Roadmap for the next decade of plant programmed cell death research.

Programmed cell death (PCD) is fundamentally important for plant development, abiotic stress responses and immunity, but our understanding of its regulation remains fragmented. Building a stronger research community is required to accelerate progress in this area through knowledge exchange and constructive debate. In this Viewpoint, we aim to initiate a collective effort to integrate data across a diverse set of experimental models to facilitate characterisation of the fundamental mechanisms underlying plant PCD and ultimately aid the development of a new plant cell death classification system in the future. We also put forward our vision for the next decade of plant PCD research stemming from discussions held during the 31st New Phytologist workshop, 'The Life and Death Decisions of Plant Cells' that took place at University College Dublin in Ireland (14-15 June 2023). We convey the key areas of significant progress and possible future research directions identified, including resolving the spatiotemporal control of cell death, isolation of its molecular and genetic regulators, and harnessing technical advances for studying PCD events in plants. Further, we review the breadth of potential impacts of plant PCD research and highlight the promising new applications of findings from this dynamically evolving field.

Transposable elements cause the loss of self-incompatibility in citrus.

Self-incompatibility (SI) is a widespread prezygotic mechanism for flowering plants to avoid inbreeding depression and promote genetic diversity. Citrus has an S-RNase-based SI system, which was frequently lost during evolution. We previously identified a single nucleotide mutation in Sm-RNase, which is responsible for the loss of SI in mandarin and its hybrids. However, little is known about other mechanisms responsible for conversion of SI to self-compatibility (SC) and we identify a completely different mechanism widely utilized by citrus. Here, we found a 786-bp miniature inverted-repeat transposable element (MITE) insertion in the promoter region of the FhiS2-RNase in Fortunella hindsii Swingle (a model plant for citrus gene function), which does not contain the Sm-RNase allele but are still SC. We demonstrate that this MITE plays a pivotal role in the loss of SI in citrus, providing evidence that this MITE insertion prevents expression of the S-RNase; moreover, transgenic experiments show that deletion of this 786-bp MITE insertion recovers the expression of FhiS2-RNase and restores SI. This study identifies the first evidence for a role for MITEs at the S-locus affecting the SI phenotype. A family-wide survey of the S-locus revealed that MITE insertions occur frequently adjacent to S-RNase alleles in different citrus genera, but only certain MITEs appear to be responsible for the loss of SI. Our study provides evidence that insertion of MITEs into a promoter region can alter a breeding strategy and suggests that this phenomenon may be broadly responsible for SC in species with the S-RNase system.

Variability of cell wall recalcitrance and composition in genotypes of Miscanthus from different genetic groups and geographical origin.

Miscanthus is a promising crop for bioenergy and biorefining in Europe. The improvement of Miscanthus as a crop relies on the creation of new varieties through the hybridization of germplasm collected in the wild with genetic variation and suitable characteristics in terms of resilience, yield and quality of the biomass. Local adaptation has likely shaped genetic variation for these characteristics and is therefore important to quantify. A key biomass quality parameter for biorefining is the ease of conversion of cell wall polysaccharides to monomeric sugars. Thus far, the variability of cell wall related traits in Miscanthus has mostly been explored in accessions from limited genetic backgrounds. Here we analysed the soil and climatic conditions of the original collection sites of 592 Miscanthus genotypes, which form eight distinct genetic groups based on discriminant analysis of principal components of 25,014 single-nucleotide polymorphisms. Our results show that species of the genus Miscanthus grow naturally across a range of soil and climate conditions. Based on a detailed analysis of 49 representative genotypes, we report generally minor differences in cell wall characteristics between different genetic groups and high levels of genetic variation within groups, with less investigated species like M. floridulus showing lower recalcitrance compared to the other genetic groups. The results emphasize that both inter- and intra- specific variation in cell wall characteristics and biomass recalcitrance can be used effectively in Miscanthus breeding programmes, while also reinforcing the importance of considering biomass yield when quantifying overall conversion efficiency. Thus, in addition to reflecting the complexity of the interactions between compositional and structural cell wall features and cell wall recalcitrance to sugar release, our results point to traits that could potentially require attention in breeding programmes targeted at improving the Miscanthus biomass crop.

