Enhanced Herbicide Metabolism and Target Site Mutation Enabled the Multiple Resistance to Cyhalofop-butyl, Florpyrauxifen-benzyl, and Penoxsulam in Echinochloa crus-galli.

This study investigated the multiple herbicide resistance (MHR) mechanism of one Echinochloa crus-galli population that was resistant to florpyrauxifen-benzyl (FPB), cyhalofop-butyl (CHB), and penoxsulam (PEX). This population carried an Ala-122-Asn mutation in the acetolactate synthase (ALS) gene but no mutation in acetyl-CoA carboxylase (ACCase) and transport inhibitor response1 (TIR1) genes. The metabolism rate of PEX was 2-fold higher, and the production of florpyrauxifen-acid and cyhalofop-acid was lower in the resistant population. Malathion and 4-chloro-7-nitrobenzoxadiazole (NBD-Cl) could reverse the resistance, suggesting that cytochrome P450 (CYP450) and glutathione S-transferase (GST) contribute to the enhanced metabolism. According to RNA-seq and qRT-PCR validation, two CYP450 genes (CYP71C42 and CYP71D55), one GST gene (GSTT2), two glycosyltransferase genes (rhamnosyltransferase 1 and IAAGLU), and two ABC transporter genes (ABCG1 and ABCG25) were induced by CHB, FPB, and PEX in the resistant population. This study revealed that the target mutant and enhanced metabolism were involved in the MHR mechanism in E. crus-galli.

Enhanced metabolic resistance mechanism endows resistance to metamifop in Echinochloa crus-galli (L.) P. Beauv.

Barnyardgrass (Echinochloa crus-galli (L.) P. Beauv.), one of the worst weeds in paddy fields in China, has been frequently reported evolving resistance to acetyl-CoA carboxylase (ACCase) inhibiting herbicides. However, in the previous research, more attention was paid to target-site resistance (TSR) mechanisms, the non-target-site resistance (NTSR) mechanisms have not been well-established. In this study, the potential mechanism of resistance in a metamifop-resistant E. crus-galli collected from Kunshan city, Jiangsu Province, China was investigated. Dose-response assays showed that the phenotypic resistant population (JS-R) has evolved 4.3-fold resistance to metamifop compared with the phenotypic susceptible population (YN-S). The ACCase CT gene sequencing and relative ACCase gene expression levels studies showed that no mutations were detected in the ACCase CT gene in both YN-S and JS-R, and there was no significant difference in the relative ACCase gene expression between YN-S and JS-R. After the pre-processing of glutathione-S-transferase (GSTs) inhibitor NBD-Cl, the resistance level of JS-R to metamifop was reversed 18.73%. Furthermore, the GSTs activity of JS-R plants was significantly enhanced compared to that of YN-S plants. UPLC-MS/MS revealed that JS-R plants had faster metabolic rates to metamifop than YN-S plants. Meanwhile, the JS-R popultion exhibited resistant to cyhalofop-butyl and penoxsulam. In summary, this study presented a novel discovery regarding the global emergence of metabolic resistance to metamifop in E. crus-galli. The low-level resistance observed in the JS-R population was not found to be related to TSR but rather appeared to be primarily associated with the overexpression of genes in the GSTs metabolic enzyme superfamily.

Target-Site and Metabolic Resistance Mechanisms to Penoxsulam in Late Watergrass (Echinochloa phyllopogon) in China.

