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Increasing plant group productivity through latent genetic variation for cooperation

Historic yield advances in the major crops have to a large part been achieved by selection for improved productivity of groups of plant individuals such as high-density stands. Research suggests that such improved group productivity depends on "cooperative" traits (e.g. erect leaves, short stems) that - while beneficial to the group - decrease individual fitness under competition. This poses a problem for some traditional breeding approaches, especially when selection occurs at the level of individuals, because "selfish" traits will be selected for and reduce yields in high-density monocultures. One approach therefore has been to select individuals based on ideotypes with traits expected to promote group productivity. However, this approach is limited to architectural and physiological traits whose effects on growth and competition are relatively easy to anticipate. Here, we developed a general and simple method for the discovery of alleles promoting cooperation in plant stands. Our method is based on the game-theoretical premise that alleles increasing cooperation incur a cost to the individual but benefit the monoculture group. Testing the approach using the model plant Arabidopsis thaliana, we found a major effect locus where the rarer allele was associated with increased cooperation and productivity in high-density stands. The allele likely affects a pleiotropic gene, since we find that it is also associated with reduced root competition but higher resistance against disease. Thus, even though cooperation is considered evolutionarily unstable, conflicting selective forces acting on a pleiotropic gene might maintain latent genetic variation for it in nature. Such variation, once identified in a crop, could be rapidly leveraged in modern breeding programs and provide efficient routes to increase yields.

plant biology

Quantification of bioagressors induced yield gap for grain crops in France

Analyzing yield losses of crops is instrumental in sustaining high productivity. Here, we develop statistical modelling of yield losses by French department based on weather, soil and epidemiological data for 13 diseases and 12 insect pests of grain crops over 8 years. Those environmental factors explain up to 90% of yield variations (wheat). Weather and soil quality are first order determinants as manifested both by direct assessment and by strongly correlated yields of even taxonomically very different crops. Bioagressors are important second order determinants: losses of 5 qx/ha (~7%) on wheat and 2 qx/ha (~15%) on winter oilseed rape. Across models based on conceptually different achievable yields, only Septoria tritici and Sitobion avenae on wheat, and Ceutorhynchus picitarsis and Ceutorhynchus assimilis on winter oilseed rape are consistently and significantly detrimental to yield. Those bioagressors seem not fully controlled which is compatible with empirical observations in the non academical litterature.\n\nHighlightsO_LIGrain crop yields of similar growing seasons are highly correlated\nC_LIO_LIEnvironmental determinants explain up to 90% of yield variations\nC_LIO_LIBioagressors induced yield variations of 5 to 20%\nC_LIO_LISpecific pests and diseases with less than perfect control are identified\nC_LI

plant biology

Molecular and transcriptional structure of the petal and leaf circadian clock in Petunia hybrida

The plant circadian clock coordinates environmental signals with internal processes. We characterized the genomic and transcriptomic structure of the Petunia hybrida W115 clock in leaves and petals. We found three levels of evolutionary differences. First, PSEUDO-RESPONSE REGULATORS PhPRR5a, PhPRR5b, PhPRR7a, PhPRR7b, and GIGANTEA PhGI1 and PhGI2, differed in gene structure including exon number and deletions including the CCT domain of the PRR family. Second, leaves showed preferential day expression while petals tended to display night expression. Under continuous dark, most genes were delayed in leaves and petals. Importantly, photoperiod sensitivity of gene expression was tissue specific as TIMING OF CAB EXPRESSION PhNTOC1 was affected in leaves but not in petals, and PhPRR5b, PhPRR7b and the ZEITLUPE ortholog CHANEL, PhCHL, were modified in petals but not leaves. Third, we identified a strong transcriptional noise at different times of the day, and high robustness at dawn in leaves and dusk in petals, coinciding with the coordination of photosynthesis and scent emission. Our results indicate multilayered evolution of the Petunia clock including gene structure, number of genes and transcription patterns. The major transcriptional reprogramming of the clock in petals, with night expression may be involved in controlling scent emission in the dark.\n\nHighlightThe petunia leaf circadian clock shows maxima during the day while petal clock does it during the night. Reaction to dark is organ specific.