Contrasting self-recognition rejection systems for self-incompatibility in Brassica and Papaver.

Self-incompatibility (SI) plays a pivotal role in whether self-pollen is accepted or rejected. Most SI systems employ two tightly linked loci encoding highly polymorphic pollen (male) and pistil (female) S-determinants that control whether self-pollination is successful or not. In recent years our knowledge of the signalling networks and cellular mechanisms involved has improved considerably, providing an important contribution to our understanding of the diverse mechanisms used by plant cells to recognise each other and elicit responses. Here, we compare and contrast two important SI systems employed in the Brassicaceae and Papaveraceae. Both use 'self-recognition' systems, but their genetic control and S-determinants are quite different. We describe the current knowledge about the receptors and ligands, and the downstream signals and responses utilized to prevent self-seed set. What emerges is a common theme involving the initiation of destructive pathways that block the key processes that are required for compatible pollen-pistil interactions.

Bacillus Calmette-Guérin vaccine for prevention of COVID-19 and other respiratory tract infections in older adults with comorbidities: a randomized controlled trial.

OBJECTIVES: To test whether Bacillus Calmette-Guérin (BCG) vaccination would reduce the incidence of COVID-19 and other respiratory tract infections (RTIs) in older adults with one or more comorbidities. METHODS: Community-dwelling adults aged 60 years or older with one or more underlying comorbidities and no contraindications to BCG vaccination were randomized 1:1 to BCG or placebo vaccination and followed for 6 months. The primary endpoint was a self-reported, test-confirmed COVID-19 incidence. Secondary endpoints included COVID-19 hospital admissions and clinically relevant RTIs (i.e. RTIs including but not limited to COVID-19 requiring medical intervention). COVID-19 and clinically relevant RTI episodes were adjudicated. Incidences were compared using Fine-Gray regression, accounting for competing events. RESULTS: A total of 6112 participants with a median age of 69 years (interquartile range, 65-74) and median of 2 (interquartile range, 1-3) comorbidities were randomized to BCG (n = 3058) or placebo (n = 3054) vaccination. COVID-19 infections were reported by 129 BCG recipients compared to 115 placebo recipients [hazard ratio (HR), 1.12; 95% CI, 0.87-1.44]. COVID-19-related hospitalization occurred in 18 BCG and 21 placebo recipients (HR, 0.86; 95% CI, 0.46-1.61). During the study period, 13 BCG recipients died compared with 18 placebo recipients (HR, 0.71; 95% CI, 0.35-1.43), of which 11 deaths (35%) were COVID-19-related: six in the placebo group and five in the BCG group. Clinically relevant RTI was reported by 66 BCG and 72 placebo recipients (HR, 0.92; 95% CI, 0.66-1.28). DISCUSSION: BCG vaccination does not protect older adults with comorbidities against COVID-19, COVID-19 hospitalization, or clinically relevant RTIs.

Transgenic ZmMYB167 Miscanthus sinensis with increased lignin to boost bioenergy generation for the bioeconomy.