Echinochloa phyllopogon, a malignant weed in Northeast China's paddy fields, is currently presenting escalating resistance concerns. Our study centered on the HJHL-715 E. phyllopogon population, which showed heightened resistance to penoxsulam, through a whole-plant bioassay. Pretreatment with a P450 inhibitor malathion significantly increased penoxsulam sensitivity in resistant plants. In order to determine the resistance mechanism of the resistant population, we purified the resistant population from individual plants and isolated target-site resistance (TSR) and nontarget-site resistance (NTSR) materials. Pro-197-Thr and Trp-574-Leu mutations in acetolactate synthase (ALS) 1 and ALS2 of the resistant population drove reduced sensitivity of penoxsulam to the target-site ALS, the primary resistance mechanisms. To fully understand the NTSR mechanism, NTSR materials were investigated by using RNA-sequencing (RNA-seq) combined with a reference genome. High-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) analysis further supported the enhanced penoxsulam metabolism in NTSR materials. Gene expression data and quantitative reverse transcription polymerase chain reaction (qRT-PCR) validation confirmed 29 overexpressed genes under penoxsulam treatment, with 16 genes concurrently upregulated with quinclorac and metamifop treatment. Overall, our study confirmed coexisting TSR and NTSR mechanisms in E. phyllopogon's resistance to ALS inhibitors.

Defensive Molecules Momilactones A and B: Function, Biosynthesis, Induction and Occurrence.

Labdane-related diterpenoids, momilactones A and B were isolated and identified in rice husks in 1973 and later found in rice leaves, straws, roots, root exudate, other several Poaceae species and the moss species Calohypnum plumiforme. The functions of momilactones in rice are well documented. Momilactones in rice plants suppressed the growth of fungal pathogens, indicating the defense function against pathogen attacks. Rice plants also inhibited the growth of adjacent competitive plants through the root secretion of momilactones into their rhizosphere due to the potent growth-inhibitory activity of momilactones, indicating a function in allelopathy. Momilactone-deficient mutants of rice lost their tolerance to pathogens and allelopathic activity, which verifies the involvement of momilactones in both functions. Momilactones also showed pharmacological functions such as anti-leukemia and anti-diabetic activities. Momilactones are synthesized from geranylgeranyl diphosphate through cyclization steps, and the biosynthetic gene cluster is located on chromosome 4 of the rice genome. Pathogen attacks, biotic elicitors such as chitosan and cantharidin, and abiotic elicitors such as UV irradiation and CuCl2 elevated momilactone production through jasmonic acid-dependent and independent signaling pathways. Rice allelopathy was also elevated by jasmonic acid, UV irradiation and nutrient deficiency due to nutrient competition with neighboring plants with the increased production and secretion of momilactones. Rice allelopathic activity and the secretion of momilactones into the rice rhizosphere were also induced by either nearby Echinochloa crus-galli plants or their root exudates. Certain compounds from Echinochloa crus-galli may stimulate the production and secretion of momilactones. This article focuses on the functions, biosynthesis and induction of momilactones and their occurrence in plant species.

Lateral transfers lead to the birth of momilactone biosynthetic gene clusters in grass.

Momilactone A, an important plant labdane-related diterpenoid, functions as a phytoalexin against pathogens and an allelochemical against neighboring plants. The genes involved in the biosynthesis of momilactone A are found in clusters, i.e., momilactone A biosynthetic gene clusters (MABGCs), in the rice and barnyardgrass genomes. In addition, we know little about the origin and evolution of MABGCs. Here, we integrated results from comprehensive phylogeny and comparative genomic analyses of the core genes of MABGC-like clusters and MABGCs in 40 monocot plant genomes, providing convincing evidence for the birth and evolution of MABGCs in grass species. The MABGCs found in the PACMAD clade of the core grass lineage (including Panicoideae and Chloridoideae) originated from a MABGC-like cluster in Triticeae (BOP clade) via lateral gene transfer (LGT) and followed by recruitment of MAS1/2 and CYP76L1 genes. The MABGCs in Oryzoideae originated from PACMAD through another LGT event and lost CYP76L1 afterwards. The Oryza MABGC and another Oryza diterpenoid cluster c2BGC are two distinct clusters, with the latter originating from gene duplication and relocation within Oryzoideae. Further comparison of the expression patterns of the MABGC genes between rice and barnyardgrass in response to pathogen infection and allelopathy provides novel insights into the functional innovation of MABGCs in plants. Our results demonstrate LGT-mediated origination of MABGCs in grass and shed lights into the evolutionary innovation and optimization of plant biosynthetic pathways.

