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During photosynthesis, reactive oxygen species (ROS) are formed, including hydrogen peroxide (H 2 O 2 ) and singlet oxygen ( 1 O 2 ), which have putative roles in signalling, but their involvement in photosynthetic acclimation is unclear. Due to extreme reactivity and a short lifetime, 1 O 2 signalling occurs via its reaction products, such as oxidised poly-unsaturated fatty acids in thylakoid membranes. The resulting lipid peroxides decay to various aldehydes and reactive electrophile species (RES). Here, we investigated the role of ROS in the signal transduction of high light (HL), focusing on GreenCut2 genes unique to photosynthetic organisms. Using RNA seq. data, the transcriptional responses of Chlamydomonas reinhardtii to 2 h HL were compared with responses under low light to exogenous RES (acrolein; 4-hydroxynonenal), β-cyclocitral, a β-carotene oxidation product, as well as Rose Bengal, a 1 O 2 -producing photosensitiser, and H 2 O 2 . HL induced significant ( p < 0.05) up- and down-regulation of 108 and 23 GreenCut2 genes, respectively. Of all HL up-regulated genes, over half were also up-regulated by RES, including RBCS1 (ribulose bisphosphate carboxylase small subunit), NPQ-related PSBS1 and LHCSR1 . Furthermore, 96% of the genes down-regulated by HL were also down-regulated by 1 O 2 or RES, including CAO1 (chlorophyllide- a oxygnease), MDH2 (NADP-malate dehydrogenase) and PGM4 (phosphoglycerate mutase) for glycolysis. In comparison, only 0-4% of HL-affected GreenCut2 genes were similarly affected by H 2 O 2 or β-cyclocitral. Overall, 1 O 2 plays a significant role in signalling during the initial acclimation of C. reinhardtii to HL by up-regulating photo-protection and carbon assimilation and down-regulating specific primary metabolic pathways. Our data support that this pathway involves RES.

Antimicrobial photodynamic therapy (aPDT) is an evolving treatment strategy against human pathogenic microbes such as the Candida species, including the emerging pathogen C. auris . Using a modified EUCAST protocol, the light-enhanced antifungal activity of the natural compound parietin was explored. The photoactivity was evaluated against three separate strains of five yeasts, and its molecular mode of action was analysed via several techniques, i.e., cellular uptake, reactive electrophilic species (RES), and singlet oxygen yield. Under experimental conditions ( λ  = 428 nm, H = 30 J/cm 2 , PI = 30 min), microbial growth was inhibited by more than 90% at parietin concentrations as low as c = 0.156 mg/L (0.55 µM) for C. tropicalis and Cryptococcus neoformans , c = 0.313 mg/L (1.10 µM) for C. auris , c = 0.625 mg/L (2.20 µM) for C. glabrata , and c = 1.250 mg/L (4.40 µM) for C. albicans . Mode-of-action analysis demonstrated fungicidal activity. Parietin targets the cell membrane and induces cell death via ROS-mediated lipid peroxidation after light irradiation. In summary, parietin exhibits light-enhanced fungicidal activity against all Candida species tested (including C. auris ) and Cryptococcus neoformans , covering three of the four critical threats on the WHO's most recent fungal priority list., Competing Interests: The authors declare that they have no conflict of interest., (The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution-NonDerivative-NonCommercial License, permitting copying and reproduction so long as the original work is given appropriate credit. Contents may not be used for commecial purposes, or adapted, remixed, transformed or built upon. (https://creativecommons.org/licenses/by-nc-nd/4.0/).)