plant biology

Characterizing the serotonin biosynthesis pathway upon aphid infestation in Setaria viridis leaves

Setaria viridis (green foxtail millet), a short life-cycle C4 plant in the Gramineae, serves as a resilient crop that provides good yield even in dry and marginal land. Although S. viridis has been studied extensively in the last decade, its defense responses, in particular the chemical defensive metabolites that protect it against insect herbivory, are unstudied. To characterize S. viridis defense responses, we conducted transcriptomic and metabolomic assays of plants infested with aphids and caterpillars. Pathway enrichment analysis indicated massive transcriptomic changes that involve genes from amino acid biosynthesis and degradation, secondary metabolites and phytohormone biosynthesis. The Trp-derived metabolite serotonin was notably induced by insect feeding. Through comparisons with known rice serotonin biosynthetic genes, we identified several predicted S. viridis Trp decarboxylases and cytochrome P450 genes that were up-regulated in response to insect feeding. The function of one Trp decarboxylase was validated by ectopic expression and detection of tryptamine accumulation in Nicotiana tabacum. To validate the defensive properties of serotonin, we used an artificial diet assay to show reduced Rhopalosiphum padi aphid survival with increasing serotonin concentrations. This demonstrated that serotonin is a defensive metabolite in S. viridis and is fundamental for understanding the adaptation of it to biotic stresses.\n\nHIGHLIGHTA combined transcriptomic and metabolomic profiling of Setaria viridis leaves response to aphid and caterpillar infestation identifies the genes related to the biosynthesis of serotonin and their function in defense.

plant biology

miRNA-encoded peptide, miPEP858, regulates plant growth and development in Arabidopsis

MicroRNAs (miRNAs), small non-coding endogenous RNAs, are processed product of primary miRNAs (pri-miRNAs) and regulate target gene expression. pri-miRNAs have also been reported to encode small peptides, miRNA-Encoded Peptides (miPEPs). Though regulatory role of miPEPs has been speculated, no detailed study has been carried out to elucidate their function through developing knock-out mutants. Here, we report that pri-miR858a of Arabidopsis thaliana encodes a small peptide (miPEP858a) which regulates the expression of pri-miR858a leading to modulation in the expression of target genes involved in the plant growth and development as well as phenylpropanoid pathway. CRISPR-based miPEP858a-edited plants developed phenotypes similar to that of mature miR858-edited plants suggesting crucial role of miPEP858a in mediating miR585a function. miPEP858a-edited and miPEP858a overexpressing lines altered plant development and accumulated modulated levels of flavonoids due to changes in expression of associated genes. Exogenous treatment of synthetic-miPEP858a to the miPEP858a-edited plants complemented phenotypes and the gene function suggesting a significant role of miPEP858a in controlling the miR858 function and plant development.\n\nOne sentence summarySmall peptide, miPEP858a, encoded by primary miRNA for miR858a regulates plant growth, development and flavonoid biosynthesis\n\nThe authors responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors

plant biology

A diversity of traits contributes to salinity tolerance of wild Galapagos tomatoes seedlings

Traits of modern crops have been heavily selected in agriculture, causing the commercial lines to be more susceptible to harsh conditions, which their wild relatives are naturally better able to withstand. Understanding the developed mechanisms of tolerance present in wild relatives can enhance crop performance under stress. In this study, salinity tolerance traits of two species of wild tomato endemic to the Galapagos Islands, Solanum cheesmaniae and Solanum galapagense, were investigated. Since these tomatoes grow well despite being constantly splashed with seawater, they could be a valuable genetic resource for improving salinity tolerance in commercial tomatoes. To explore their potential, over 20 traits reflecting plant growth, physiology and ion content were recorded in 67 accessions of S. cheesmaniae and S. galapagense and two commercial tomato lines of Solanum lycopersicum. Salt treatments of 200 mM NaCl were applied for ten days, using supported hydroponics. Great natural variation was evident in the responses of the Galapagos tomatoes to salt stress and they also displayed greater tolerance to salt stress than the commercial lines tested, based on multivariate trait analyses. Although Galapagos tomatoes in general exhibited better tolerance to salt stress than the commercial lines tested, the accessions LA0317, LA1449 and LA1403 showed particularly high salinity tolerance based on growth maintenance under stress. Thus, Galapagos tomatoes should be further explored using forward genetic studies to identify and investigate the genes underlying their high tolerance and be used as a resource for increasing salinity tolerance of commercial tomatoes. The generated data, along with useful analysis tools, have been packaged and made publicly available via an interactive online application (https://github.com/mmjulkowska/La_isla_de_tomato) to facilitate trait selection and the use of Galapagos tomatoes for the development of salt tolerant commercial tomatoes.