Up-regulation of bZIP88 transcription factor is involved in resistance to three different herbicides in both Echinochloa crus-galli and E. glabrescens.

The resistance of weeds to herbicides poses a major threat to agricultural production, and non-target-site resistance (NTSR) is often a serious problem as its mechanisms can in some cases confer resistance to herbicides with different modes of action. In this study, we hypothesized that bZIP transcription factors (TFs), which regulate abiotic stress responses in many plants, play a regulatory role in NTSR. Whole-plant assays indicated that the wild grasses Echinochloa crus-galli and E. glabrescens are resistant to the herbicides penoxsulam, cyhalofop-butyl, and quintrione. Transcriptome sequencing then identified 101 and 49 bZIP TFs with differential expression following penoxsulam treatment in E. crus-galli and E. glabrescens, respectively. Twelve of these genes had >60% homology with rice genes. The expression of bZIP88 was considerably up-regulated 6 h after treatment with the three different herbicides, and it was similar between resistant and susceptible populations; however, the relative expression levels before herbicide treatment and 24 h after were the same. We used rice (Oryza sativa ssp. japonica cv Nipponbare) as a model system for functional validation and found that CRISPR-Cas9-knockout of the rice bZIP88 ortholog increased the sensitivity to herbicide, whereas overexpression reduced it. The OsbZIP88 protein was localized to the nucleus. Using ChIP coupled with high-throughput sequencing, OsbZIP88 was found to form a network regulatory center with other TFs such as bZIP20/52/59 to regulate OsKS1, OsCOE1, and OsIM1, which are related to auxin, abscisic acid, brassinosteroids, and gibberellic acid. Based on these results, we have established a database of bZIP TFs corresponding to herbicide stress, and resolved the mechanisms of the positive regulation of herbicide resistance by bZIP88, thereby providing new insights for NTSR.

Structural analysis and engineering of aldo-keto reductase from glyphosate-resistant Echinochloa colona.

Glyphosate is a dominant organophosphate herbicide that inhibits 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) of the shikimate pathway. Glyphosate is extensively applied since manufactured, which has led to the emergence of various glyphosate-resistant crops and weeds. However, the molecular mechanism of many glyphosate-resistance machineries remains unclear. Recently, the upregulated expression of two homologous aldo-keto reductases (AKRs), designated as AKR4C16 and AKR4C17, were found to contribute to the glyphosate resistance in Echinochloa colona. This represents the first naturally evolved glyphosate-degrading machinery reported in plants. Here, we report the three-dimensional structure of these two AKR enzymes in complex with cofactor by performing X-ray crystallography. Furthermore, the binding-mode of glyphosate were elucidated in a ternary complex of AKR4C17. Based on the structural information and the previous study, we proposed a possible mechanism of action of AKR-mediated glyphosate degradation. In addition, a variant F291D of AKR4C17 that was constructed based on structure-based engineering showed a 70% increase in glyphosate degradation. In conclusion, these results demonstrate the structural features and glyphosate-binding mode of AKR4C17, which increases our understanding of the enzymatic mechanism of glyphosate bio-degradation and provides an important basis for the designation of AKR-based glyphosate-resistance for further applications.

Cross-resistance of barnyardgrass [Echinochloa crus-galli (L.) P. Beauv.] to aryloxyphenoxypropionate herbicides.