BACKGROUND: Perennial C4 grasses from the genus Miscanthus are widely regarded as leading and promising dedicated bioenergy crops due to their high biomass accumulation on marginal land with low environmental impacts and maintenance requirements over its productive life. There is an urgent socio-political and environmental need to ramp up the production of alternative, affordable and green bioenergy sources and to re-direct the net zero carbon emissions trajectory. Hence, up-scaling of Miscanthus cultivation as a source of biomass for renewable energy could play an important role to strategically address sustainable development goals for a growing bio-based economy. Certain Miscanthus sinensis genotypes are particularly interesting for their biomass productivity across a wide range of locations. As the aromatic biomass component lignin exhibits a higher energy density than cell wall polysaccharides and is generally used as an indicator for heating or calorific value, genetic engineering could be a feasible strategy to develop M. sinensis biomass with increased lignin content and thus improving the energetic value of the biomass. RESULTS: For this purpose, transgenic M. sinensis were generated by Agrobacterium-mediated transformation for expression of ZmMYB167, a MYB transcription factor known for regulating lignin biosynthesis in C3 and C4 grasses. Four independent transgenic ZmMYB167 Miscanthus lines were obtained. Agronomic traits such as plant height, tillering and above-ground dry weight biomass of the transgenic plants were not different to that of wild-type control plants. Total lignin content of the transgenic plants was ~ 15-24% higher compared with control plants. However, the structural carbohydrates, glucan and xylan, were decreased by ~ 2-7% and ~ 8-10%, respectively, in the transgenic plants. Moreover, expression of ZmMYB167 in transgenic plants did not alter lignin composition, phenolic compounds or enzymatic saccharification efficiency yields but importantly improved total energy levels in Miscanthus biomass, equivalent to 10% higher energy yield per hectare. CONCLUSIONS: This study highlights ZmMYB167 as a suitable target for genetic lignin bioengineering interventions aimed at advancing and developing lignocellulosic biomass supply chains for sustainable production of renewable bioenergy.

Pollen Coat Proteomes of Arabidopsis thaliana, Arabidopsis lyrata, and Brassica oleracea Reveal Remarkable Diversity of Small Cysteine-Rich Proteins at the Pollen-Stigma Interface.

The pollen coat is the outermost domain of the pollen grain and is largely derived from the anther tapetum, which is a secretory tissue that degenerates late in pollen development. By being localised at the interface of the pollen-stigma interaction, the pollen coat plays a central role in mediating early pollination events, including molecular recognition. Amongst species of the Brassicaceae, a growing body of data has revealed that the pollen coat carries a range of proteins, with a number of small cysteine-rich proteins (CRPs) being identified as important regulators of the pollen-stigma interaction. By utilising a state-of-the-art liquid chromatography/tandem mass spectrometry (LC-MS/MS) approach, rich pollen coat proteomic profiles were obtained for Arabidopsis thaliana, Arabidopsis lyrata, and Brassica oleracea, which greatly extended previous datasets. All three proteomes revealed a strikingly large number of small CRPs that were not previously reported as pollen coat components. The profiling also uncovered a wide range of other protein families, many of which were enriched in the pollen coat proteomes and had functions associated with signal transduction, cell walls, lipid metabolism and defence. These proteomes provide an excellent source of molecular targets for future investigations into the pollen-stigma interaction and its potential evolutionary links to plant-pathogen interactions.

Valorisation Potential of Invasive Acacia dealbata, A. longifolia and A. melanoxylon from Land Clearings.

Acacia spp. are invasive in Southern Europe, and their high propagation rates produce excessive biomass, exacerbating wildfire risk. However, lignocellulosic biomass from Acacia spp. may be utilised for diverse biorefinery applications. In this study, attenuated total reflectance Fourier transform infrared spectroscopy (FTIR-ATR), high-performance anion-exchange chromatography pulsed amperometric detection (HPAEC-PAD) and lignin content determinations were used for a comparative compositional characterisation of A. dealbata, A. longifolia and A. melanoxylon. Additionally, biomass was treated with three white-rot fungi species (Ganoderma lucidum, Pleurotus ostreatus and Trametes versicolor), which preferentially degrade lignin. Our results showed that the pre-treatments do not significantly alter neutral sugar composition while reducing lignin content. Sugar release from enzymatic saccharification was enhanced, in some cases possibly due to a synergy between white-rot fungi and mild alkali pretreatments. For example, in A. dealbata stems treated with alkali and P. ostreatus, saccharification yield was 702.3 nmol mg-1, which is higher than the samples treated only with alkali (608.1 nmol mg-1), and 2.9-fold higher than the non-pretreated controls (243.9 nmol mg-1). By characterising biomass and pretreatments, generated data creates value for unused biomass resources, contributing to the implementation of sustainable biorefining systems. In due course, the generated value will lead to economic incentives for landowners to cut back invasive Acacia spp. more frequently, thus reducing excess biomass, which exacerbates wildfire risk.

