoxazinoids (BX), known for their antifeedant, insecticidal, antimicrobial, and allelopathic activities. We demonstrate that HA root content is positively correlated with S. hermonthica resistance in the resistant parent and its progenies and in unrelated maize lines. Downregulation of HA genes causes increased susceptibility to S. hermonthica infestation in loss-of-function maize mutants. While the mechanism of BX action in parasitic plant resistance is yet to be uncovered, the potential of this discovery for developing effective control and breeding strategies is enormous.

Resolving intergenotypic Striga resistance in sorghum.

Genetic underpinnings of host-pathogen interactions in the parasitic plant Striga hermonthica, a root parasitic plant that ravages cereals in sub-Saharan Africa, are unclear. We performed a comparative transcriptome study on five genotypes of sorghum exhibiting diverse resistance responses to S. hermonthica using weighted gene co-expression network analysis (WGCNA). We found that S. hermonthica elicits both basal and effector-triggered immunity-like a bona fide pathogen. The resistance response was genotype specific. Some resistance responses followed the salicylic acid-dependent signaling pathway for systemic acquired resistance characterized by cell wall reinforcements, lignification, and callose deposition, while in others the WRKY-dependent signaling pathway was activated, leading to a hypersensitive response. In some genotypes, both modes of resistance were activated, while in others either mode dominated the resistance response. Cell wall-based resistance was common to all sorghum genotypes but strongest in IS2814, while a hypersensitive response was specific to N13, IS9830, and IS41724. WGCNA further allowed for pinpointing of S. hermonthica resistance causative genes in sorghum, including glucan synthase-like 10 gene, a pathogenesis-related thaumatin-like family gene, and a phosphoinositide phosphatase gene. Such candidate genes will form a good basis for subsequent functional validation and possibly future resistance breeding.

Genome-wide association studies of Striga resistance in extra-early maturing quality protein maize inbred lines.

Identification of genes associated with Striga resistance is invaluable for accelerating genetic gains in breeding for Striga resistance in maize. We conducted a genome-wide association study to identify genomic regions associated with grain yield and other agronomic traits under artificial Striga field infestation. One hundred and forty-one extra-early quality protein maize inbred lines were phenotyped for key agronomic traits. The inbred lines were also genotyped using 49,185 DArTseq markers from which 8,143 were retained for population structure analysis and genome wide-association study. Cluster analysis and population structure revealed the presence of 3 well-defined genetic groups. Using the mixed linear model, 22 SNP markers were identified to be significantly associated with grain yield, Striga damage at 10 weeks after planting, number of emerged Striga plants at 8 and 10 weeks after planting and ear aspect. The identified SNP markers would be useful for breeders for marker-assisted selection to accelerate the genetic enhancement of maize for Striga resistance in sub-Saharan Africa after validation.

Comparative genomics of orobanchaceous species with different parasitic lifestyles reveals the origin and stepwise evolution of plant parasitism.

Orobanchaceae is the largest family of parasitic plants, containing autotrophic and parasitic plants with all degrees of parasitism. This makes it by far the best family for studying the origin and evolution of plant parasitism. Here we provide three high-quality genomes of orobanchaceous plants, the autotrophic Lindenbergia luchunensis and the holoparasitic plants Phelipanche aegyptiaca and Orobanche cumana. Phylogenomic analysis of these three genomes together with those previously published and the transcriptomes of other orobanchaceous species created a robust phylogenetic framework for Orobanchaceae. We found that an ancient whole-genome duplication (WGD; about 73.48 million years ago), which occurred earlier than the origin of Orobanchaceae, might have contributed to the emergence of parasitism. However, no WGD events occurred in any lineage of orobanchaceous parasites except for Striga after divergence from their autotrophic common ancestor, suggesting that, in contrast with previous speculations, WGD is not associated with the emergence of holoparasitism. We detected evident convergent gene loss in all parasites within Orobanchaceae and between Orobanchaceae and dodder Cuscuta australis. The gene families in the orobanchaceous parasites showed a clear pattern of recent gains and expansions. The expanded gene families are enriched in functions related to the development of the haustorium, suggesting that recent gene family expansions may have facilitated the adaptation of orobanchaceous parasites to different hosts. This study illustrates a stepwise pattern in the evolution of parasitism in the orobanchaceous parasites and will facilitate future studies on parasitism and the control of parasitic plants in agriculture.

