Genomic prediction for agronomic traits in a diverse Flax (Linum usitatissimum L.) germplasm collection.

Breeding programs require exhaustive phenotyping of germplasms, which is time-demanding and expensive. Genomic prediction helps breeders harness the diversity of any collection to bypass phenotyping. Here, we examined the genomic prediction's potential for seed yield and nine agronomic traits using 26,171 single nucleotide polymorphism (SNP) markers in a set of 337 flax (Linum usitatissimum L.) germplasm, phenotyped in five environments. We evaluated 14 prediction models and several factors affecting predictive ability based on cross-validation schemes. Models yielded significant variation among predictive ability values across traits for the whole marker set. The ridge regression (RR) model covering additive gene action yielded better predictive ability for most of the traits, whereas it was higher for low heritable traits by models capturing epistatic gene action. Marker subsets based on linkage disequilibrium decay distance gave significantly higher predictive abilities to the whole marker set, but for randomly selected markers, it reached a plateau above 3000 markers. Markers having significant association with traits improved predictive abilities compared to the whole marker set when marker selection was made on the whole population instead of the training set indicating a clear overfitting. The correction for population structure did not increase predictive abilities compared to the whole collection. However, stratified sampling by picking representative genotypes from each cluster improved predictive abilities. The indirect predictive ability for a trait was proportionate to its correlation with other traits. These results will help breeders to select the best models, optimum marker set, and suitable genotype set to perform an indirect selection for quantitative traits in this diverse flax germplasm collection.

Weed-induced changes in the maize root transcriptome reveal transcription factors and physiological processes impacted early in crop-weed interactions.

A new paradigm suggests weeds primarily reduce crop yield by altering crop developmental and physiological processes long before the weeds reduce resources through competition. Multiple studies have implicated stress response pathways are activated when crops such as maize are grown in close proximity with weeds during the first 4-8 weeks of growth-the point at which weeds have their greatest impact on subsequent crop yields. To date, these studies have mostly focused on the response of above-ground plant parts and have not examined the early signal transduction processes associated with maize root response to weeds. To investigate the impact of signals from a below-ground competitor on the maize root transcriptome when most vulnerable to weed pressure, a system was designed to expose maize to only below-ground signals. Gene set enrichment analyses identified over-represented ontologies associated with oxidative stress signalling throughout the time of weed exposure, with additional ontologies associated with nitrogen use and transport and abscisic acid (ABA) signalling, and defence responses being enriched at later time points. Enrichment of promoter motifs indicated over-representation of sequences known to bind FAR-RED IMPAIRED RESPONSE 1 (FAR1), several AP2/ERF transcription factors and others. Likewise, co-expression networks were identified using Weighted-Gene Correlation Network Analysis (WGCNA) and Spatiotemporal Clustering and Inference of Omics Networks (SC-ION) algorithms. WGCNA highlighted the potential roles of several transcription factors including a MYB 3r-4, TB1, WRKY65, CONSTANS-like5, ABF3, HOMEOBOX 12, among others. These studies also highlighted the role of several specific proteins involved in ABA signalling as being important for the initiation of the early response of maize to weeds. SC-ION highlighted potential roles for NAC28, LOB37, NAC58 and GATA2 transcription factors, among many others.

Overwintering Camelina and Canola/Rapeseed Show Promise for Improving Integrated Weed Management Approaches in the Upper Midwestern U.S.