Managing emerged weeds that have evolved resistance to acetyl CoA carboxylase (ACCase)-inhibiting herbicides is a challenging task. A dose-response experiment was conducted on barnyardgrass biotypes resistant (R) and susceptible (S) to three aryloxyphenoxypropionate herbicides cyhalofop-butyl (CyB), fenoxaprop-ethyl (FeE), and quizalofop-ethyl (QuE) along with investigations into the potential resistance mechanism of these biotypes. The tested R barnyardgrass biotypes had strong resistance to CyB and FeE (resistant/susceptible ratio: 7.9-14.4) but weak resistance to QuE (resistant/susceptible ratio: 2.4-3.1). Absorption, translocation, and total metabolism of CyB and QuE were not associated with differences among S and R barnyardgrass biotypes. However, differences between S and R barnyardgrass were observed in production of active acid forms of each herbicide (cyhalofop-acid and quizalofop-acid). Production of cyhalofop-acid was >1.6-fold less in R barnyardgrass (3-8%) for 24 h after herbicide application than in the S barnyardgrass (8-16%). Meanwhile, production of quizalofop-acid was less in R barnyardgrass (< 14%) throughout the study period than in the S barnyardgrass (< 22%). Sequencing results of ACCase gene showed no difference between S and R barnyardgrass. Overall results show that a non-target-site resistance mechanism altering metabolism of CyB and QuE likely contributes to resistance of the barnyardgrass biotypes to these herbicides.

Study on the selectivity and phloem mobility of Fenoxaprop-P amino acid ester conjugates on rice and barnyard grass.

To improve the selectivity of the fenoxaprop herbicide to rice and barnyard grass, a series of fenoxaprop-P-ethyl-amino acid ester conjugates were designed and synthesized, and tested for biological activity as well as their phloem mobility. The bioassay results indicated that the target compounds possessed better activity against barnyard grass (Echinochloa crusgalli) than rape (Brassica campestris L.) at the concentration of 0.5 mmol/L. Compounds 3h and 3i, showed more than 70% control efficiency against barnyard grass, while less than 30% for rape. The compounds showed less impact on rice after spray treatment than in the germination test. Compounds 3i, 3j, and 3k showed excellently herbicidal activities against barnyard grass and low phytotoxicity to rice. Compound 3k showed 6.1% phytotoxicity to rice at a spray concentration of 0.25 mmol/L, better than fenoxaprop-P-ethyl (61.6%) at the same concentration. The selectivity results of the target compounds revealed that most of compounds obviously reduced phytotoxicity to rice while retaining herbicidal activity of barnyard grass. The herbicidal activity of compound 3d compared to FPE was increased by 50%, while its safety on rice was also increased by 50%. The concentration of the compounds in barnyard grass roots was higher than in rice roots, showing greater phloem mobility. In particular, the concentration of compound 3d on barnyard grass exhibited 142.72 mg/kg which was 3 times as much as Fenoxaprop, while its concentration on rice exhibited 3.65 mg/kg, the results revealed that the difference of phloem mobility might be the important reason for causing the selectivity.

High Resistance to Quinclorac in Multiple-Resistant Echinochloa colona Associated with Elevated Stress Tolerance Gene Expression and Enriched Xenobiotic Detoxification Pathway.

Echinochloa colona and other species in this genus are a threat to global rice production and food security. Quinclorac, an auxin mimic, is a common herbicide for grass weed control in rice, and Echinochloa spp. have evolved resistance to it. The complete mode of quinclorac action and subsequent evolution of resistance is not fully understood. We analyzed the de novo transcriptome of multiple-herbicide-resistant (ECO-R) and herbicide-susceptible genotypes in response to quinclorac. Several biological processes were constitutively upregulated in ECO-R, including carbon metabolism, photosynthesis, and ureide metabolism, indicating improved metabolic efficiency. The transcriptional change in ECO-R following quinclorac treatment indicates an efficient response, with upregulation of trehalose biosynthesis, which is also known for abiotic stress mitigation. Detoxification-related genes were induced in ECO-R, mainly the UDP-glycosyltransferase (UGT) family, most likely enhancing quinclorac metabolism. The transcriptome data also revealed that many antioxidant defense elements were uniquely elevated in ECO-R to protect against the auxin-mediated oxidative stress. We propose that upon quinclorac treatment, ECO-R detoxifies quinclorac utilizing UGT genes, which modify quinclorac using the sufficient supply of UDP-glucose from the elevated trehalose pathway. Thus, we present the first report of upregulation of trehalose synthesis and its association with the herbicide detoxification pathway as an adaptive mechanism to herbicide stress in Echinochloa, resulting in high resistance.

