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Results for “Plant Biology”

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bioRxiv2019

Morpho-anatomical observations on Phyllanthus of Southwestern Bangladesh with two new records for Bangladesh

An extensive floristic survey was done to annotate Phyllanthus of southwestern Bangladesh from 2015 to 2018. In total, 2189 individuals of Phyllanthus were counted and identified as eight different species (five herbs, two trees and a shrub). All species were examined following both morphological and anatomical methods, based on taxonomic notes. The listed species were Phyllanthus acidus, Phyllanthus amarus, Phyllanthus debilis, Phyllanthus emblica, Phyllanthus niruri, Phyllanthus urinaria, Phyllanthus reticulatus and Phyllanthus virgatus. Among them, Phyllanthus amarus and Phyllanthus debilis were listed for the first time from Bangladesh during this study period.

plant biology↗

bioRxiv2019

Extension of the in vivo haploid induction system from diploid maize to hexaploid wheat

Doubled haploid breeding technology has been one of the most important techniques for accelerating crop breeding. In compare to in vivo haploid induction in maize, which is efficient and background independent, wheat haploid production by interspecific hybridization pollinated with maize is influenced by genetic background and requires rescue of young embryos. Here, we analyzed the homologues of maize haploid induction gene MTL/ZmPLA1/NLD in several crop species systematically, the homologues are highly conserved in sorghum, millet and wheat etc. Since wheat is a very important polyploidy crop, as a proof of concept, we demonstrated that the in vivo haploid induction method could be extended from diploid maize to hexaploid wheat by knocking out the wheat homologues (TaPLAs). Result showed that double knock-out mutation could trigger wheat haploid induction at ~ 2%-3%, accompanied by 30% - 60% seed setting rate. The performance of haploid wheat individual showed shorter plant, narrower leaves and male sterile. Our results also revealed that knockout of TaPLA-A and TaPLA-D do not affect pollen viability. This study not only confirmed the function of the induction gene and explored a new approach for haploid production in wheat, but also provided an example that the in vivo haploid induction could be applied in more crop species with different ploidy levels. Furthermore, by combining with gene editing, it would be a fast and powerful platform for traits improvement in polyploidy crops breeding.

plant biology↗

bioRxiv2019

Progress toward the identification and stacking of crucial domestication traits in pennycress

The oilseed species Thlaspi arvense (pennycress) is being domesticated as a new crop that can provide both important ecosystem services and intensify farmland output. Through the use of high throughput sequencing and phenotyping, along with classical mutagenesis key traits needed for pennycress domestication have been identified. Domestication traits identified herein include reduced pod shatter, early maturity, reduced seed glucosinolate levels, and improved oil fatty acid content. By taking advantage of pennycress close genetic relationship with Arabidopsis thaliana, the causative mutations responsible for each of these traits have been identified. These mutations have been used to develop molecular markers to begin to stack the traits into individual lines.

plant biology↗

bioRxiv2019

Arabidopsis glutathione reductase 2 is indispensable in plastids, while mitochondrial glutathione is safeguarded by additional reduction and transport systems

O_LIA highly negative glutathione redox potential (EGSH) is maintained in the cytosol, plastids and mitochondria of plant cells to support fundamental processes, including antioxidant defence, redox regulation and iron-sulfur cluster biogenesis. Out of two glutathione reductase (GR) proteins in Arabidopsis, GR2 is predicted to be dual-targeted to plastids and mitochondria, but its differential roles in these organelles remain unclear.\nC_LIO_LIWe dissected the role of GR2 in organelle glutathione redox homeostasis and plant development using a combination of genetic complementation and stacked mutants, biochemical activity studies, immunogold labelling and in vivo biosensing.\nC_LIO_LIOur data demonstrate that GR2 is dual-targeted to plastids and mitochondria, but embryo lethality of gr2 null mutants is caused specifically in plastids. Whereas lack of mitochondrial GR2 leads to a partially oxidised glutathione pool in the matrix, the ABC transporter ATM3 and the mitochondrial thioredoxin system provide functional backup and maintain plant viability.\nC_LIO_LIWe identify GR2 as essential in the plastid stroma, where it counters GSSG accumulation and developmental arrest. By contrast a functional triad of GR2, ATM3 and the thioredoxin system in the mitochondria provides resilience to excessive glutathione oxidation.\nC_LI

plant biology↗

bioRxiv2019

Intronic heterochromatin prevents cryptic transcription initiation in Arabidopsis