Genome-enabled discovery of candidate virulence loci in Striga hermonthica, a devastating parasite of African cereal crops.

Parasites have evolved proteins, virulence factors (VFs), that facilitate plant colonisation, however VFs mediating parasitic plant-host interactions are poorly understood. Striga hermonthica is an obligate, root-parasitic plant of cereal hosts in sub-Saharan Africa, causing devastating yield losses. Understanding the molecular nature and allelic variation of VFs in S. hermonthica is essential for breeding resistance and delaying the evolution of parasite virulence. We assembled the S. hermonthica genome and identified secreted proteins using in silico prediction. Pooled sequencing of parasites growing on a susceptible and a strongly resistant rice host allowed us to scan for loci where selection imposed by the resistant host had elevated the frequency of alleles contributing to successful colonisation. Thirty-eight putatively secreted VFs had very different allele frequencies with functions including host cell wall modification, protease or protease inhibitor and kinase activities. These candidate loci had significantly higher Tajima's D than the genomic background, consistent with balancing selection. Our results reveal diverse strategies used by S. hermonthica to overcome different layers of host resistance. Understanding the maintenance of variation at virulence loci by balancing selection will be critical to managing the evolution of virulence as part of a sustainable control strategy.

Optimizing use of U.S. Ex-PVP inbred lines for enhancing agronomic performance of tropical Striga resistant maize inbred lines.

BACKGROUND: Temperate maize inbred lines with expired Plant Variety Protection Act certificates (Ex-PVP) are potential sources of desirable alleles for tropical germplasm improvement. Up to now, the usefulness of the Ex-PVP inbred lines as a potential source of novel beneficial alleles for Striga hermonthica resistance breeding to enhance genetic gain in tropical maize has not been reported. RESULTS: This study was thus conducted to characterize the combining ability of 24 Ex-PVP inbred lines in crosses with two tropical Striga resistant inbred testers under Striga-infested and non-infested conditions and across three locations for 2 years. Many testcrosses between Ex-PVP inbred lines and the first tester (T1) produced competitive or significantly higher grain yields compared to the hybrid between the two resistant testers under Striga infested and non-infested conditions and across multiple test locations. Also, most of the testcrosses with positive heterosis for grain yield and negative heterosis for Striga damage and emerged Striga count involved T1 as a tester. Our study identified six Ex-PVP inbred lines with positive GCA effects for grain yield under Striga infested and non-infested conditions and across multiple test locations. Amongst these, inbred lines HB8229-1 and WIL900-1 also displayed negative GCA effects for emerged Striga count and Striga damage rating. The inbred line HB8229-1 showed positive SCA effects for grain yield with T2, whereas WIL900-1 had positive SCA effects for grain yield with T1. Over 70% of the Ex-PVP inbred lines were consistently assigned to specific heterotic groups using yield-based classifying methods (mean grain yield and SCA effects). CONCLUSIONS: These results could facilitate systematic introgression of the Ex-PVP inbred lines into the existing Striga resistant heterotic groups in IITA. The Ex-PVP inbred lines with positive GCA effects and producing high grain yields in hybrid combinations could be useful parents for enhancing Striga resistance and agronomic performance of tropical maize hybrids.

Identification of dicistro-like viruses in the transcriptome data of Striga asiatica and other plants.

Dicistroviruses (the family Dicistroviridae) are positive-sense single-stranded RNA viruses of the order Picornavirales, which is a rapidly growing viral group. They have been detected in a wide range of animals, predominantly in insects and crustaceans. In this study, we identified the genome sequences of 14 dicistro-like viruses in the transcriptome data from 12 plant species, including Striga asiatica dicistro-like virus 1 and 2 identified in the transcriptome data of Striga asiatica. Sequence comparison and phylogenetic analysis indicated that these 14 plant-associated dicistro-like viruses were novel members of the family Dicistroviridae, five of which are placed within the genera Aparavirus and Cripavirus, which mainly consist of viruses infecting animals, including insects. The other nine plant dicistro-like viruses formed clades with unclassified dicistroviruses. Our study implies that a wide range of plant species may serve as hosts for dicistroviruses or reservoirs for their transmission. Keywords: dicistrovirus; Dicistroviridae; plant; transcriptome; Striga asiatica.