plant biology

PICKLE recruits RETINOBLASTOMA RELATED 1 to Control Lateral Root Formation in Arabidopsis

Lateral root (LR) formation is an example of plant post-embryonic organogenesis event. LRs are issued from non-dividing cells entering consecutive steps of formative divisions, proliferation and elongation. The chromatin remodeling protein PICKLE negatively regulates auxin-mediated LR formation through a mechanism that is not yet known. Here we show that PICKLE interacts with RETINOBLASTOMA-RELATED 1 (RBR1) to repress the LATERAL ORGAN BOUNDARIES-DOMAIN 16 (LBD16) promoter activity. Since LBD16 function is required for the formative division of LR founder cells, repression mediated by the PKL-RBR1 complex negatively regulates formative division and LR formation. Inhibition of LR formation by PKL-RBR1 is counteracted by auxin indicating that in addition to auxin-mediated transcriptional responses, the fine-tuned process of LR formation is also controlled at the chromatin level in an auxin-signaling dependent manner.

plant biology

Fast, Efficient, and Precise Gene Editing in the Moss Physcomitrella patens

Recent years, the bryophyte moss Physcomitrella patens has become an emerging model organism for studying conserved signaling pathways and developmental processes during plant evolution. Its short life cycle, ease of cultivation, and high rate of homologous recombination have made it an ideal system for genetic analysis. However, the presence of highly redundant genes and the difficulty of isolating hypomorphic mutants have limited its broader use. Here we developed a simple, fast, and efficient method to generate customized mutants in P. patens. We show that transient cotransformation of CRISPR/Cas9 and oligonucleotide templates enables microindel knock-in with high efficiency and accuracy. Using this method, we generated strains carrying various types of mutations, including amino acid substitution, out-of-frame deletion/insertion, splice site alteration, and small tag integration. We also demonstrate that multiplex gene editing can be efficiently achieved to generate putative null and hypomorphic mutants of redundant genes in one step. Thus our method will not only simplify multiple-gene knockout, but also allows the generation of hypomorphic mutants of genes of interest, especially those that are essential for viability.

plant biology

A carbohydrate-binding protein, FLOURY ENDOSPERM 6 influences the initiation of A- and B-type starch granules in wheat.

Previously, we identified a quantitative trait locus on the group 4 chromosomes of Aegilops and bread wheat that controls B-type starch-granule content. Here, we identify a candidate gene by fine-mapping in Aegilops and confirm its function using wheat TILLING mutants. This gene is orthologous to the FLOURY ENDOSPERM 6 (FLO6) gene of rice and barley and the PTST2 gene of Arabidopsis. In Triticeae endosperm, reduction in the gene dose of functional FLO6 alleles results in reduction, or loss, of B-granules. This is due to repression of granule initiation in late-grain development, but has no deleterious impact on the synthesis of A-granules. The complete absence of functional FLO6, however, results in reduced numbers of normal A-type and B-type granules and the production of highly-abnormal granules that vary in size and shape. This polymorphous starch seen in a wheat flo6 triple mutant is similar to that observed in the barley mutant Franubet. Analysis of Franubet (fractured Nubet) starch suggests that the mutant A-granules are not fractured but compound, due to stimulation of granule initiation in plastids during early-grain development. Thus, in different situations in Triticeae, FLO6 either stimulates or represses granule initiation.

plant biology

An optimised protocol for isolation of RNA through laser capture microdissection of leaf material