Winter oilseed cash cover crops are gaining popularity in integrated weed management programs for suppressing weeds. A study was conducted at two field sites (Fargo, North Dakota, and Morris, Minnesota) to determine the freezing tolerance and weed-suppressing traits of winter canola/rapeseed (Brassica napus L.) and winter camelina [Camelina sativa (L.) Crantz] in the Upper Midwestern USA. The top 10 freezing tolerant accessions from a phenotyped population of winter canola/rapeseed were bulked and planted at both locations along with winter camelina (cv. Joelle) as a check. To phenotype our entire winter B. napus population (621 accessions) for freezing tolerance, seeds were also bulked and planted at both locations. All B. napus and camelina were no-till seeded at Fargo and Morris at two planting dates, late August (PD1) and mid-September (PD2) 2019. Data for winter survival of oilseed crops (plants m-2) and their corresponding weed suppression (plants m-2 and dry matter m-2) were collected on two sampling dates (SD) in May and June 2020. Crop and SD were significant (p < 0.05) for crop plant density at both locations, and PD in Fargo and crop x PD interaction in Morris were significant for weed dry matter. At Morris and Fargo, PD1 produced greater winter B. napus survival (28% and 5%, respectively) and PD2 produced higher camelina survival (79% and 72%, respectively). Based on coefficient of determination (r2), ~50% of weed density was explained by camelina density, whereas ≤20% was explained by B. napus density at both locations. Camelina from PD2 suppressed weed dry matter by >90% of fallow at both locations, whereas weed dry matter in B. napus was not significantly different from fallow at either PD. Genotyping of overwintering canola/rapeseed under field conditions identified nine accessions that survived at both locations, which also had excellent freezing tolerance under controlled conditions. These accessions are good candidates for improving freezing tolerance in commercial canola cultivars.

Homozygosity mapping identified loci and candidate genes responsible for freezing tolerance in Camelina sativa.

Homozygosity mapping is an effective tool for detecting genomic regions responsible for a given trait when the phenotype is controlled by a limited number of dominant or co-dominant loci. Freezing tolerance is a major attribute in agricultural crops such as camelina. Previous studies indicated that freezing tolerance differences between a tolerant (Joelle) and susceptible (CO46) variety of camelina were controlled by a small number of dominant or co-dominant genes. We performed whole genome homozygosity mapping to identify markers and candidate genes responsible for freezing tolerance difference between these two genotypes. A total of 28 F3 RILs were sequenced to ∼30× coverage, and parental lines were sequenced to >30-40× coverage with Pacific Biosciences high fidelity technology and 60× coverage using Illumina whole genome sequencing. Overall, about 126k homozygous single nucleotide polymorphism markers were identified that differentiate both parents. Moreover, 617 markers were also homozygous in F3 families fixed for freezing tolerance/susceptibility. All these markers mapped to two contigs forming a contiguous stretch of chromosome 11. The homozygosity mapping detected 9 homozygous blocks among the selected markers and 22 candidate genes with strong similarity to regions in or near the homozygous blocks. Two such genes were differentially expressed during cold acclimation in camelina. The largest block contained a cold-regulated plant thionin and a putative rotamase cyclophilin 2 gene previously associated with freezing resistance in arabidopsis (Arabidopsis thaliana). The second largest block contains several cysteine-rich RLK genes and a cold-regulated receptor serine/threonine kinase gene. We hypothesize that one or more of these genes may be primarily responsible for freezing tolerance differences in camelina varieties.

Weed-induced crop yield loss: a new paradigm and new challenges.

Direct competition for resources is generally considered the primary mechanism for weed-induced yield loss. A re-evaluation of physiological evidence suggests weeds initially impact crop growth and development through resource-independent interference. We suggest weed perception by crops induce a shift in crop development, before resources become limited, which ultimately reduce crop yield, even if weeds are subsequently removed. We present the mechanisms by which crops perceive and respond to weeds and discuss the technologies used to identify these mechanisms. These data lead to a fundamental paradigm shift in our understanding of how weeds reduce crop yield and suggest new research directions and opportunities to manipulate or engineer crops and cropping systems to reduce weed-induced yield losses.

Genetic loci associated with freezing tolerance in a European rapeseed (Brassica napus L.) diversity panel identified by genome-wide association mapping.