Radiation-associated sarcomas (RASs) are rare tumors with limited contemporary data to inform prognostication and management. We sought to identify the clinical presentation, patterns of care, and prognostic factors of RASs. RAS patients treated at a single institution from 2015 to 2021 were retrospectively reviewed for clinicopathologic variables, treatment strategies, and outcomes. Thirty-eight patients were identified with a median follow-up of 30.5 months. The median age at RAS diagnosis was 68.4 years (27.9-85.4), with a median latency from index radiotherapy (RT) of 9.1 years (3.7-46.3). RAS histologies included angiosarcoma (26%), undifferentiated pleomorphic sarcoma (21%), and osteosarcoma (18%). Most were high-grade (76%). Genomic profiling revealed low tumor mutational burden, frequent inactivating TP53 mutations (44%), CDKN2A deletions (26%), and MYC amplifications (22%), particularly in breast angiosarcomas. Of 38 patients, 33 presented with localized disease, 26 of whom were treated with curative intent. Overall, the median progression-free survival (PFS) was 9.5 months (1.4-34.7), and the overall survival (OS) was 11.1 months (0.6-31.6). Patients with localized vs. metastatic RASs had a longer PFS (HR, 3.0 [1.1-8.5]; p = 0.03) and OS (HR, 3.0 [1.04-8.68]; p = 0.03). Among localized RAS patients, high grade was associated with shorter OS (HR, 4.6 [1.04-20.30]; p = 0.03) and resection with longer OS (mean 58.8 vs. 6.1 months, HR, 0.1 [0.03-0.28]; p < 0.001). Among patients undergoing resection, negative margins were associated with improved OS (mean 71.0 vs. 15.5 months, HR, 5.1 [1.4-18.2]; p = 0.006). Patients with localized disease, particularly those undergoing R0 resection, demonstrated significantly better outcomes. Novel strategies are urgently needed to improve treatment outcomes in this challenging group of diseases.

The maintenance of regulatory T (T reg ) cells critically prevents autoimmunity. Pre-B cell leukemia transcription factor 1 ( Pbx1 ) variants are associated with lupus susceptibility, particularly through the expression of a dominant negative isoform Pbx1-d in CD4 + T cells. Pbx1-d overexpression impaired T reg cell homeostasis and promoted inflammatory CD4 + T cells. Here, we showed a high expression of Pbx1 in human and murine T reg cells, which is decreased in lupus patients and mice. Pbx1 deficiency or Pbx1-d overexpression reduced the number, stability, and suppressive activity of T reg cells, which increased murine responses to immunization and autoimmune induction. Mechanistically, Pbx1 deficiency altered the expression of genes implicated in cell cycle and apoptosis in T reg cells. Intriguingly, Rtkn2 , a Rho-GTPase previously associated with T reg homeostasis, was directly transactivated by Pbx1. Our results suggest that the maintenance of T reg cell homeostasis and stability by Pbx1 through cell cycle progression prevent the expansion of inflammatory T cells that otherwise exacerbates lupus progression in the hosts.

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease in which poorly characterized genetic factors lead to the production of proinflammatory or autoreactive T cells. Pre-B cell leukemia homeobox 1 (PBX1) is a transcription factor whose dominant negative isoform (PBX1-D) is overexpressed in the CD4 + T cells of SLE patients and lupus-prone mice. Pbx1-D overexpression favors the expansion of proinflammatory T cells and impairs regulatory T (Treg) cell development. Here we show that Pbx1 deficiency and Pbx1-D overexpression decreased STAT3 expression and activation in T cells. Accordingly, Pbx1 deficiency in T cells and Pbx1-D overexpression reduced STAT3-dependent T H 17 cell polarization in vitro, but it had no effect in vivo at steady state. STAT3-dependent follicular helper T (T FH ) cell polarization in vitro and splenic T FH cell frequency were not affected by either Pbx1 deficiency or Pbx1-D overexpression. Pbx1 deficiency also increased the expression of cell cycle arrest and pro-apoptotic genes, with an increased apoptosis in T cells. Our results suggest a complex interplay between PBX1 and STAT3, which may contribute to lupus pathogenesis through dysregulation of the cell cycle and apoptosis., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Identifying traits that exhibit improved drought resistance is highly important to cope with the challenges of predicted climate change. We investigated the response of state transition mutants to drought. Compared with the wild type, state transition mutants were less affected by drought. Photosynthetic parameters in leaves probed by chlorophyll fluorescence confirmed that mutants possess a more reduced plastoquinone (PQ) pool, as expected due to the absence of state transitions. Seedlings of the mutants showed an enhanced growth of the primary root and more lateral root formation. The photosystem II inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea, leading to an oxidised PQ pool, inhibited primary root growth in wild type and mutants, while the cytochrome b 6 f complex inhibitor 2,5-dibromo-3-methyl-6-isopropylbenzoquinone, leading to a reduced PQ pool, stimulated root growth. A more reduced state of the PQ pool was associated with a slight but significant increase in singlet oxygen production. Singlet oxygen may trigger a, yet unknown, signalling cascade promoting root growth. We propose that photosynthetic mutants with a deregulated ratio of photosystem II to photosystem I activity can provide a novel path for improving crop drought resistance., (© 2023 The Authors. Plant, Cell & Environment published by John Wiley & Sons Ltd.)