Callus Induction from Diverse Explants and Genotypes Enables Robust Transformation of Perennial Ryegrass (Lolium perenne L.).

Genetic transformation of perennial ryegrass (Lolium perenne L.) is critical for fundamental and translational research in this important grass species. It often relies on Agrobacterium-mediated transformation of callus tissue. However, callus induction is restricted to a few genotypes that respond well to tissue culture. Here, we report callus induction from different perennial ryegrass genotypes and explants, such as shoot tips, seeds, and anthers, which were transformed with several plasmids for functional genomics. ß-glucuronidase (GUS) histochemical staining showed the LmdsRNAbp promoter sequence was active in stigmas, spikelets, anthers, and leaves. We also transformed calli with plasmids allowing gene silencing and gene knock-out using RNA interference and CRISPR/Cas9, respectively, for which genotypic and phenotypic investigations are ongoing. Using 19 different constructs, 262 transgenic events were regenerated. Moreover, the protocol regenerated a doubled haploid transgenic event from anther-derived calli. This work provides a proof-of-concept method for expanding the range of genotypes amenable to transformation, thus, serving research and breeding initiatives to improve this important grass crop for forage and recreation.

Depletion plays a pivotal role in self-incompatibility, revealing a link between cellular energy status, cytosolic acidification and actin remodelling in pollen tubes.

Self-incompatibility (SI) involves specific interactions during pollination to reject incompatible ('self') pollen, preventing inbreeding in angiosperms. A key event observed in pollen undergoing the Papaver rhoeas SI response is the formation of punctate F-actin foci. Pollen tube growth is heavily energy-dependent, yet ATP levels in pollen tubes have not been directly measured during SI. Here we used transgenic Arabidopsis lines expressing the Papaver pollen S-determinant to investigate a possible link between ATP levels, cytosolic pH ([pH]cyt ) and alterations to the actin cytoskeleton. We identify for the first time that SI triggers a rapid and significant ATP depletion in pollen tubes. Artificial depletion of ATP triggered cytosolic acidification and formation of actin aggregates. We also identify in vivo, evidence for a threshold [pH]cyt of 5.8 for actin foci formation. Imaging revealed that SI stimulates acidic cytosolic patches adjacent to the plasma membrane. In conclusion, this study provides evidence that ATP depletion plays a pivotal role in SI upstream of programmed cell death and reveals a link between the cellular energy status, cytosolic acidification and alterations to the actin cytoskeleton in regulating Papaver SI in pollen tubes.

Self-incompatibility requires GPI anchor remodeling by the poppy PGAP1 ortholog HLD1.

Glycosylphosphatidylinositol-anchored proteins (GPI-APs) are tethered to the outer leaflet of the plasma membrane where they function as key regulators of a plethora of biological processes in eukaryotes. Self-incompatibility (SI) plays a pivotal role regulating fertilization in higher plants through recognition and rejection of "self" pollen. Here, we used Arabidopsis thaliana lines that were engineered to be self-incompatible by expression of Papaver rhoeas SI determinants for an SI suppressor screen. We identify HLD1/AtPGAP1, an ortholog of the human GPI-inositol deacylase PGAP1, as a critical component required for the SI response. Besides a delay in flowering time, no developmental defects were observed in HLD1/AtPGAP1 knockout plants, but SI was completely abolished. We demonstrate that HLD1/AtPGAP1 functions as a GPI-inositol deacylase and that this GPI-remodeling activity is essential for SI. Using GFP-SKU5 as a representative GPI-AP, we show that the HLD1/AtPGAP1 mutation does not affect GPI-AP production and targeting but affects their cleavage and release from membranes in vivo. Our data not only implicate GPI-APs in SI, providing new directions to investigate SI mechanisms, but also identify a key functional role for GPI-AP remodeling by inositol deacylation in planta.

Clinical characteristics and risk factors of preventable hospital readmissions within 30 days.