Winter biotypes of rapeseed (Brassica napus L.) require a vernalization treatment to enter the reproductive phase and generally produce greater yields than spring rapeseed. To find genetic loci associated with freezing tolerance in rapeseed, we first performed genotyping-by-sequencing (GBS) on a diversity panel consisting of 222 rapeseed accessions originating primarily from Europe, which identified 69,554 high-quality single-nucleotide polymorphisms (SNPs). Model-based cluster analysis suggested that there were eight subgroups. The diversity panel was then phenotyped for freezing survival (visual damage and Fv/Fo and Fv/Fm) after 2 months of cold acclimation (5°C) and a freezing treatment (-15°C for 4 h). The genotypic and phenotypic data for each accession in the rapeseed diversity panel was then used to conduct a genome-wide association study (GWAS). GWAS results showed that 14 significant markers were mapped to seven chromosomes for the phenotypes scored. Twenty-four candidate genes located within the mapped loci were identified as previously associated with lipid, photosynthesis, flowering, ubiquitination, and cytochrome P450 in rapeseed or other plant species.

Fatty Acid Data and Crop Surveys Indicate Sources of Red Sunflower Seed Weevil (Coleoptera: Curculionidae), Populations and Suggest Strategies for Management.

The red sunflower seed weevil, Smicronyx fulvus L., is a univoltine seed-feeding pest of cultivated sunflower, Helianthus annuus L. Artificial infestations of S. fulvus onto sunflowers with traditional (<25% oleic acid), mid-oleic (55-75%), or high oleic (>80%) fatty acid profiles were used to test if fatty acids could be used as natural markers to estimate the proportion of weevils developing on oilseed sunflowers rather than wild Helianthus spp. and confection (non-oil) types. Oleic acid (%) in S. fulvus confirmed the fatty acid compositions of mature larvae and weevil adults reflected their diets, making primary (oleic or linoleic) fatty acids feasible as natural markers for this crop-insect combination. Oleic acid in wild S. fulvus populations in North Dakota suggests at least 84 and 90% of adults originated from mid-oleic or high oleic sunflower hybrids in 2017 and 2018, respectively. Surveys in 2017 (n = 156 fields) and 2019 (n = 120 fields) extended information provided by S. fulvus fatty acid data; no significant spatial patterns of S. fulvus damage were detected in samples, damage to oilseed sunflowers was greater than confection (non-oil) types, and the majority of damage occurred in ≈10% of surveyed fields. Combined, data suggest a few unmanaged or mismanaged oilseed sunflower fields are responsible for producing most S. fulvus in an area. Improved management seems possible with a combination of grower education and expanded use of non-insecticidal tactics, including cultural practices and S. fulvus-resistant hybrids.

Genome-Wide Association Studies and Transcriptome Changes during Acclimation and Deacclimation in Divergent Brassica napus Varieties.

Information concerning genes and signals regulating cold acclimation processes in plants is abundant; however, less is known about genes and signals regulating the deacclimation process. A population of primarily winter B. napus varieties was used to conduct a genome-wide association study and to compare the transcriptomes from two winter B. napus varieties showing time-dependent differences in response to cold acclimation and deacclimation treatments. These studies helped to identify loci, candidate genes, and signaling processes impacting deacclimation in B. napus. GWAS identified polymorphisms at five different loci associated with freezing tolerance following deacclimation. Local linkage decay rates near these polymorphisms identified 38 possible candidate genes. Several of these genes have been reported as differentially regulated by cold stress in arabidopsis (Arabidopsis thaliana), including a calcium-binding EF-hand family protein (encoded by BnaCnng10250D) that was also differentially expressed during deacclimation in this study. Thousands of other genes differentially expressed during the acclimation and deacclimation treatments implicated processes involving oxidative stress, photosynthesis, light-regulated diurnal responses, and growth regulation. Generally, responses observed during acclimation were reversed within one week of deacclimation. The primary differences between the two winter B. napus varieties with differential deacclimation responses involved protection from oxidative stress and the ability to maintain photosynthesis.

Genes associated with chloroplasts and hormone-signaling, and transcription factors other than CBFs are associated with differential survival after low temperature treatments of Camelina sativa biotypes.