Pre-B cell leukemia homeobox 1 (PBX1) controls chromatin accessibility to a large number of genes in various cell types. Its dominant negative splice isoform, PBX1D, which lacks the DNA and Hox-binding domains, is expressed more frequently in the CD4+ T cells from lupus-prone mice and patients with systemic lupus erythematosus than healthy control subjects. PBX1D overexpression in CD4+ T cells impaired regulatory T cell homeostasis and expanded inflammatory CD4+ T cells. In this study, we showed that PBX1 message expression is downregulated by activation in CD4+ T cells as well as in B cells. PBX1D protein was less stable than the normal isoform, PBX1B, and it is degraded through the ubiquitin-proteasome-dependent pathway. The DNA binding domain lacking in PBX1D has two putative ubiquitin binding sites, K292 and K293, that are predicted to be in direct contact with DNA. Mutation of K292-293 reduced PBX1B stability to a level similar to PBX1D and abrogated DNA binding. In addition, contrary to PBX1B, PBX1D is retained in the cytoplasm without the help of the cofactors MEIS or PREP1, indicating a different requirement for nuclear translocation. Overall, these findings suggest that multiple post-transcriptional mechanisms are responsible for PBX1D loss of function and induction of CD4+ T cell inflammatory phenotypes in systemic lupus erythematosus., (Copyright © 2023 by The American Association of Immunologists, Inc.)

Fungal infections adversely influence the production and quality of seeds. Previously, Fusarium solani was reported as the causal agent of roselle ( Hibiscus sabdariffa L.) seed rot. This study was designed to evaluate the effect of F. solani infection on the germination, biochemical composition, energy reserves, and antioxidant activity of roselle seeds because there is currently a lack of information on the relationship between seed metabolism and infection with F. solani . The results showed that roselle seeds infected with F. solani exhibited a ca. 55% reduction in overall germination. Additionally, the fungal infection decreased antioxidant activity, total phenolic content, protein, sugar (sucrose, fructose, and glucose), and some amino acid (glutamine, serine, and arginine) contents. In contrast, some metabolites were more abundant in infected seeds, including alanine (2.1-fold) and some fatty acids (palmitic acid and heptadecanoic acid by 1.1- and 1.4-fold, respectively). The infection-associated changes in fatty acid profile resulted in the ratio of unsaturated/saturated fatty acids being 2.1-fold higher in infected seeds. Therefore, our results reveal that F. solani infection remarkably altered the biochemical composition of roselle seeds, which may have contributed to the loss of germination and quality of roselle seeds., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Tahmasebi, Roach, Shin and Lee.)