Intronic transposable elements (TEs) comprise a large proportion in eukaryotic genomes, but how they regulate the host genes remains to be explored. Our forward genetic screen disclosed the plant specific RNA polymerases IV and V in suppressing intronic TE-mediated cryptic transcription initiation of a chimeric transcripts at FLC (FLCTE). Initiation of FLCTE transcription is blocked by the locally formed intronic heterochromatin, which is directly associated with RNA Pol V to inhibit the entry of RNA Pol II and the occupancy of H3K4 methylation. Genome-wide Pol II Ser5p native elongation transcription sequencing revealed that this is a common mechanism among intronic heterochromatin-containing genes. This study sheds light on deeply understanding the function of intronic heterochromatin on host genes expression in eukaryotic genome.

plant biology↗

bioRxiv2019

Plasma membrane associated Receptor Like Kinases relocalise to plasmodesmata in response to osmotic stress.

Plasmodesmata act as key elements in intercellular communication, coordinating processes related to plant growth, development and responses to environmental stresses. While many of the developmental, biotic and abiotic signals are primarily perceived at the plasma membrane (PM) by receptor proteins, plasmodesmata also cluster receptor-like activities and whether or not these two pathways interact is currently unknown.\n\nHere we show that specific PM-located Leucine-Rich-Repeat Receptor-Like-Kinases (LRR-RLKs), KIN7 and IMK2, which under optimal growth conditions are absented from plasmodesmata, rapidly relocate and cluster to the pores in response to osmotic stress. This process is remarkably fast, it is not a general feature of PM-associated proteins and is independent of sterol- and sphingolipid-membrane composition. Focusing on KIN7, previously reported to be involved in stress responses, we show that relocalisation upon mannitol depends on KIN7 phosphorylation. Loss-of-function mutation in KIN7 induces delay in lateral root (LR) development and the mutant is affected in the root response to mannitol stress. Callose-mediated plasmodesmata regulation is known to regulate LR development. We found that callose levels are reduced in kin7 mutant background with a root phenotype resembling ectopic expression of PdBG1, an enzyme that degrades callose at the pores. Both the LR and callose phenotypes can be complemented by expression of KIN7 -wild-type and -phosphomimic variants but not by KIN7 phosphodead mutant which fails to relocalise at plasmodesmata. Together the data indicate that re-organisation of RLKs to plasmodesmata is important for the regulation of callose and LR development as part of the plant response to osmotic stress.

plant biology↗

bioRxiv2019

Identifying and validating housekeeping hybrid Prunus sp. genes for root gene-expression studies

Prunus rootstock belonging to subgenera Amygdalus (peach), Prunus (plum) and Cerasus (cherry) are either from the same species as the scion or another one. The number of inter-species (including inter-subgenera) hybrids have increased as a result of efforts to broaden the genetic basis for biotic and abiotic resistance/tolerance. Identifying genes associated with important traits and responses requires expression analysis. Relative quantification is the simplest and most popular alternative, which requires reference genes (housekeeping) to normalize RT-qPCR data. However, there is a scarcity of validated housekeeping genes for hybrid Prunus rootstock species. This research aims to increase the number of housekeeping genes suitable for Prunus rootstock expression analysis.\n\nTwenty-one candidate housekeeping genes were pre-selected from previous RNAseq data that compared the response of root transcriptomes of two rootstocks subgenera to hypoxia treatment, Mariana 2624 (P. cerasifera Ehrh.x P. munsoniana W. Wight & Hedrick), and Mazzard F12/1 (P. avium L.). Representing groups of low, intermediate or high levels of expression, the genes were assayed by RT-qPCR at 72 hours of hypoxia treatment and analyzed with NormFinder software. A sub-set of seven housekeeping genes that presented the highest level of stability were selected, two with low levels of expression (Unknown 3, Unknown 7) and five with medium levels (GTB 1, TUA 3, ATPase P, PRT 6, RP II). The stability of these genes was evaluated under different stress conditions, cold and heat with the hybrid Mariana 2624 and N nutrition with the hybrids Colt (P. avium x P. pseudocerasus Lindl.) and Garnem [P. dulcis Mill.x (P. persica L.x P. davidiana Carr.)]. The algorithms of geNorm and BestKeeper software also were used to analyze the performance of these genes as housekeepers.\n\nStability rankings varied according to treatments, genotypes and the software for evaluation, but the gene GBT 1 often had the highest ranking. However, most of the genes are suitable depending on the stressor and/or genotype to be evaluated. No optimal number of reference genes could be determined with geNorm software when all conditions and genotypes were considered. These results strongly suggest that relative RT-qPCR should be analyzed separately with their respective best housekeeper according to the treatment and/or genotypes in Prunus spp rootstocks.