Winter annual biotypes of Camelina sativa regularly survive after winter conditions experienced in northern regions of the U.S., whereas summer annual biotypes do not. To determine potential molecular mechanisms associated with these biotype differences in survival after low temperature treatments, we examined genetic and transcript variations in both a winter- (Joelle) and a summer- (CO46) biotype. It was determined that as few as one or two dominant genes may control differential survival after low temperature treatments. Of the 1797 genes that were differentially expressed in response to cold in both the winter and summer biotypes many COR genes were identified, indicating that the CBF regulon is functional in both. However, only 153 and 76 genes from Joelle and CO46, respectively, were either differentially expressed or not expressed at all in one biotype versus the other following cold acclimation. We hypothesize that these 229 genes play a significant role in, or are primarily responsive to, differences in survival after freezing between these two biotypes. Promoter analysis provided few clues as to the regulation or these genes; however, genes that were down-regulated specifically in the winter biotype Joelle were enriched with the sequence TGGCCCTCGCTCAC, which is over-represented among genes associated with chloroplasts in Arabidopsis. Additionally, several genes involved in auxin signaling were down-regulated specifically in Joelle. A transcription factor with strong similarity to MYB47, known to be up-regulated by salt, drought, and jasmonic acid, but not cold in Arabidopsis, was essentially off in the freezing sensitive biotype CO46, but was cold-induced in the winter biotype Joelle. Several other transcription factors genes including three with similarity to WRKY70, that may be involved in SA/JA-dependent responses, a HOMEOBOX 6 gene involved in ABA signaling, and two others (NUCLEAR FACTOR Y and CONSTANS-like 2) known to be implicated in photoperiodic flowering were also differentially expressed between the two biotypes.

Varying Weed Densities Alter the Corn Transcriptome, Highlighting a Core Set of Weed-Induced Genes and Processes with Potential for Manipulating Weed Tolerance.

CORE IDEAS: Corn increases the number of differentially expressed genes and the intensity of differential gene expression in response to increasing weed density. Genes associated with kinase signaling and transport functions are upregulated by weeds. Genes associated with protein production are downregulated by weeds. A sugar transporter (PMT5) and NUCLEOREDOXIN 1 are upregulated by weeds under diverse conditions. The phenological responses of corn (Zea mays L.) to competition with increasing densities of winter canola (Brassica napus L.) as the weedy competitor were investigated. Changes in the corn transcriptome resulting from varying weed densities were used to identify genes and processes responsive to competition under controlled conditions where light, nutrients, and water were not limited. Increasing densities of weeds resulted in decreased corn growth and development and increased the number and expression intensity of competition-responsive genes. The physiological processes identified in corn that were consistently induced by competition with weeds included protein synthesis and various transport functions. Likewise, numerous genes involved in these processes, as well as several genes implicated in phytochrome signaling and defense responses, were noted as differentially expressed. The results obtained in this study, conducted under controlled (greenhouse) conditions, were compared with a previously published study where the response of corn to competition with other species was evaluated under field conditions. Approximately one-third of the genes were differentially expressed in response to competition under both field and controlled conditions. These competition-responsive genes represent a resource for investigating the signaling processes by which corn recognizes and responds to competition. These results also highlight specific physiological processes that might be targets for mitigating the response of crops to weeds or other competitive plants under field conditions.

Weed presence altered biotic stress and light signaling in maize even when weeds were removed early in the critical weed-free period.