Mechanisms of pre-attachment Striga resistance in sorghum through genome-wide association studies.

Witchweeds (Striga spp.) greatly limit production of Africa's most staple crops. These parasitic plants use strigolactones (SLs)-chemical germination stimulants, emitted from host's roots to germinate, and locate their hosts for invasion. This information exchange provides opportunities for controlling the parasite by either stimulating parasite seed germination without a host (suicidal germination) or by inhibiting parasite seed germination (pre-attachment resistance). We sought to determine genetic factors that underpin Striga pre-attachment resistance in sorghum using the genome wide association study (GWAS) approach. Results revealed that Striga germination was associated with genes encoding hormone signaling functions, e.g., the Novel interactor of jaz (NINJA) and, Abscisic acid-insensitive 5 (ABI5). This pointed toward abscisic acid (ABA) and gibberellic acid (GA) as probable determinants of Striga germination. To test this hypothesis, we conditioned Striga using: ABA, ABA + its inhibitor fluridone (FLU), GA or water. Unexpectedly, Striga conditioned with FLU germinated after 4 days without SL. Upon germination stimulation using sorghum root exudate or the synthetic SL GR24, we found that ABA conditioned seeds had above 20-fold reduction in germination. Conversely, FLU conditioned seeds recorded above 20-fold increase in germination. Conditioning with GA reduced Striga seed germination 1.5-fold only in the GR24 treatment. Germination assays using seeds of a related parasitic plant (Alectra vogelii) showed similar degrees of stimulation and reduction of germination by the hormones further affirming the hormonal crosstalk. Our findings have far-reaching implications in the control of some of the most noxious pathogens of crops in Africa.

Association analysis for resistance to Striga hermonthica in diverse tropical maize inbred lines.

Striga hermonthica is a widespread, destructive parasitic plant that causes substantial yield loss to maize productivity in sub-Saharan Africa. Under severe Striga infestation, yield losses can range from 60 to 100% resulting in abandonment of farmers' lands. Diverse methods have been proposed for Striga management; however, host plant resistance is considered the most effective and affordable to small-scale famers. Thus, conducting a genome-wide association study to identify quantitative trait nucleotides controlling S. hermonthica resistance and mining of relevant candidate genes will expedite the improvement of Striga resistance breeding through marker-assisted breeding. For this study, 150 diverse maize inbred lines were evaluated under Striga infested and non-infested conditions for two years and genotyped using the genotyping-by-sequencing platform. Heritability estimates of Striga damage ratings, emerged Striga plants and grain yield, hereafter referred to as Striga resistance-related traits, were high under Striga infested condition. The mixed linear model (MLM) identified thirty SNPs associated with the three Striga resistance-related traits based on the multi-locus approaches (mrMLM, FASTmrMLM, FASTmrEMMA and pLARmEB). These SNPs explained up to 14% of the total phenotypic variation. Under non-infested condition, four SNPs were associated with grain yield, and these SNPs explained up to 17% of the total phenotypic variation. Gene annotation of significant SNPs identified candidate genes (Leucine-rich repeats, putative disease resistance protein and VQ proteins) with functions related to plant growth, development, and defense mechanisms. The marker-effect prediction was able to identify alleles responsible for predicting high yield and low Striga damage rating in the breeding panel. This study provides valuable insight for marker validation and deployment for Striga resistance breeding in maize.

A chromosome-scale Gastrodia elata genome and large-scale comparative genomic analysis indicate convergent evolution by gene loss in mycoheterotrophic and parasitic plants.