Laser Capture Microdissection is a powerful tool that allows thin slices of specific cells types to be separated from one another. However, the most commonly used protocol, which involves embedding tissue in paraffin wax, results in severely degraded RNA. Yields from low abundance cell types of leaves are particularly compromised. We reasoned that the relatively high temperature used for sample embedding, and aqueous conditions associated with sample preparation prior to microdissection contribute to RNA degradation. Here we describe an optimized procedure to limit RNA degradation that is based on the use of low melting point wax as well as modifications to sample preparation prior to dissection, and isolation of paradermal, rather than transverse sections. Using this approach high quality RNA suitable for down-stream applications such as quantitative reverse transcriptase polymerase chain reactions or RNA-sequencing is recovered from microdissected bundle sheath strands and mesophyll cells of leaf tissue.

plant biology

GRANAR, a new computational tool to better understand the functional importance of root anatomy

Root hydraulic conductivity is an important determinant of plant water uptake capacity. In particular, the root radial conductivity is often thought to be a limiting factor along the water pathways between the soil and the leaf. The root radial conductivity is itself defined by cell scale hydraulic properties and anatomical features. However, quantifying the influence of anatomical features on the radial conductivity remains challenging due to complex, and time-consuming, experimental procedures.\n\nWe present a new computation tool, the Generator of Root ANAtomy in R (GRANAR) that can be used to rapidly generate digital versions of root anatomical networks. GRANAR uses a limited set of root anatomical parameters, easily acquired with existing image analysis tools. The generated anatomical network can then be used in combination with hydraulic models to estimate the corresponding hydraulic properties.\n\nWe used GRANAR to re-analyse large maize (Zea mays) anatomical datasets from the literature. Our model was successful at creating virtual anatomies for each experimental observation. We also used GRANAR to generate anatomies not observed experimentally, over wider ranges of anatomical parameters. The generated anatomies were then used to estimate the corresponding radial conductivities with the hydraulic model MECHA. This enabled us to quantify the effect of individual anatomical features on the root radial conductivity. In particular, our simulations highlight the large importance of the width of the stele and the cortex.\n\nGRANAR is an open-source project available here: http://granar.github.io\n\nOne-Sentence summaryGenerator of Root ANAtomy in R (GRANAR) is a new open-source computational tool that can be used to rapidly generate digital versions of root anatomical networks.

plant biology

Anaesthesia with diethyl ether impairs jasmonate signalling in the carnivorous plant Venus flytrap (Dionaea muscipula).

General anaesthetics are compounds that induce loss of responsiveness to environmental stimuli in animals and humans. The primary site of general anaesthetic action is the nervous system, where anaesthetics inhibit neuronal transmission. Although plants do not have neurons, they generate electrical signals in response to biotic and abiotic stresses. Here, we investigated the effect of the general volatile anaesthetic diethyl ether on the ability to sense potential prey or herbivore attacks in the carnivorous plant Venus flytrap (Dionaea muscipula). We monitored trap movement, electrical signalling, phytohormone accumulation and gene expression in response to the mechanical stimulation of trigger hairs and wounding under diethyl ether treatment. Diethyl ether completely inhibited the generation of action potentials and trap closing reactions, which were easily and rapidly restored when the anaesthetic was removed. Diethyl ether also inhibited the later response: jasmonate (JA) accumulation and expression of JA-responsive genes. However, external application of JA bypassed the inhibited action potentials and restored gene expression under diethyl ether anaesthesia, indicating that downstream reactions from JA are not inhibited. Thus, the Venus flytrap cannot sense prey or a herbivore attack under diethyl ether treatment. This is an intriguing parallel to the effect of anaesthesia on animals and humans.\n\nHighlightCarnivorous plant Venus flytrap (Dionaea muscipula) is unresponsive to insect prey or herbivore attack due to impaired electrical and jasmonate signalling under general anaesthesia induced by diethyl ether.

plant biology

Tomato root transformation followed by inoculation with Ralstonia solanacearum for straightforward genetic analysis of bacterial wilt disease