plant biology↗

bioRxiv2019

Protein engineering expands the effector recognition profile of a rice NLR immune receptor

Plant NLR receptors detect pathogen effectors and initiate an immune response. Since their discovery, NLRs have been the focus of protein engineering to improve disease resistance. However, this has proven challenging, in part due to their narrow response specificity. Here, we used structure-guided engineering to expand the response profile of the rice NLR Pikp to variants of the rice blast pathogen effector AVR-Pik. A mutation located within an effector binding interface of the integrated Pikp-HMA domain increased the binding affinity for AVR-Pik variants in vitro and in vivo. This translates to an expanded cell death response to AVR-Pik variants previously unrecognized by Pikp in planta. Structures of the engineered Pikp-HMA in complex with AVR-Pik variants revealed the mechanism of expanded recognition. These results provide a proof-of-concept that protein engineering can improve the utility of plant NLR receptors where direct interaction between effectors and NLRs is established, particularly via integrated domains.

plant biology↗

bioRxiv2019

GIGANTEA recruits deubiquitylases, UBP12 and UBP13, to regulate accumulation of the ZTL photoreceptor complex

To remain synchronous with the environment, plants constantly survey daily light conditions using an array of photoreceptors and adjust their circadian rhythms accordingly. ZEITLUPE (ZTL), a blue light photoreceptor with E3 ubiquitin ligase activity, communicates end-of-day light conditions to the circadian clock. To function properly, ZTL protein must accumulate but not destabilize target clock transcription factors before dusk, while in the dark ZTL mediates degradation of target proteins. It is not clear how ZTL can accumulate to high levels in the light while its targets remain stable. Two deubiquitylating enzymes, UBIQUITIN-SPECIFIC PROTEASE 12 and UBIQUITIN-SPECIFIC PROTEASE 13 (UBP12 and UBP13), which have opposite genetic and biochemical functions to ZTL, were shown to associate with the ZTL protein complex. Here we demonstrate that the ZTL light-dependent interacting partner, GIGANTEA (GI), recruits UBP12 and UBP13 to the ZTL photoreceptor complex. We show that loss of UBP12 and UBP13 reduces ZTL and GI protein levels through a post-transcriptional mechanism. Furthermore, the ZTL target protein TOC1 is unable to accumulate to normal levels in ubp mutants, indicating that UBP12 and UBP13 are necessary to stabilize clock transcription factors during the day. Our results demonstrate that the ZTL photoreceptor complex contains both ubiquitin-conjugating and -deconjugating enzymes, and that these two opposing enzyme types are necessary for the complex to properly regulate the circadian clock. This work also shows that deubiquitylating enzymes are a core design element of circadian clocks that is conserved from plants to animals.

plant biology↗

bioRxiv2019

Chitin perception in plasmodesmata identifies subcellular, context-specific immune signalling in plants