CYP81A68 confers metabolic resistance to ALS and ACCase-inhibiting herbicides and its epigenetic regulation in Echinochloa crus-galli.

Long-term and excessive herbicide use has led to some environmental concerns and especially, herbicide resistance evolution in weeds. Here, we confirmed acetolactate synthase (ALS) inhibiting herbicide penoxsulam resistance and cross resistance to acetyl-coenzyme carboxylase (ACCase) inhibiting herbicides (cyhalofop-butyl and metamifop) in a global weed Echinochloa crus-galli population resistant to these herbicides (R). Penoxsulam metabolism study indicated that degradation rate was significantly higher in R than susceptible E. crus-galli population (S). RNA-sequencing revealed that a cytochrome P450 (P450) gene, CYP81A68, expressed higher in R versus S. Rice seedlings overexpressing this CYP81A68 gene are resistant to penoxsulam, cyhalofop-butyl and metamifop, and penoxsulam resistance is due to enhanced metabolism via O-demethylation. Deletion analysis of the CYP81A68 gene promoter identified an efficient region, in which differential methylation of CpG islands occurred between R and S. Collectively, these results demonstrate that upregulation of E. crus-galli CYP81A68 gene endows generalist metabolic resistance to commonly used ALS- and ACCase-inhibiting herbicides in rice fields and epigenetic regulation may play a role in the resistance evolution. This research could contribute to strategies reducing herbicide environmental impacts by judicious selection of alternative herbicide and non-chemical control tactics.

Unravelling Phytotoxicity and Mode of Action of Tripyrasulfone, a Novel Herbicide.

Tripyrasulfone is a novel herbicide post-emergence applied in paddy fields. In this study, tripyrasulfone phytotoxicity and its mode of action were investigated. Within 3-7 days after treatment (DAT), tripyrasulfone caused strong bleaching symptoms on newly developed leaves of Echinochloa crus-galli followed by necrosis prior to death within 14 DAT. By investigating pigment composition, photosynthetic activity and energy dissipation of E. crus-galli treated with tripyrasulfone, the accumulation of phytoene and significant decreases in total carotenoids were observed; the photosystem II complex (PSII) reaction center and PSII-PSI electron transport chain were damaged; and the non-photochemical energy quenching and reactive oxygen species were significantly increased. Based on the reversion of bleaching symptoms in treated Spirodela polyrrhiza by the addition of homogentisic acid, it was hypothesized that tripyrasulfone blocks the biosynthesis of HGA, possibly by the inhibition of 4-hydroxyphenylpyruvate dioxygenase (HPPD). However, based on its chemical structure, tripyrasulfone may tend to be hydrolyzed in plants. Indeed, the hydrolyzed tripyrasulfone (HDT) inhibited the activity of HPPD from Arabidopsis thaliana produced by Escherichia coli, which was approximately 6 times less effective than mesotrione. Molecular docking showed that the HDT formed a stable bidentate interaction with the active center Fe2+ chelation of A. thaliana HPPD.

Enantioselective and Synergistic Herbicidal Activities of Common Amino Acids Against Amaranthus tricolor and Echinochloa crus-galli.