Weed presence early in the life cycle of maize (typically, from emergence through the 8 to 12 leaf growth stage) can reduce crop growth and yield and is known as the critical weed-free period (CWFP). Even if weeds are removed during or just after the CWFP, crop growth and yield often are not recoverable. We compared transcriptome responses of field-grown hybrid maize at V8 in two consecutive years among plants grown under weed-free and two weed-stressed conditions (weeds removed at V4 or present through V8) using RNAseq analysis techniques. Compared with weed-free plant responses, physiological differences at V8 were identified in all weed-stressed plants and were most often associated with altered photosynthetic processes, hormone signaling, nitrogen use and transport, and biotic stress responses. Even when weeds were removed at V4 and tissues sampled at V8, carbon: nitrogen supply imbalance, salicylic acid signals, and growth responses differed between the weed-stressed and weed-free plants. These underlying processes and a small number of developmentally important genes are potential targets for decreasing the maize response to weed pressure. Expression differences of several novel, long noncoding RNAs resulting from exposure of maize to weeds during the CWFP were also observed and could open new avenues for investigation into the function of these transcription units.

Expression of FLOWERING LOCUS C and a frameshift mutation of this gene on chromosome 20 differentiate a summer and winter annual biotype of Camelina sativa.

The nature of the vegetative to reproductive transition in the shoot apical meristem of Camelina sativa summer annual cultivar CO46 and winter annual cultivar Joelle was confirmed by treating seedlings with or without 8 weeks of vernalization. True to their life cycle classification, Joelle required a vernalization treatment to induce bolting and flowering, whereas CO46 did not. In this study, whole genome sequence, RNAseq, and resequencing of PCR-amplified transcripts for a key floral repressor were used to better understand factors involved in the flowering habit of summer and winter biotypes at the molecular level. Analysis of transcriptome data indicated that abundance for one of the three genes encoding the floral repressor FLOWERING LOCUS C (FLC; Csa20 g015400) was 16-fold greater in Joelle compared to CO46 prior to vernalization. Abundance of this transcript decreased only slightly in CO46 postvernalization, compared to a substantial decrease in Joelle. The results observed in the winter annual biotype Joelle are consistent with repression of FLC by vernalization. Further characterization of FLC at both the genome and transcriptome levels identified a one base deletion in the 5th exon coding for a keratin-binding domain in chromosome 20 of CO46 and Joelle. The one base deletion detected in chromosome 20 FLC is predicted to result in a frameshift that would produce a nonfunctional protein. Analysis of whole genome sequence indicated that the one base deletion in chromosome 20 FLC occurred at a greater ratio in the summer biotype CO46 (2:1) compared to the winter biotype Joelle (1:4); similar trends were also observed for RNAseq and cDNA transcripts mapping to chromosome 20 FLC of CO46 and Joelle.

Comparison of phytohormone levels and transcript profiles during seasonal dormancy transitions in underground adventitious buds of leafy spurge.

Leafy spurge (Euphorbia esula L.) is an herbaceous perennial weed that maintains its perennial growth habit through generation of underground adventitious buds (UABs) on the crown and lateral roots. These UABs undergo seasonal phases of dormancy under natural conditions, namely para-, endo-, and ecodormancy in summer, fall, and winter, respectively. These dormancy phases can also be induced in growth chambers by manipulating photoperiod and temperature. In this study, UABs induced into the three phases of dormancy under controlled conditions were used to compare changes in phytohormone and transcriptome profiles. Results indicated that relatively high levels of ABA, the ABA metabolite PA, and IAA were found in paradormant buds. When UABs transitioned from para- to endodormancy, ABA and PA levels decreased, whereas IAA levels were maintained. Additionally, transcript profiles associated with regulation of soluble sugars and ethylene activities were also increased during para- to endodormancy transition, which may play some role in maintaining endodormancy status. When crown buds transitioned from endo- to ecodormancy, the ABA metabolites PA and DPA decreased significantly along with the down-regulation of ABA biosynthesis genes, ABA2 and NCED3. IAA levels were also significantly lower in ecodormant buds than that of endodormant buds. We hypothesize that extended cold treatment may trigger physiological stress in endodormant buds, and that these stress-associated signals induced the endo- to ecodormancy transition and growth competence. The up-regulation of NAD/NADH phosphorylation and dephosphorylation pathway, and MAF3-like and GRFs genes, may be considered as markers of growth competency.