Heat and drought stresses are increasingly relevant topics in the context of climate change, particularly in the Alps, which are warming faster than the global average. Previously, we have shown that alpine plants, including Primula minima , can be gradually heat hardened under field conditions in situ to achieve maximum tolerance within a week. Here, we investigated the antioxidant mechanisms of P. minima leaves that had been heat hardened (H) without or with (H+D) additional drought stress. Lower free-radical scavenging and ascorbate concentrations were found in H and H+D leaves, while concentrations of glutathione disulphide (GSSG) were higher under both treatments without any change in glutathione (GSH) and little change in glutathione reductase activity. In contrast, ascorbate peroxidase activity in H leaves was increased, and H+D leaves had >two-fold higher catalase, ascorbate peroxidase and glucose-6-phosphate dehydrogenase activities compared with the control. In addition, the glutathione reductase activity was higher in H+D compared with H leaves. Our results highlight that the stress load from heat acclimation to maximum tolerance is associated with a weakened low-molecular-weight antioxidant defence, which may be compensated for by an increased activity of antioxidant enzymes, particularly under drought conditions.

The photosynthetic apparatus of lichen photobionts has been well-characterized by chlorophyll fluorescence analysis (e.g., by pulse amplitude modulation [PAM]), which provides a proxy of the activity of photosystem II (PSII) and its antenna. However, such kinetics are unable to directly characterize photosystem I (PSI) activity and the associated alternative electron pathways that may be involved in photoprotection. Instead, PSI can be probed in vivo by near-infrared absorption, measured at the same time as standard chlorophyll fluorescence (e.g., using the WALZ Dual PAM). Here, we used the Dual PAM to investigate cyclic electron flow and photoprotection in a range of mostly temperate lichens sampled from shaded to more open microhabitats. Sun species displayed lower acceptor side limitation of PSI (Y[NA]) early in illumination when compared to shade species, indicative of higher flavodiiron-mediated pseudocyclic electron flow. In response to high irradiance, some lichens accumulate melanin, and Y[NA] was lower and NAD(P)H dehydrogenase (NDH-2)-type cyclic flow was higher in melanised than pale forms. Furthermore, non-photochemical quenching (NPQ) was higher and faster relaxing in shade than sun species, while all lichens displayed high rates of photosynthetic cyclic electron flow. In conclusion, our data suggest that (1) low acceptor side limitation of PSI is important for sun-exposed lichens; (2) NPQ helps shade species tolerate brief exposure to high irradiance; and (3) cyclic electron flow is a prominent feature of lichens regardless of habitat, although NDH-2-type flow is associated with high light acclimation., (© 2023 The Authors. Physiologia Plantarum published by John Wiley & Sons Ltd on behalf of Scandinavian Plant Physiology Society.)

High seed quality is key to agricultural production, which is increasingly affected by climate change. We studied the effects of drought and elevated temperature during seed production on key seed quality traits of two genotypes of malting barley ( Hordeum sativum L.). Plants of a "Hana-type" landrace (B1) were taller, flowered earlier and produced heavier, larger and more vigorous seeds that resisted ageing longer compared to a semi-dwarf breeding line (B2). Accordingly, a NAC domain-containing transcription factor (TF) associated with rapid response to environmental stimuli, and the TF ABI5, a key regulator of seed dormancy and vigour, were more abundant in B1 seeds. Drought significantly reduced seed yield in both genotypes, and elevated temperature reduced seed size. Genotype B2 showed partial thermodormancy that was alleviated by drought and elevated temperature. Metabolite profiling revealed clear differences between the embryos of B1 and B2. Drought, but not elevated temperature, affected the metabolism of amino acids, organic acids, osmolytes and nitrogen assimilation, in the seeds of both genotypes. Our study may support future breeding efforts to produce new lodging and drought resistant malting barleys without trade-offs that can occur in semi-dwarf varieties such as lower stress resistance and higher dormancy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Nagel, Arc, Rajjou, Cueff, Bailly, Clément, Sanchez-Vicente, Bailly, Seal, Roach, Rolletschek, Lorenzo, Börner and Kranner.)