Knowledge regarding preventable hospital readmissions is scarce. Our aim was to compare the clinical characteristics of potentially preventable readmissions (PPRs) with non-PPRs. Additionally, we aimed to identify risk factors for PPRs. Our study included readmissions within 30 days after discharge from 1 of 7 hospital departments. Preventability was assessed by multidisciplinary meetings. Characteristics of the readmissions were collected and 23 risk factors were analyzed. Of the 1120 readmissions, 125 (11%) were PPRs. PPRs occurred equally among different departments (p = 0.21). 29.6% of PPRs were readmitted by a practitioner of a different medical specialty than the initial admission (IA) specialist. The PPR group had more readmissions within 7 days (PPR 54% vs. non-PPR 44%, p = 0.03). The median LOS was 1 day longer for PPRs (p = 0.16). Factors associated with PPR were higher age (p = 0.004), higher socio-economic status (p = 0.049), fewer prior hospital admissions (p = 0.004), and no outpatient visit prior to readmission (p = 0.025). This study found that PPRs can occur at any department in the hospital. There is not a single type of patient that can easily be pinpointed to be at risk of a PPR, probably due to the multifactorial nature of PPRs.

Biorefining Potential of Wild-Grown Arundo donax, Cortaderia selloana and Phragmites australis and the Feasibility of White-Rot Fungi-Mediated Pretreatments.

Arundo donax, Cortaderia selloana and Phragmites australis are high-biomass-producing perennial Poalean species that grow abundantly and spontaneously in warm temperate regions, such as in Mediterranean-type climates, like those of Southern Europe, Western United States coastal areas, or in regions of South America, South Africa and Australia. Given their vigorous and spontaneous growth, biomass from the studied grasses often accumulates excessively in unmanaged agro-forestry areas. Nonetheless, this also creates the demand and opportunity for the valorisation of these biomass sources, particularly their cell wall polymers, for biorefining applications. By contrast, a related crop, Miscanthus × giganteus, is a perennial grass that has been extensively studied for lignocellulosic biomass production, as it can grow on low-input agricultural systems in colder climates. In this study Fourier transform mid-infrared spectroscopy (FTIR), high-performance anion-exchange chromatography (HPAEC) and lignin content determinations were used for a comparative compositional characterisation of A. donax, C. selloana and P. australis harvested from the wild, in relation to a trial field-grown M. × giganteus high-yielding genotype. A high-throughput saccharification assay showed relatively high sugar release values from the wild-grown grasses, even with a 0.1M NaOH mild alkali pretreatment. In addition to this alkaline pretreatment, biomass was treated with white-rot fungi (WRF), which preferentially degrade lignin more readily than holocellulose. Three fungal species were used: Ganoderma lucidum, Pleurotus ostreatus and Trametes versicolor. Our results showed that neutral sugar contents are not significantly altered, while some lignin is lost during the pretreatments. Furthermore, sugar release upon enzymatic saccharification was enhanced, and this was dependent on the plant biomass and fungal species used in the treatment. To maximise the potential for lignocellulose valorisation, the liquid fractions from the pretreatments were analysed by high performance liquid chromatography - photodiode array detection - electrospray ionisation tandem mass spectrometry (HPLC-PDA-ESI-MS n ). This study is one of the first to report on the composition of WRF-treated grass biomass, while assessing the potential relevance of breakdown products released during the treatments, beyond more traditional sugar-for-energy applications. Ultimately, we expect that our data will help promote the valorisation of unused biomass resources, create economic value, while contributing to the implementation of sustainable biorefining systems.

Plant biology: Stigmatic ROS decide whether pollen is accepted or rejected.

A new study reports that self-incompatibility in Brassica triggers the production of stigmatic ROS that are responsible for the rejection of incompatible pollen.

Mechanical stimulation in wheat triggers age- and dose-dependent alterations in growth, development and grain characteristics.