The plasma membrane (PM) that lines plasmodesmata has a distinct protein and lipid composition, underpinning specific regulation of these connections between cells. The plasmodesmal PM can integrate extracellular signals differently from the cellular PM, but it is not known how this specificity is established or how a single stimulus can trigger independent signalling cascades in neighbouring membrane domains. Here we have used the fungal elicitor chitin to investigate signal integration and responses at the plasmodesmal PM. We found that the plasmodesmal PM employs a receptor complex composed of the LysM receptors LYM2 and LYK4 which respectively change their location and interactions in response to chitin. Downstream, signalling is transmitted via a specific phosphorylation signature of an NADPH oxidase and localised callose synthesis that causes plasmodesmata closure. This demonstrates the plasmodesmal PM deploys both plasmodesmata-specific components and differential activation of PM-common components to independently integrate an immune signal.

plant biology↗

bioRxiv2019

Intraspecific root and leaf trait variation with tropical forest successional status: consequences for community-weighted patterns

O_LIQuantifying the dimensions and magnitude of intraspecific root trait variation is key to understanding the functional trade-offs in the belowground plant strategies of tropical forest trees. Additionally, accurately measuring how belowground functional trait variation relates to soil environment and forest age is crucial to tropical forest modeling efforts.\nC_LIO_LIWe sampled leaf and root morphologies from 423 juvenile trees of 72 species from 14 Angiosperm families along a 6.6 km transect that corresponded to an environmental gradient in decreasing soil fertility and texture with increasing forest age.\nC_LIO_LIWe observed within-lineage conservative functional trait-shifts in root and leaf morphological traits along the transect. From secondary to primary forest, average leaf area increased 7 cm2 and average root system diameter increased 0.4 mm. Mean specific leaf area decreased by 0.8 m2 kg-1, specific root length decreased by 3.5 m kg-1, and root branching intensity decreased by 0.3 tips cm-1. Leaf thickness and root tissue density showed no change.\nC_LIO_LIWe coupled trait measurements to a network of 164 1/16th-ha plots across a Chinese tropical forest reserve, to scale individual trait measurements up to the community-level, accounting for forest age.\nC_LIO_LIFor most traits, intraspecific trait variation negatively covaried with species compositional turnover between plots in younger versus older forest to compound and create greater community-weighted differences in trait values than would be observed if intraspecific variation in traits with forest age was not accounted for.\nC_LIO_LISummary Root morphologies are variable with local scale variation in soil fertility and texture. Accurately understanding broader (i.e. forest)-scale patterns in root functional traits, requires attention to underlying environmental variation in soil resources, which interacts with environmental filtering of plant communities.\nC_LI

plant biology↗

bioRxiv2019

Comparative analysis of perennial and annual Phaseolus seed nutrient concentrations

Malnutrition is a global public health concern and identifying mechanisms to elevate the nutrient output of crops may minimize nutrient deficiencies. Perennial grains within an agroforestry context offers one solution. The development and integration of perennial crops for food has critically influenced dialogue on the ecological intensification of agriculture and agroforestry. However, the nutrient compositions of wild, perennial, herbaceous species, such as those related to the common bean (Phaseolus vulgaris) are not well known. In this study, seed amino acid and ion concentrations of perennial and annual Phaseolus species were quantified using ionomics and mass spectrometry. No statistical difference was observed for Zn, toxic ions (e.g. As) or essential amino acid concentrations (except threonine) between perennial and annual Phaseolus species. However, differences were observed for some nutritionally important ions among and within lifespan groups. Ca, Cu, Fe, Mg, Mn, and P concentrations were higher in annual species. Intraspecific variability in ion concentrations and amino acids was observed within species; further, ion concentrations and amino acids differ among annual species and among perennial species. Ion and amino acid concentration appear to be largely independent of each other. These results suggest variability in ion and amino acid concentrations exist in nature. As new crop candidates are considered for ecological services, nutritional quality should be optimized to maximize nutrient output of sustainable food crops.

plant biology↗

bioRxiv2019

The de novo reference genome and transcriptome assemblies of the wild tomato species Solanum chilense