Systemic lupus erythematosus (SLE) is an autoimmune disease in which the overactivation of the immune system has been associated with metabolic alterations. Targeting the altered immunometabolism has been proposed to treat SLE patients based on their results obtained and mouse models of the disease. Here, we review the recent literature to discuss the possible origins of the alterations in the metabolism of immune cells in lupus, the dominant role of mitochondrial defects, technological advances that may move the field forward, as well as how targeting lupus immunometabolism may have therapeutic potential., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Senescence in plants enables resource recycling from senescent leaves to sink organs. Under stress, increased production of reactive oxygen species (ROS) and associated signalling activates senescence. However, senescence is not always associated with stress since it has a prominent role in plant development, in which the role of ROS signalling is less clear. To address this, we investigated lipid metabolism and patterns of lipid peroxidation related to signalling during sequential senescence in first-emerging barley leaves grown under natural light conditions. Leaf fatty acid compositions were dominated by linolenic acid (75% of total), the major polyunsaturated fatty acid (PUFA) in galactolipids of thylakoid membranes, known to be highly sensitive to peroxidation. Lipid catabolism during senescence, including increased lipoxygenase activity, led to decreased levels of PUFA and increased levels of short-chain saturated fatty acids. When normalised to leaf area, only concentrations of hexanal, a product from the 13-lipoxygenase pathway, increased early upon senescence, whereas reactive electrophile species (RES) from ROS-associated lipid peroxidation, such as 4-hydroxynonenal, 4-hydroxyhexenal and acrolein, as well as β-cyclocitral derived from oxidation of β-carotene, decreased. However, relative to total chlorophyll, amounts of most RES increased at late-senescence stages, alongside increased levels of α-tocopherol, zeaxanthin and non-photochemical quenching, an energy dissipative pathway that prevents ROS production. Overall, our results indicate that lipid peroxidation derived from enzymatic oxidation occurs early during senescence in first barley leaves, while ROS-derived lipid peroxidation associates weaker with senescence., (© 2022 The Authors. Physiologia Plantarum published by John Wiley & Sons Ltd on behalf of Scandinavian Plant Physiology Society.)

Understanding the genetic factors involved in seed longevity is of paramount importance in agricultural and ecological contexts. The polygenic nature of this trait suggests that many of them remain undiscovered. Here, we exploited the contrasting seed longevity found amongst Arabidopsis thaliana accessions to further understand this phenomenon. Concentrations of glutathione were higher in longer-lived than shorter-lived accessions, supporting that redox poise plays a prominent role in seed longevity. However, high seed permeability, normally associated with shorter longevity, is also present in long-lived accessions. Dry seed transcriptome analysis indicated that the contribution to longevity of stored messenger RNA (mRNAs) is complex, including mainly accession-specific mechanisms. The detrimental effect on longevity caused by other factors may be counterbalanced by higher levels of specific mRNAs stored in dry seeds, for instance those of heat-shock proteins. Indeed, loss-of-function mutant analysis demonstrated that heat-shock factors HSF1A and 1B contributed to longevity. Furthermore, mutants of the stress-granule zinc-finger protein TZF9 or the spliceosome subunits MOS4 or MAC3A/MAC3B, extended seed longevity, positioning RNA as a novel player in the regulation of seed viability. mRNAs of proteins with putative relevance to longevity were also abundant in shorter-lived accessions, reinforcing the idea that resistance to ageing is determined by multiple factors., (© 2022 The Authors. Plant, Cell & Environment published by John Wiley & Sons Ltd.)