Foliar Glyphosate Treatment Alters Transcript and Hormone Profiles in Crown Buds of Leafy Spurge and Induces Dwarfed and Bushy Phenotypes throughout its Perennial Lifecycle.

Leafy spurge ( L.) is an invasive weed of North America and its perennial nature attributed to underground adventitious buds (UABs) that undergo seasonal cycles of para-, endo-, and ecodormancy. Recommended rates of glyphosate (∼1 kg ha) destroy aboveground shoots but plants still regenerate vegetatively; therefore, it is considered glyphosate-tolerant. However, foliar application of glyphosate at higher rates (2.2-6.7 kg ha) causes sublethal effects that induce UABs to produce stunted, bushy phenotypes. We investigated the effects of glyphosate treatment (±2.24 kg ha) on vegetative growth, phytohormone, and transcript profiles in UABs under controlled environments during one simulated seasonal cycle. Because shoots derived from UABs of foliar glyphosate-treated plants produced stunted, bushy phenotypes, we could not directly determine if these UABs transitioned through seasonally induced endo- and ecodormancy. However, transcript abundance for leafy spurge dormancy marker genes and principal component analyses suggested that UABs of foliar glyphosate-treated plants transitioned through endo- and ecodormancy. Glyphosate treatment increased shikimate abundance in UABs 7 d after treatment; however, the abundance of shikimate gradually decreased as UABs transitioned through endo- and ecodormancy. The dissipation of shikimate over time suggests that glyphosate's target site was no longer affected, but these changes did not reverse the altered phenotypes observed from UABs of foliar glyphosate-treated leafy spurge. Transcript profiles further indicated that foliar glyphosate treatment significantly affected phytohormone biosynthesis and signaling, particularly auxin transport; gibberellic acid, abscisic acid and jasmonic acid biosynthesis; ethylene responses; and detoxification and cell cycle processes in UABs. These results correlated well with the available phytohormone profiles and altered phenotypes.

Natural variation in expression of genes associated with carotenoid biosynthesis and accumulation in cassava (Manihot esculenta Crantz) storage root.

BACKGROUND: Cassava (Manihot esculenta Crantz) storage root provides a staple food source for millions of people worldwide. Increasing the carotenoid content in storage root of cassava could provide improved nutritional and health benefits. Because carotenoid accumulation has been associated with storage root color, this study characterized carotenoid profiles, and abundance of key transcripts associated with carotenoid biosynthesis, from 23 landraces of cassava storage root ranging in color from white-to-yellow-to-pink. This study provides important information to plant breeding programs aimed at improving cassava storage root nutritional quality. RESULTS: Among the 23 landraces, five carotenoid types were detected in storage root with white color, while carotenoid types ranged from 1 to 21 in storage root with pink and yellow color. The majority of storage root in these landraces ranged in color from pale-to-intense yellow. In this color group, total ß-carotene, containing all-E-, 9-Z-, and 13-Z-ß-carotene isomers, was the major carotenoid type detected, varying from 26.13 to 76.72 %. Although no α-carotene was observed, variable amounts of a α-ring derived xanthophyll, lutein, was detected; with greater accumulation of α-ring xanthophylls than of ß-ring xanthophyll. Lycopene was detected in a landrace (Cas51) with pink color storage root, but it was not detected in storage root with yellow color. Based on microarray and qRT-PCR analyses, abundance of transcripts coding for enzymes involved in carotenoid biosynthesis were consistent with carotenoid composition determined by contrasting HPLC-Diode Array profiles from storage root of landraces IAC12, Cas64, and Cas51. Abundance of transcripts encoding for proteins regulating plastid division were also consistent with the observed differences in total ß-carotene accumulation. CONCLUSIONS: Among the 23 cassava landraces with varying storage root color and diverse carotenoid types and profiles, landrace Cas51 (pink color storage root) had low LYCb transcript abundance, whereas landrace Cas64 (intense yellow storage root) had decreased HYb transcript abundance. These results may explain the increased amounts of lycopene and total ß-carotene observed in landraces Cas51 and Cas64, respectively. Overall, total carotenoid content in cassava storage root of color class representatives were associated with spatial patterns of secondary growth, color, and abundance of transcripts linked to plastid division. Finally, a partial carotenoid biosynthesis pathway is proposed.