Amino acids have a wide range of biological activities, which usually rely on the stereoisomer presented. In this study, glycine and 21 common α-amino acids were investigated for their herbicidal property against Chinese amaranth (Amaranthus tricolor L.) and barnyard grass (Echinochloa crus-galli (L.) Beauv.). Both d- and l-isomers, as well as a racemic mixture, were tested and found that most compounds barely inhibited germination but moderately suppressed seedling growth. Various ratios of d:l-mixture were studied and synergy between enantiomers was found. For Chinese amaranth, the most toxic d:l-mixtures were at 3:7 (for glutamine), 8:2 (for methionine), and 5:5 (for tryptophan). For barnyard grass, rac-glutamine was more toxic than the pure forms; however, d-tryptophan exhibited greater activity than racemate and l-isomer, indicating the sign of enantioselective toxicity. The mode of action was unclear, but d-tryptophan caused bleaching of leaves, indicating pigment synthesis of the grass was inhibited. The results highlighted the enantioselective and synergistic toxicity of some amino acids, which relied upon plant species, chemical structures, and concentrations. Overall, our finding clarifies the effect of stereoisomers, and provides a chemical clue of amino acid herbicides, which may be useful in the development of herbicides from natural substances.

An ABCC-type transporter endowing glyphosate resistance in plants.

Glyphosate is the most widely used herbicide in world agriculture and for general vegetation control in a wide range of situations. Global and often intensive glyphosate selection of very large weedy plant populations has resulted in widespread glyphosate resistance evolution in populations of many weed species. Here, working with a glyphosate-resistant (GR) Echinochloa colona population that evolved in a Western Australia agricultural field, we identified an ATP-binding cassette (ABC) transporter (EcABCC8) that is consistently up-regulated in GR plants. When expressed in transgenic rice, this EcABCC8 transporter endowed glyphosate resistance. Equally, rice, maize, and soybean overexpressing the EcABCC8 ortholog genes were made resistant to glyphosate. Conversely, CRISPR/Cas9-mediated knockout of the EcABCC8 ortholog gene OsABCC8 increased rice susceptibility to glyphosate. Subcellular localization analysis and quantification of glyphosate cellular levels in treated ABCC8 transgenic rice plants and isolated leaf protoplasts as well as structural modeling support that EcABCC8 is likely a plasma membrane-localized transporter extruding cytoplasmic glyphosate to the apoplast, lowering the cellular glyphosate level. This is a report of a membrane transporter effluxing glyphosate in a GR plant species, and its function is likely conserved in crop plant species.

Synthesis and Evaluation of Halogenated 5-(2-Hydroxyphenyl)pyrazoles as Pseudilin Analogues Targeting the Enzyme IspD in the Methylerythritol Phosphate Pathway.

This work reports halogenated 5-(2-hydroxyphenyl)pyrazoles as pseudilin analogues with the potential to target the enzyme IspD in the methylerythritol phosphate (MEP) pathway. Such analogues were designed using the bioisosteric replacement of the pseudilin core structure and synthesized via an efficient three-step route. With AtIspD-based screening and pre- and post-emergence herbicidal tests, these compounds were demonstrated to have considerable activities against AtIspD, with IC50 up to 3.27 µM, and against model plants rape and barnyard grass, with moderate to excellent activities. At a rate of 150 g/ha in the greenhouse test, three compounds exhibited higher or comparable herbicidal activities than pseudilin. Molecular docking of representative compounds into the allosteric site of AtIspD revealed a binding mode similar to that of pseudilin. The established bioisosterism and synthesis method in this work may serve as an important tool for the development of new herbicides and antimicrobials targeting IspD in the MEP pathway.

Ethylene Biosynthesis Inhibition Combined with Cyanide Degradation Confer Resistance to Quinclorac in Echinochloa crus-galli var. mitis.