BACKGROUND AND AIMS: Wheat crops are exposed to a range of mechanical stimulations in their natural environment, yet we know very little about their response to such conditions. The aim of this study was to better understand the effect of mechanical stimulation on wheat growth and development, stem mechanical properties and grain measures. We focused on the following questions: (1) Does plant age affect the response to mechanical stimulation? (2) Is there a minimum threshold for the perception of mechanical stimuli? (3) Is the effect of manual brushing different to natural wind stimulation? METHODS: For age- and dose-response experiments, wheat plants were grown under controlled glasshouse conditions with brushing treatments applied using a purpose-built rig. The results of the controlled experiments are compared with those from an outside experiment where wheat plants were exposed to natural wind, with or without additional brushing. Detailed phenotypic measurements were conducted and treatment effects on grain characteristics were determined using micro-computed tomography imaging. KEY RESULTS: Two-week-old wheat plants were particularly sensitive to mechanical stimulation by controlled brushing treatments. Amongst others, plants exhibited a large reduction in height and grain yield, and an increase in tillers, above-ground biomass and stiffness of stem segments. Plants responded significantly to doses as small as one daily brushstroke. Outdoor experiments by and large confirmed results from controlled environment experiments. CONCLUSIONS: The morphological and developmental response to mechanical brushing treatment, in relation to vegetative above-ground biomass and grain yield, is dependent on plant age as well as the dose of the treatments. This study shows that mechanical stimulation of wheat impacts on a multitude of agriculturally relevant traits and provides a much needed advancement of our understanding of wheat thigmomorphogenesis and the potential applications of mechanical conditioning to control relevant traits.

A Comparison of Differential Gene Expression in Response to the Onset of Water Stress Between Three Hybrid Brachiaria Genotypes.

Brachiaria (Trin.) Griseb. (syn. Urochloa P. Beauv.) is a C4 grass genus belonging to the Panicoideae. Native to Africa, these grasses are now widely grown as forages in tropical areas worldwide and are the subject of intensive breeding, particularly in South America. Tolerance to abiotic stresses such as aluminum and drought are major breeding objectives. In this study, we present the transcriptomic profiling of leaves and roots of three Brachiaria interspecific hybrid genotypes with the onset of water stress, Br12/3659-17 (gt-17), Br12/2360-9 (gt-9), and Br12/3868-18 (gt-18), previously characterized as having good, intermediate and poor tolerance to drought, respectively, in germplasm evaluation programs. RNA was extracted from leaf and root tissue of plants at estimated growing medium water contents (EWC) of 35, 15, and 5%. Differentially expressed genes (DEGs) were compared between different EWCs, 35/15, 15/5, and 35/5 using DESeq2. Overall, the proportions of DEGs enriched in all three genotypes varied in a genotype-dependent manner in relation to EWC comparison, with intermediate and sensitive gt-9 and gt-18 being more similar to each other than to drought tolerant gt-17. More specifically, GO terms relating to carbohydrate and cell wall metabolism in the leaves were enriched by up-regulated DEGs in gt-9 and gt-18, but by down-regulated DEGs in gt-17. Across all genotypes, analysis of DEG enzyme activities indicated an excess of down-regulated putative apoplastic peroxidases in the roots as water stress increased. This suggests that changes in root cell-wall architecture may be an important component of the response to water stress in Brachiaria.

Supplemental Breast MRI for Women with Extremely Dense Breasts: Results of the Second Screening Round of the DENSE Trial.