Ralstonia solanacearum is a devastating soil borne vascular pathogen that is able to infect a large range of plant species, causing an important threat to agriculture. However, the Ralstonia model is considerably under-explored in comparison to other models involving bacterial plant pathogens, such as Pseudomonas syringae in Arabidopsis. Research targeted to understanding the interaction between Ralstonia and crop plants is essential to develop sustainable solutions to fight against bacterial wilt disease, but is currently hindered by the lack of straightforward experimental assays to characterize the different components of the interaction in native host plants. In this scenario, we have developed an easy method to perform genetic analysis of Ralstonia infection of tomato, a natural host of Ralstonia. This method is based on Agrobacterium rhizogenes-mediated transformation of tomato roots, followed by Ralstonia soil-drenching inoculation of the resulting plants, containing transformed roots expressing the construct of interest. The versatility of the root transformation assay allows performing either gene overexpression or gene silencing mediated by RNAi. As a proof of concept, we used this method to show that RNAi-mediated silencing of SlCESA6 of tomato roots conferred resistance to Ralstonia. Here, we describe this method in detail, enabling genetic approaches to understand bacterial wilt disease in a relative short time and with small requirements of equipment and plant growth space.\n\nSUMMARYA versatile method for tomato root transformation followed by inoculation with Ralstonia solanacearum to perform straightforward genetic analysis for the study of bacterial wilt disease.

plant biology

Brassinosteroid signaling controls PILS-dependent nuclear auxin input in Arabidopsis thaliana

Auxin and brassinosteroids (BR) are crucial growth regulators and display overlapping functions during plant development. Here, we reveal an alternative phytohormone crosstalk mechanism, revealing that brassinosteroid signaling controls nuclear abundance of auxin. We performed a forward genetic screen for imperial pils (imp) mutants that enhance the overexpression phenotypes of PIN-LIKES (PILS) putative intracellular auxin transport facilitator. Here we report that the imp1 mutant is defective in the brassinosteroid-receptor BRI1. Our data reveals that BR signaling transcriptionally and posttranslationally represses accumulation of PILS proteins at the endoplasmic reticulum, thereby increasing nuclear abundance and signaling of auxin. We demonstrate that this alternative phytohormonal crosstalk mechanism integrates BR signaling into auxin-dependent organ growth rates and likely has widespread importance for plant development.

plant biology

The Synchytrium endobioticum AvrSen1 triggers a Hypersensitive Response in Sen1 potatoes while natural variants evade detection

Synchytrium endobioticum is an obligate biotrophic fungus of the phylum Chytridiomycota. It causes potato wart disease, has a world-wide quarantine status and is included on the HHS and USDA Select Agent list. S. endobioticum isolates are grouped in pathotypes based on their ability to evade host-resistance in a set of differential potato varieties. So far, thirty-nine pathotypes are reported. A single dominant gene (Sen1) governs pathotype 1 resistance and we anticipated that the underlying molecular model would involve a pathogen effector (AvrSen1) that is recognized by the host. The S. endobioticum specific secretome of fourteen isolates representing six different pathotypes was screened for effectors specifically present in pathotype 1(D1) isolates but absent in others. We identified a single AvrSen1 candidate. Expression of this candidate in potato Sen1 plants showed a specific hypersensitive response, which co-segregated with the Sen1 resistance in potato populations. No HR was obtained with truncated genes found in pathotypes that evaded recognition by Sen1. These findings established that our candidate gene was indeed Avrsen1. The S. endobioticum AvrSen1 is a single copy gene and encodes a 376 amino acid protein without predicted function or functional domains, and is the first effector gene identified in Chytridiomycota, an extremely diverse yet underrepresented basal lineage of fungi.\n\nAuthor SummaryPlant pathogens can have a great social and economic impact, and are a continuous threat to food security. A clear example is Synchytrium endobioticum, the fungus causing potato wart disease. The impact of the pathogen, lack of effective chemical control agents and the longevity of resting spores produced by the pathogen led to a world-wide quarantine status for S. endobioticum. Strict phytosanitary measures and the use of resistance potato varieties are currently the only way to prevent the spread of the disease. The emergence of new pathotypes that overcome resistance urged to study the underlying molecular mechanisms of S. endobioticum recognition by the plant. Here we describe the identification of the first effector (AvrSen1) of S. endobioticum that is recognized by the Sen1 resistance gene product. Also, we report the loss of AvrSen1 in other pathotypes thus avoiding recognition by the plant and triggering immune responses. AvrSen1 represents the first effector to be identified in the basal fungal lineage Chytridiomycota. The discovery of AvrSen1 provides an important tool to manage potato wart disease. Moreover, knowledge about Chytridiomycota effectors will shed light on other (pathogenic) interactions and the co-evolution of Chytridiomycota species with their hosts.