Phytohormone balance and stress-related cellular responses are involved in the transition from bud to shoot growth in leafy spurge.

BACKGROUND: Leafy spurge (Euphorbia esula L.) is an herbaceous weed that maintains a perennial growth pattern through seasonal production of abundant underground adventitious buds (UABs) on the crown and lateral roots. During the normal growing season, differentiation of bud to shoot growth is inhibited by physiological factors external to the affected structure; a phenomenon referred to as paradormancy. Initiation of shoot growth from paradormant UABs can be accomplished through removal of the aerial shoots (hereafter referred to as paradormancy release). RESULTS: In this study, phytohormone abundance and the transcriptomes of paradormant UABs vs. shoot-induced growth at 6, 24, and 72 h after paradormancy release were compared based on hormone profiling and RNA-seq analyses. Results indicated that auxin, abscisic acid (ABA), and flavonoid signaling were involved in maintaining paradormancy in UABs of leafy spurge. However, auxin, ABA, and flavonoid levels/signals decreased by 6 h after paradormancy release, in conjunction with increase in gibberellic acid (GA), cytokinin, jasmonic acid (JA), ethylene, and brassinosteroid (BR) levels/signals. Twenty four h after paradormancy release, auxin and ABA levels/signals increased, in conjunction with increase in GA levels/signals. Major cellular changes were also identified in UABs at 24 h, since both principal component and Venn diagram analysis of transcriptomes clearly set the 24 h shoot-induced growth apart from other time groups. In addition, increase in auxin and ABA levels/signals and the down-regulation of 40 over-represented AraCyc pathways indicated that stress-derived cellular responses may be involved in the activation of stress-induced re-orientation required for initiation of shoot growth. Seventy two h after paradormancy release, auxin, cytokinin, and GA levels/signals were increased, whereas ABA, JA, and ethylene levels/signals were decreased. CONCLUSION: Combined results were consistent with different phytohormone signals acting in concert to direct cellular changes involved in bud differentiation and shoot growth. In addition, shifts in balance of these phytohormones at different time points and stress-related cellular responses after paradormancy release appear to be critical factors driving transition of bud to shoot growth.

Glyphosate's impact on vegetative growth in leafy spurge identifies molecular processes and hormone cross-talk associated with increased branching.

BACKGROUND: Leafy spurge (Euphorbia esula) is a perennial weed that is considered glyphosate tolerant, which is partially attributed to escape through establishment of new vegetative shoots from an abundance of underground adventitious buds. Leafy spurge plants treated with sub-lethal concentrations of foliar-applied glyphosate produce new vegetative shoots with reduced main stem elongation and increased branching. Processes associated with the glyphosate-induced phenotype were determined by RNAseq using aerial shoots derived from crown buds of glyphosate-treated and -untreated plants. Comparison between transcript abundance and accumulation of shikimate or phytohormones (abscisic acid, auxin, cytokinins, and gibberellins) from these same samples was also done to reveal correlations. RESULTS: Transcriptome assembly and analyses confirmed differential abundance among 12,918 transcripts (FDR ≤ 0.05) and highlighted numerous processes associated with shoot apical meristem maintenance and stem growth, which is consistent with the increased number of actively growing meristems in response to glyphosate. Foliar applied glyphosate increased shikimate abundance in crown buds prior to decapitation of aboveground shoots, which induces growth from these buds, indicating that 5-enolpyruvylshikimate 3-phosphate (EPSPS) the target site of glyphosate was inhibited. However, abundance of shikimate was similar in a subsequent generation of aerial shoots derived from crown buds of treated and untreated plants, suggesting EPSPS is no longer inhibited or abundance of shikimate initially observed in crown buds dissipated over time. Overall, auxins, gibberellins (precursors and catabolites of bioactive gibberellins), and cytokinins (precursors and bioactive cytokinins) were more abundant in the aboveground shoots derived from glyphosate-treated plants. CONCLUSION: Based on the overall data, we propose that the glyphosate-induced phenotype resulted from complex interactions involving shoot apical meristem maintenance, hormone biosynthesis and signaling (auxin, cytokinins, gibberellins, and strigolactones), cellular transport, and detoxification mechanisms.