Background In the first (prevalent) supplemental MRI screening round of the Dense Tissue and Early Breast Neoplasm Screening (DENSE) trial, a considerable number of breast cancers were found at the cost of an increased false-positive rate (FPR). In incident screening rounds, a lower cancer detection rate (CDR) is expected due to a smaller pool of prevalent cancers, and a reduced FPR, due to the availability of prior MRI examinations. Purpose To investigate screening performance indicators of the second round (incidence round) of the DENSE trial. Materials and Methods The DENSE trial (ClinicalTrials.gov: NCT01315015) is embedded within the Dutch population-based biennial mammography screening program for women aged 50-75 years. MRI examinations were performed between December 2011 and January 2016. Women were eligible for the second round when they again had a negative screening mammogram 2 years after their first MRI. The recall rate, biopsy rate, CDR, FPR, positive predictive values, and distributions of tumor characteristics were calculated and compared with results of the first round using 95% CIs and χ2 tests. Results A total of 3436 women (median age, 56 years; interquartile range, 48-64 years) underwent a second MRI screening. The CDR was 5.8 per 1000 screening examinations (95% CI: 3.8, 9.0) compared with 16.5 per 1000 screening examinations (95% CI: 13.3, 20.5) in the first round. The FPR was 26.3 per 1000 screening examinations (95% CI: 21.5, 32.3) in the second round versus 79.8 per 1000 screening examinations (95% CI: 72.4, 87.9) in the first round. The positive predictive value for recall was 18% (20 of 110 participants recalled; 95% CI: 12.1, 26.4), and the positive predictive value for biopsy was 24% (20 of 84 participants who underwent biopsy; 95% CI: 16.0, 33.9), both comparable to that of the first round. All tumors in the second round were stage 0-I and node negative. Conclusion The incremental cancer detection rate in the second round was 5.8 per 1000 screening examinations-compared with 16.5 per 1000 screening examinations in the first round. This was accompanied by a strong reduction in the number of false-positive results. © RSNA, 2021 Online supplemental material is available for this article. See also the editorial by Moy and Gao in this issue.

Production of oligosaccharides and biofuels from Miscanthus using combinatorial steam explosion and ionic liquid pretreatment.

Pretreatment strategies are fundamental to effectively deconstruct lignocellulosic biomass and economically produce biofuels, biomaterials and bio-based chemicals. This study evaluated individual and combinatorial steam explosion (SE) and ionic liquid (IL) pretreatments for production of high-value oligosaccharides from a novel seed-based Miscanthus hybrid (Mx2779). The two ILs used for pretreatment were triethylammonium hydrogen sulphate [TEA][HSO4] and 1-ethyl-3-methylimidazolium acetate [C2mim][OAc]. The results showed that each pretreatment leads to distinct effects on the fragmentation (cellulose and xylan dissolution, delignification, deacetylation) and physicochemical modification (cellulose and lignin properties) of lignocellulose. This, in turn, dictated enzymatic hydrolysis efficiencies of the cellulose pulp to glucose or gluco-oligosaccharides for downstream applications. Our findings suggest that the stand-alone SE or [C2mim][OAc] pretreatments may offer cost advantages over [TEA][HSO4] through the production of oligosaccharides such as xylo- and gluco-oligosaccharides. This study also highlights technical and economic pretreatment process challenges related to the production of oligosaccharides from Miscanthus Mx2779 biomass.

Ectopic Expression of a Self-Incompatibility Module Triggers Growth Arrest and Cell Death in Vegetative Cells.

Self-incompatibility (SI) is used by many angiosperms to reject self-pollen and avoid inbreeding. In field poppy (Papaver rhoeas), SI recognition and rejection of self-pollen is facilitated by a female S-determinant, PrsS, and a male S-determinant, PrpS PrsS belongs to the cysteine-rich peptide family, whose members activate diverse signaling networks involved in plant growth, defense, and reproduction. PrsS and PrpS are tightly regulated and expressed solely in pistil and pollen cells, respectively. Interaction of cognate PrsS and PrpS triggers pollen tube growth inhibition and programmed cell death (PCD) of self-pollen. We previously demonstrated functional intergeneric transfer of PrpS and PrsS to Arabidopsis (Arabidopsis thaliana) pollen and pistil. Here, we show that PrpS and PrsS, when expressed ectopically, act as a bipartite module to trigger a self-recognition:self-destruct response in Arabidopsis independently of its reproductive context in vegetative cells. The addition of recombinant PrsS to seedling roots expressing the cognate PrpS resulted in hallmark features of the P rhoeas SI response, including S-specific growth inhibition and PCD of root cells. Moreover, inducible expression of PrsS in PrpS-expressing seedlings resulted in rapid death of the entire seedling. This demonstrates that, besides specifying SI, the bipartite PrpS-PrsS module can trigger growth arrest and cell death in vegetative cells. Heterologous, ectopic expression of a plant bipartite signaling module in plants has not been shown previously and, by extrapolation, our findings suggest that cysteine-rich peptides diversified for a variety of specialized functions, including the regulation of growth and PCD.