Mycoheterotrophic and parasitic plants are heterotrophic and parasitize on fungi and plants, respectively, to obtain nutrients. Large-scale comparative genomics analysis has not been conducted in mycoheterotrophic or parasitic plants or between these two groups of parasites. We assembled a chromosome-level genome of the fully mycoheterotrophic plant Gastrodia elata (Orchidaceae) and performed comparative genomic analyses on the genomes of G. elata and four orchids (initial mycoheterotrophs), three parasitic plants (Cuscuta australis, Striga asiatica, and Sapria himalayana), and 36 autotrophs from various angiosperm lineages. It was found that while in the hemiparasite S. asiatica and initial mycoheterotrophic orchids, approximately 4-5% of the conserved orthogroups were lost, the fully heterotrophic G. elata and C. australis both lost approximately 10% of the conserved orthogroups, indicating that increased heterotrophy is positively associated with gene loss. Importantly, many genes that are essential for autotrophs, including those involved in photosynthesis, the circadian clock, flowering time regulation, immunity, nutrient uptake, and root and leaf development, were convergently lost in both G. elata and C. australis. The high-quality genome of G. elata will facilitate future studies on the physiology, ecology, and evolution of mycoheterotrophic plants, and our findings highlight the critical role of gene loss in the evolution of plants with heterotrophic lifestyles.

GWAS provides biological insights into mechanisms of the parasitic plant (Striga) resistance in sorghum.

BACKGROUND: Sorghum yields in sub-Saharan Africa (SSA) are greatly reduced by parasitic plants of the genus Striga (witchweed). Vast global sorghum genetic diversity collections, as well as the availability of modern sequencing technologies, can be potentially harnessed to effectively manage the parasite. RESULTS: We used laboratory assays - rhizotrons to screen a global sorghum diversity panel to identify new sources of resistance to Striga; determine mechanisms of resistance, and elucidate genetic loci underlying the resistance using genome-wide association studies (GWAS). New Striga resistant sorghum determined by the number, size and biomass of parasite attachments were identified. Resistance was by; i) mechanical barriers that blocked parasite entry, ii) elicitation of a hypersensitive reaction that interfered with parasite development, and iii) the inability of the parasite to develop vascular connections with hosts. Resistance genes underpinning the resistance corresponded with the resistance mechanisms and included pleiotropic drug resistance proteins that transport resistance molecules; xylanase inhibitors involved in cell wall fortification and hormonal regulators of resistance response, Ethylene Response Factors. CONCLUSIONS: Our findings are of fundamental importance to developing durable and broad-spectrum resistance against Striga and have far-reaching applications in many SSA countries where Striga threatens the livelihoods of millions of smallholder farmers that rely on sorghum as a food staple.

Role of substrate recognition in modulating strigolactone receptor selectivity in witchweed.

Witchweed, or Striga hermonthica, is a parasitic weed that destroys billions of dollars' worth of crops globally every year. Its germination is stimulated by strigolactones exuded by its host plants. Despite high sequence, structure, and ligand-binding site conservation across different plant species, one strigolactone receptor in witchweed, ShHTL7, uniquely exhibits a picomolar EC50 for downstream signaling. Previous biochemical and structural analyses have hypothesized that this unique ligand sensitivity can be attributed to a large binding pocket volume in ShHTL7 resulting in enhanced ability to bind substrates, but additional structural details of the substrate-binding process would help explain its role in modulating the ligand selectivity. Using long-timescale molecular dynamics simulations, we demonstrate that mutations at the entrance of the binding pocket facilitate a more direct ligand-binding pathway to ShHTL7, whereas hydrophobicity at the binding pocket entrance results in a stable "anchored" state. We also demonstrate that several residues on the D-loop of AtD14 stabilize catalytically inactive conformations. Finally, we show that strigolactone selectivity is not modulated by binding pocket volume. Our results indicate that while ligand binding is not the sole modulator of strigolactone receptor selectivity, it is a significant contributing factor. These results can be used to inform the design of selective antagonists for strigolactone receptors in witchweed.

Molecular basis for high ligand sensitivity and selectivity of strigolactone receptors in Striga.