Polyphenol oxidase as a biochemical seed defense mechanism.

Seed dormancy and resistance to decay are fundamental survival strategies, which allow a population of seeds to germinate over long periods of time. Seeds have physical, chemical, and biological defense mechanisms that protect their food reserves from decay-inducing organisms and herbivores. Here, we hypothesize that seeds also possess enzyme-based biochemical defenses, based on induction of the plant defense enzyme, polyphenol oxidase (PPO), when wild oat (Avena fatua L.) caryopses and seeds were challenged with seed-decaying Fusarium fungi. These studies suggest that dormant seeds are capable of mounting a defense response to pathogens. The pathogen-induced PPO activity from wild oat was attributed to a soluble isoform of the enzyme that appeared to result, at least in part, from proteolytic activation of a latent PPO isoform. PPO activity was also induced in wild oat hulls (lemma and palea), non-living tissues that cover and protect the caryopsis. These results are consistent with the hypothesis that seeds possess inducible enzyme-based biochemical defenses arrayed on the exterior of seeds and these defenses represent a fundamental mechanism of seed survival and longevity in the soil. Enzyme-based biochemical defenses may have broader implications since they may apply to other defense enzymes as well as to a diversity of plant species and ecosystems.

The resemblance and disparity of gene expression in dormant and non-dormant seeds and crown buds of leafy spurge (Euphorbia esula).

BACKGROUND: Leafy spurge (Euphorbia esula L.) is a herbaceous perennial weed and dormancy in both buds and seeds is an important survival mechanism. Bud dormancy in leafy spurge exhibits three well-defined phases of para-, endo- and ecodormancy; however, seed dormancy for leafy spurge is classified as physiological dormancy that requires after-ripening and alternating temperature for maximal germination. Overlaps in transcriptome profiles between different phases of bud and seed dormancy have not been determined. Thus, we compared various phases of dormancy between seeds and buds to identify common genes and molecular processes, which should provide new insights about common regulators of dormancy. RESULTS: Cluster analysis of expression profiles for 201 selected genes indicated bud and seed samples clustered separately. Direct comparisons between buds and seeds are additionally complicated since seeds incubated at a constant temperature of 20°C for 21 days (21d C) could be considered paradormant (Para) because seeds may be inhibited by endosperm-generated signals, or ecodormant (Eco) because seeds germinate after being subjected to alternating temperature of 20:30°C. Since direct comparisons in gene expression between buds and seeds were problematic, we instead examined commonalities in differentially-expressed genes associated with different phases of dormancy. Comparison between buds and seeds ('Para to Endo buds' and '21d C to 1d C seeds'), using endodormant buds (Endo) and dormant seeds (1d C) as common baselines, identified transcripts associated with cell cycle (HisH4), stress response/transcription factors (ICE2, ERFB4/ABR1), ABA and auxin response (ABA1, ARF1, IAA7, TFL1), carbohydrate/protein degradation (GAPDH_1), and transport (ABCB2). Comparison of transcript abundance for the 'Eco to Endo buds' and '21d C to 1d C seeds' identified transcripts associated with ABA response (ATEM6), auxin response (ARF1), and cell cycle (HisH4). These results indicate that the physiological state of 21d C seeds is more analogous to paradormant buds than that of ecodormant buds. CONCLUSION: Combined results indicate that common molecular mechanisms associated with dormancy transitions of buds and seeds involve processes associated with ABA and auxin signaling and transport, cell cycle, and AP2/ERF transcription factors or their up-stream regulators.