Echinochloa crus-galli var. mitis has rarely been reported for herbicide resistance, and no case of quinclorac resistance has been reported so far. Synthetic auxin-type herbicide quinclorac is used extensively to control rice weeds worldwide. A long history of using quinclorac in Chinese rice fields escalated the resistance in E. crus-galli var. mitis against this herbicide. Bioassays in Petri plates and pots exhibited four biotypes that evolved into resistance to quinclorac ranking as JS01-R > AH01-R > JS02-R > JX01-R from three provinces of China. Ethylene production in these biotypes was negatively correlated with resistance level and positively correlated with growth inhibition. Determination of the related ethylene response pathway exhibited resistance in biotypes that recorded a decline in 1-aminocyclopropane-1-carboxylic acid (ACC) content, ACC synthase oxidase activities, and less inducible ACS and ACO genes expressions than the susceptible biotype, suggesting that there was a positive correlation between quinclorac resistance and ethylene biosynthesis inhibition. Cyanides produced during the ethylene biosynthesis pathway mainly degraded by the activity of ß-cyanoalanine synthase (ß-CAS). Resistant biotypes exhibited higher ß-CAS activity than the susceptible ones. Nucleotide changes were found in the EcCAS gene of resistant biotypes as compared to sensitive ones that caused three amino acid substitutions (Asn-105-Lys, Gln-195-Glu, and Gly-298-Val), resulting in alteration of enzyme structure, increased binding residues in the active site with its cofactor, and decreased binding free energy; hence, its activity was higher in resistant biotypes. Moreover, these mutations increased the structural stability of the enzyme. In view of the positive correlation between ethylene biosynthesis inhibition and cyanide degradation with resistance level, it is concluded that the alteration in ethylene response pathway or at least variation in ACC synthase and ACC oxidase enzyme activities-due to less relative expression of ACS and ACO genes and enhanced ß-CAS activity, as well as mutation and increased relative expression of EcCAS gene-can be considered as a probable mechanism of quinclorac resistance in E. crus-galli var. mitis.

Groups of multi-cellular passage cells in the root exodermis of Echinochloa crus-galli varieties lack not only suberin lamellae but also lignin deposits.

Passage cells are frequently found in the exodermis and the endodermis of the roots. Because passage cells lack an apoplastic diffusion barrier, they are thought to provide pathways for the transport of nutrients and the entrance of endomycorrhizal fungi. Exodermal passage cells possess Casparian strips but not suberin lamellae. So far, exodermal passage cells have not been associated with a particular internal structure. In some wetland plants, the outer part of the root (i.e., epidermis, exodermis, and sclerenchyma) of emerging lateral root primordia has an oxygen leaky zone called a window. The exodermis at the window site also lacks suberin lamellae, but it remains unclear whether the exodermis at the window site also lacks Casparian strips. Here, we report that several of the exodermal cells in the window of Echinochloa crus-galli grown under aerated or deoxygenated stagnant agar nutrient solution also lack lignin, which is a major constituent of Casparian strips. The sclerenchyma cells that form part of the window also lacked lignin deposits. Sites at which lateral root primordia developed were highly permeable to an apoplastic tracer (periodic acid). These observations indicate that windows consist of a novel type of passage cell at the exodermis that lacks lignin as well as suberin lamellae.

The response of glyphosate-resistant and glyphosate-susceptible biotypes of Echinochloa colona to carbon dioxide, soil moisture and glyphosate.

Physiological and growth responses of two Australian Echinochloa colona biotypes (glyphosate-resistant and susceptible, produced from a single population) to different concentrations of carbon dioxide (CO2) (ambient ~450 ppm and elevated ~750 ppm) and soil moisture (well-watered and water-stressed) were analyzed. Elevated CO2 and well-watered conditions resulted in E. colona plants with greater biomass, height and numbers of tillers and leaves in both biotypes; however, no significant response was observed for seed production or the amount of photosynthesis pigments with increasing CO2 at both soil moisture levels. In addition, water availability was more influential for growth than CO2 concentration. The mean shoot biomass of the susceptible biotype under elevated CO2 and well-watered conditions was significantly greater than the resistant biotype. Although the susceptible biotype showed more vegetative and reproductive growth than the resistant biotype, no significant difference was observed for seed production between the biotypes in the water-stressed condition. In a second experiment, different doses of glyphosate (0, 180, 360, 720 and 1440 g a.e ha-1) were applied to both biotypes grown at two soil moisture levels (well-watered and water-stressed). In the water-stressed condition, glyphosate efficacy was decreased in both biotypes. The resistant biotype in the well-watered condition had only 19% survival at 1440 g ha-1 glyphosate (double the recommended rate), but this value increased in the water-stressed condition by 62%. Our study suggests that future climate change can affect the physiological and growth processes of weeds and their responses to herbicides. Knowledge of their adapting behaviors will be critical to weed management strategies.