Seeds of the root parasitic plant Striga hermonthica can sense very low concentrations of strigolactones (SLs) exuded from host roots. The S. hermonthica hyposensitive to light (ShHTL) proteins are putative SL receptors, among which ShHTL7 reportedly confers sensitivity to picomolar levels of SL when expressed in Arabidopsis thaliana. However, the molecular mechanism underlying ShHTL7 sensitivity is unknown. Here we determined the ShHTL7 crystal structure and quantified its interactions with various SLs and key interacting proteins. We established that ShHTL7 has an active-site pocket with broad-spectrum response to different SLs and moderate affinity. However, in contrast to other ShHTLs, we observed particularly high affinity of ShHTL7 for F-box protein AtMAX2. Furthermore, ShHTL7 interacted with AtMAX2 and with transcriptional regulator AtSMAX1 in response to nanomolar SL concentration. ShHTL7 mutagenesis analyses identified surface residues that contribute to its high-affinity binding to AtMAX2 and residues in the ligand binding pocket that confer broad-spectrum response to SLs with various structures. Crucially, yeast-three hybrid experiments showed that AtMAX2 confers responsiveness of the ShHTL7-AtSMAX1 interaction to picomolar levels of SL in line with the previously reported physiological sensitivity. These findings highlight the key role of SL-induced MAX2-ShHTL7-SMAX1 complex formation in determining the sensitivity to SL. Moreover, these data suggest a strategy to screen for compounds that could promote suicidal seed germination at physiologically relevant levels.

The Effect of Virulence and Resistance Mechanisms on the Interactions between Parasitic Plants and Their Hosts.

Parasitic plants have a unique heterotrophic lifestyle based on the extraction of water and nutrients from host plants. Some parasitic plant species, particularly those of the family Orobanchaceae, attack crops and cause substantial yield losses. The breeding of resistant crop varieties is an inexpensive way to control parasitic weeds, but often does not provide a long-lasting solution because the parasites rapidly evolve to overcome resistance. Understanding mechanisms underlying naturally occurring parasitic plant resistance is of great interest and could help to develop methods to control parasitic plants. In this review, we describe the virulence mechanisms of parasitic plants and resistance mechanisms in their hosts, focusing on obligate root parasites of the genera Orobanche and Striga. We noticed that the resistance (R) genes in the host genome often encode proteins with nucleotide-binding and leucine-rich repeat domains (NLR proteins), hence we proposed a mechanism by which host plants use NLR proteins to activate downstream resistance gene expression. We speculated how parasitic plants and their hosts co-evolved and discussed what drives the evolution of virulence effectors in parasitic plants by considering concepts from similar studies of plant-microbe interaction. Most previous studies have focused on the host rather than the parasite, so we also provided an updated summary of genomic resources for parasitic plants and parasitic genes for further research to test our hypotheses. Finally, we discussed new approaches such as CRISPR/Cas9-mediated genome editing and RNAi silencing that can provide deeper insight into the intriguing life cycle of parasitic plants and could potentially contribute to the development of novel strategies for controlling parasitic weeds, thereby enhancing crop productivity and food security globally.

Comparative Studies of Potential Binding Pocket Residues Reveal the Molecular Basis of ShHTL Receptors in the Perception of GR24 in Striga.

Root parasitic weeds such as Striga spp. have caused significant losses in agriculture production worldwide. The seed germination of the weeds depends on strigolactones (SLs) that target a series of HYPOSENSITIVE TO LIGHT/KARRIKIN INSENSITIVE2 in Striga hermonthica (ShHTL) proteins. In the present study, 60 ShHTL7 mutants were constructed, and the equilibrium dissociation constants for GR24 (a synthetic SL analogue, commonly used as a standard in SL germination studies) against these mutants were measured by surface plasmon resonance. Based on these data, the SL binding pocket residues were distinguished. Of them, some specific residues for ShHTL7 were found, such as T142, T190, and M219. A model showing quite well internal and external predictive abilities was established by the mutation-dependent biomacromolecular quantitative structure-activity relationship method. It provided an expanded understanding for GR24 binding to a series of ShHTL receptors and should help design broad-spectrum agrochemicals with cross interactions with several members of SL receptors.