BackgroundWild tomato species, like Solanum chilense, are important germplasm resources for enhanced biotic and abiotic stress resistance in tomato breeding. In addition, S. chilense serves as a model system to study adaptation of plants to drought and to investigate the evolution of seed banks. However to date, the absence of a well annotated reference genome in this compulsory outcrossing, very diverse species limits in-depth studies on the genes involved.\n\nFindingsWe generated [~]134 Gb of DNA and 157 Gb of RNA sequence data of S chilense, which yielded a draft genome with an estimated length of 914 Mb in total encoding 25,885 high-confidence (hc) predicted gene models, which show homology to known protein-coding genes of other tomato species. Approximately 71% (18,290) of the hc gene models are additionally supported by RNA-seq data derived from leaf tissue samples. A benchmarking with Universal Single-Copy Orthologs (BUSCO) analysis of predicted gene models retrieved 93.3% BUSCO genes, which is in the current range of high-quality genomes for non-inbred plants. To further verify the genome annotation completeness and accuracy, we manually inspected the NLR resistance gene family and assessed its assembly quality. We revealed the existence of unique gene families of NLRs to S. chilense. Comparative genomics analyses of S. chilense, cultivated tomato S. lycopersicum and its wild relative S. pennellii revealed similar levels of highly syntenic gene clusters between the three species.\n\nConclusionsWe generated the first genome and transcriptome sequence assembly for the wild tomato species Solanum chilense and demonstrated its value in comparative genomics analyses. We make these genomes available for the scientific community as an important resource for studies on adaptation to biotic and abiotic stress in Solanaceae, on evolution of self-incompatibility, and for tomato breeding.

plant biology↗

bioRxiv2019

Impact of within-tree organ distances on floral induction and fruit growth in apple tree: implication of carbon and gibberellin organ contents

In plants, organs are inter-dependent for growth and development. Here, we aimed to investigate the distance at which interaction between organs operates and the relative contribution of within-tree variation in carbohydrate and hormonal contents on floral induction and fruit growth, in a fruit tree case study. Manipulations of leaf and fruit numbers were performed in two years on Golden delicious apple trees, at the shoot or branch scale or one side of Y-shape trees. For each treatment, floral induction proportion and mean fruit weight were recorded. Gibberellins content in shoot apical meristems, photosynthesis, and non-structural carbohydrate concentrations in organs were measured. Floral induction was promoted by leaf presence and fruit absence but was not associated with non-structural content in meristems. This suggests a combined action of promoting and inhibiting signals originating from leaves and fruit, and involving gibberellins. Nevertheless, these signals act at short distance only since leaf or fruit presence at long distances had no effect on floral induction. Conversely, fruit growth was affected by leaf presence even at long distances when sink demands were imbalanced within the tree, suggesting long distance transport of carbohydrates. We thus clarified the inter-dependence and distance effect among organs, therefore their degree of autonomy that appeared dependent on the process considered, floral induction or fruit growth.

plant biology↗

bioRxiv2019

Spatiotemporal restriction of FUSCA3 expression by class I BPC promotes ovule development and coordinates embryo and endosperm growth

Spatiotemporal regulation of gene expression plays an important role in developmental timing in plants and animals. FUSCA3 regulates the transition between different phases of development by acting as a link between different hormonal pathways in Arabidopsis. However, the mechanisms governing its spatiotemporal expression patterns are poorly understood. Here, we show that FUS3 is expressed in the chalaza and funiculus of the mature ovule and seed, but is repressed in the embryo sac, integuments and endosperm. FUS3 repression requires class I BASIC PENTACYSTEINE (BPC) proteins, which directly bind to the FUS3 locus and restrict its expression pattern. During vegetative and reproductive development, derepression of FUS3 in bpc1/2 or pML1:FUS3 misexpression lines results in dwarf plants carrying defective flowers and aborted ovules. Post-fertilization, ectopic FUS3 expression in the endosperm increases endosperm nuclei proliferation and seed size and delays or arrests embryo development. These phenotypes are rescued in bpc1/2 fus3-3. Lastly, class I BPCs interact with FIS-PRC2 (FERTILIZATION-INDEPENDENT SEED-Polycomb Repressive Complex 2), which represses FUS3 in the endosperm. We propose that BPC1/2 promotes the transition from reproductive to seed development by repressing FUS3 in ovule integuments. After fertilization, BPC1/2 and FIS-PRC2 repress FUS3 in the endosperm to coordinate endosperm and embryo growth.