Gene Modules Co-regulated with Biosynthetic Gene Clusters for Allelopathy between Rice and Barnyardgrass.

Allelopathy is a central process in crop-weed interactions and is mediated by the release of allelochemicals that result in adverse growth effects on one or the other plant in the interaction. The genomic mechanism for the biosynthesis of many critical allelochemicals is unknown but may involve the clustering of non-homologous biosynthetic genes involved in their formation and regulatory gene modules involved in controlling the coordinated expression within these gene clusters. In this study, we used the transcriptomes from mono- or co-cultured rice and barnyardgrass to investigate the nature of the gene clusters and their regulatory gene modules involved in the allelopathic interactions of these two plants. In addition to the already known biosynthetic gene clusters in barnyardgrass we identified three potential new clusters including one for quercetin biosynthesis and potentially involved in allelopathic interaction with rice. Based on the construction of gene networks, we identified one gene regulatory module containing hub transcription factors, significantly positively co-regulated with both the momilactone A and phytocassane clusters in rice. In barnyardgrass, gene modules and hub genes co-expressed with the gene clusters responsible for 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA) biosynthesis were also identified. In addition, we found three genes in barnyardgrass encoding indole-3-glycerolphosphate synthase that regulate the expression of the DIMBOA cluster. Our findings offer new insights into the regulatory mechanisms of biosynthetic gene clusters involved in allelopathic interactions between rice and barnyardgrass, and have potential implications in controlling weeds for crop protection.

Transcriptome profiling to identify cytochrome P450 genes involved in penoxsulam resistance in Echinochloa glabrescens.

Cytochrome P450s (P450s) confer resistance against herbicides, and this is increasingly becoming a concern for weed control. As a widespread Gramineae weed in paddy fields, Echinocloa glabrescens has become resistant to the acetolactate synthase (ALS)-inhibiting triazolopyrimidine herbicide penoxsulam. In this study, we found that the GR50 of the resistant population (SHQP-R) decreased substantially from 25.6 to 5.0 and 6.2 g a.i. ha-1 after treatment with the P450 inhibitors piperonyl butoxide (PBO) and malathion, respectively. However, P450 inhibitors almost had no effects on the susceptibility of the sensitive population (JYJD-S) to penoxsulam. To investigate the mechanisms of metabolic resistance, transcriptome sequencing analysis was performed to find candidate genes that may confer resistance to penoxsulam in E. glabrescens. A total of 233 P450 differentially expressed genes (DEGs) were identified by transcriptome sequencing. We found that the metabolic process and metabolic pathways were the most highly enriched in DEGs. Further, twenty-seven candidate P450 DEGs were selected for qPCR validation analyses. After penoxsulam treatment, the relative expression levels were significantly higher in SHQP-R than in JYJD-S. Among these, the relative expression of twenty-three P450 DEGs (eighteen from the CYP72A-71C-74A-96A-734A subfamily; five from CYP81E1-94C1-94B3-714C1-714C2) were upregulated and four P450 DEGs (from CYP724B1-711A1-707A7-97B2) were downregulated. Changes in the expression of these candidate P450 genes in E. glabrescens were in response to penoxsulam, which provides preliminary evidence for the role of P450s in herbicide metabolism in E. glabrescens. However, further functional studies on metabolic resistance to penoxsulam in a resistant E. glabrescens population are required.