plant biology

X-ray phase contrast imaging of Vitis spp. buds shows freezing pattern and correlation between volume and cold hardiness

Grapevine (Vitis spp.) buds must survive winter temperatures in order to resume growth when suitable conditions return in spring. They do so by developing cold hardiness through deep supercooling, but the mechanistic process of supercooling in buds remains largely unknown. Here we use synchrotron X-ray phase contrast imaging to study cold hardiness-related characteristics of V. amurensis, V. riparia, and V. vinifera buds: time-resolved 2D imaging was used to visualize freezing; and microtomography was used to evaluate morphological changes during deacclimation. Bud cold hardiness was determined (low temperature exotherms; LTEs) using needle thermocouples during 2D imaging as buds were cooled with a N2 gas cryostream. Resolution in 2D imaging did not allow for ice crystal identification, but freezing was assessed due to movement of tissues coinciding with LTE values. Freezing was observed to propagate from the center of the bud toward the outer bud scales. The freezing events observed lasted several minutes. Additionally, loss of supercooling ability appears to be correlated with increases in bud tissue volume during the process of deacclimation, but major increases in volume occur after most of the supercooling ability is lost, suggesting growth resumption processes are limited by deacclimation state.\n\nHighlightX-ray phase contrast imaging shows freezing occurs over several minutes and propagates from center toward tip of Vitis spp. buds. Incremental increase in bud volume correlates with cold deacclimation

plant biology

Breeding for broad-spectrum disease resistance alters the maize leaf microbiome

O_LIPlant genotype strongly affects disease resistance, and also influences the composition of the leaf microbiome. However, these processes have not been studied and linked in the microevolutionary context of breeding for improved disease resistance. We hypothesized that broad-spectrum disease resistance alleles also affect colonization by non-pathogenic symbionts.\nC_LIO_LIQuantitative trait loci (QTL) conferring resistance to multiple fungal pathogens were introgressed into a disease-susceptible maize inbred line. Bacterial and fungal leaf microbiomes of the resulting near-isogenic lines were compared to the microbiome of the disease-susceptible parent line at two timepoints in multiple fields.\nC_LIO_LIIntrogression of QTL from disease-resistant lines strongly shifted the relative abundance of diverse fungal and bacterial taxa in both 3-week-old and 7-week-old plants. Nevertheless, the effects on overall community structure and diversity were minor and varied among fields and years. Contrary to our expectations, host genotype effects were not any stronger in fields with high disease pressure than in uninfected fields, and microbiome succession over time was similar in heavily infected plants and uninfected plants.\nC_LIO_LIThese results show that introgressed QTL can greatly improve broad-spectrum disease resistance while having only limited and inconsistent pleiotropic effects on the leaf microbiome in maize.\nC_LI

plant biology

Structural imaging of native cryo-preserved secondary cell walls reveals presence of macrofibrils composed of cellulose, lignin and xylan.

The woody secondary cell walls of plants are the largest repository of renewable carbon biopolymers on the planet. These walls are made principally from cellulose and hemicelluloses and are impregnated with lignin. Despite their importance as the main load bearing structure for plant growth, as well as their industrial importance as both a material and energy source, the precise arrangement of these constituents within the cell wall is not yet fully understood. We have adapted low temperature scanning electron microscopy (cryo-SEM) for imaging the nanoscale architecture of angiosperm and gymnosperm cell walls in their native hydrated state. Our work confirms that cell wall macrofibrils, cylindrical structures with a diameter exceeding 10 nm, are a common feature of the native hardwood and softwood samples. We have observed these same structures in Arabidopsis thaliana secondary cell walls, enabling macrofibrils to be compared between mutant lines that are perturbed in cellulose, hemicellulose and lignin formation. Our analysis indicates that the macrofibrils in Arabidopsis cell walls are composed, at least partially, of cellulose, xylan and lignin. This study is a useful additional approach for investigating the native nanoscale architecture and composition of hardwood and softwood secondary cell walls and demonstrates the applicability of Arabidopsis genetic resources to relate fibril structure with wall composition and biosynthesis.

plant biology