plant biology↗

bioRxiv2019

A mobile ELF4 delivers circadian temperature information from shoots to roots

The circadian clock is synchronized by environmental cues, mostly by light and temperature. Elucidating how the plant circadian clock responds to temperature oscillations is crucial to understand plant responsiveness to the environment. Here we found a prevalent temperature-dependent function of the Arabidopsis clock component ELF4 (EARLY FLOWERING 4) in the root clock. Micrografting assays and mathematical analyses show that ELF4 moves from shoots to regulate rhythms in roots. ELF4 movement does not convey photoperiodic information but trafficking is essential to control the period of the root clock in a temperature-dependent manner. At low temperatures, ELF4 mobility is favored and results in a slow-paced root clock while high temperatures decrease movement, leading to a fast clock. Hence, the mobile ELF4 delivers temperature information and establishes a shoot-to-root dialogue that sets the pace of the clock in roots.

plant biology↗

bioRxiv2019

Defective photosynthetic adaptation mechanism in winter restricts the introduction of overwintering plant to high latitudes

Because of the need for agriculture and landscaping, many overwintering evergreen and biennial species that maintain green leaves over winter were introduced to higher latitudes. The green leaves of introduced overwintering species have to withstand a harsher winter, especially lower temperature, than in their native region of origin. Although the responses and adaptability of photosynthetic apparatus to winter conditions in native overwintering species were widely studied, the experimental results on the introduced overwintering species are very limited. Here, the photosynthetic adaptability during winter was analyzed in two native overwintering species, pine (woody plants), winter wheat (herb), and two introduced overwintering species, bamboo (woody plants), lilyturf (herb). The native species exhibited higher capacity for photosynthetic CO2 fixation and lower susceptibility for photoinhibition than introduced species during winter. Photosynthesis related proteins, such as PsbA, PsaA, Rubisco and Lhcb1, were marginally affected in native species, but significantly degraded in introduced species during winter. More interestingly, the PSII photoinhibition was mainly caused by up-regulation of photoprotection mechanism, non-photochemical quenching, in native species, but by photodamage in introduced species. This study indicates that the growth and survival of introduced overwintering species is limited by their photosynthetic adaptability to the harsher winter conditions at high latitudes.

plant biology↗

bioRxiv2019

Phosphatase PP2C6 negatively regulates phosphorylation status of Pti1b kinase, a regulator of flagellin-triggered immunity in tomato

Plant immune responses, including the production of reactive oxygen species (ROS), are triggered when pattern recognition receptors (PRR) become activated upon detection of microbe-associated molecular patterns (MAMPs). Receptor-like cytoplasmic kinases are key components of PRR-dependent signaling pathways. In tomato two such kinases, Pti1a and Pti1b, are important positive regulators of the plant immune response. However, it is unknown how these kinases control plant immunity at the molecular level, and how their activity is regulated. To investigate these issues, we used mass spectrometry to search for interactors of Pti1b in Nicotiana benthamiana leaves and identified a protein phosphatase, PP2C6. An in vitro pull-down assay and in vivo split luciferase complementation assay verified this interaction. Pti1b was found to autophosphorylate on threonine-233 and this phosphorylation was abolished in the presence of PP2C6. An arginine-to-cysteine substitution at position 240 in the Arabidopsis MARIS kinase was previously reported to convert it into a constitutive-active form. The analogous substitution in Pti1b made it resistant to PP2C6 phosphatase activity, although it still interacted with PP2C6. Treatment of N. benthamiana leaves with the MAMP flg22 induced threonine phosphorylation of Pti1b. Expression of PP2C6, but not a phosphatase-inactive variant of this protein, in N. benthamiana leaves greatly reduced ROS production in response to treatment with MAMPs flg22 or csp22. The results indicate that PP2C6 acts as a negative regulator by dephosphorylating the Pti1b kinase, thereby interfering with its ability to activate plant immune responses.

plant biology↗