Seed germination critically determines successful plant establishment and agricultural productivity. In the plant holobiont's life cycle, seeds are hubs for microbial communities' assembly, but what exactly shapes the holobiont during germination remains unknown. Here, 16S rRNA gene amplicon sequencing characterized the bacterial communities in embryonic compartments (cotyledons and axes) and on seed coats pre- and post-germination of four soybean ( Glycine max ) cultivars, in the presence or absence of exogenous abscisic acid (ABA), which prevented germination and associated metabolism of seeds that had imbibed. Embryonic compartments were metabolically profiled during germination to design minimal media mimicking the seed endosphere for bacterial growth assays. The distinction between embryonic and seed coat bacterial microbiomes of dry seeds weakened during germination, resulting in the plumule, radicle, cotyledon, and seed coat all hosting the same most abundant and structurally influential genera in germinated seeds of every cultivar. Treatment with ABA prevented the increase of bacterial microbiomes' richness, but not taxonomic homogenization across seed compartments. Growth assays on minimal media containing the most abundant metabolites that accumulated in germinated seeds revealed that seed reserve mobilization promoted enrichment of copiotrophic bacteria. Our data show that seed imbibition enabled distribution of seed-coat-derived epiphytes into embryos irrespective of germination, while germinative metabolism promoted proliferation of copiotrophic taxa, which predominated in germinated seeds., Competing Interests: The author(s) declare no conflict of interest.

Seeds offer an internal microbial niche, termed the endosphere, colonized by communities of endophytic bacteria. To elucidate the functions of seed endophytes during germination and early plant growth, studies with culturable isolates are essential. Conventional growth media favor few fast-growing taxa, while micro organisms with restricted nutrient requirements are usually outcompeted prior to isolation. Consequently, current knowledge of the interaction between seeds and their endophytes remains limited to only few bacterial taxa, despite a "black box" of unculturable isolates colonizing the endosphere. Here, we designed various solid media to mimic the endosphere of germinating soybean (Glycine max L.) seeds and assessed their effect on the diversity of culturable endophytic bacteria. The embryonic axis (i.e., the future plant) possessed higher richness and harbored more unique genera (i.e., Brevundimonas , Methylobacterium , Microbacterium , Pseudoclavibacter , and Rathayibacter ) than cotyledons (i.e., seed storage organs). Overall, media containing germinating and ground seeds enabled culturing and isolation of the broadest diversity of endophytic bacteria, viewed through the molecular identification of 246 isolates. The use of multiple tailored media helped uncover trophic adaptation of the core taxa. Furthermore, comparison of seeds from four lots of distinct cultivars and origin revealed few overlapping taxa, indicating that the parental environment, including soil and fertilization regime, influenced seed endophytic diversity. Extended diversity of native seed endophytic bacteria revealed the functional relevance of unique Arthrobacter , Bacillus , and Curtobacterium strains to seed germination under salt stress, exemplifying the importance of enhanced culturing approaches to elucidate the role of microbiota in seed germination. IMPORTANCE Plant growth-promoting endophytic isolates that appear to advance seed germination are often obtained from plant niches other than the seed endosphere. Isolating pure cultures of native endophytes from seeds during germination is crucial to investigate their function during early plant growth. Here, the diversity of endophytic bacteria isolated from seeds during soybean germination was enhanced by combining media tailored to the nutritional composition of the seed endosphere, including pregerminated seeds themselves. Our results show that isolation from distinct soybean seed compartments affected such diversity, with the embryonic axis harboring more unique taxa while displaying higher endophytic richness. Furthermore, using pools of seeds from separate lots, each corresponding to a certain cultivar and field site, supported isolation of further unique strains that often unveiled substantial effects on germination performance. Such findings are relevant to assist studies on the interactions between seeds and their native endophytic bacteria.

Light stress signalling in algae and plants is partially orchestrated by singlet oxygen ( 1 O 2 ), a reactive oxygen species (ROS) that causes significant damage within the chloroplast, such as lipid peroxidation. In the vicinity of the photosystem II reaction centre, a major source of 1 O 2 , are two β-carotene molecules that quench 1 O 2 to ground-state oxygen. 1 O 2 can oxidise β-carotene to release β-cyclocitral, which has emerged as a 1 O 2 -mediated stress signal in the plant Arabidopsis thaliana . We investigated if β-cyclocitral can have similar retrograde signalling properties in the unicellular alga Chlamydomonas reinhardtii . Using RNA-Seq, we show that genes up-regulated in response to exogenous β-cyclocitral included CAROTENOID CLEAVAGE DIOXYGENASE 8 ( CCD8 ), while down-regulated genes included those associated with porphyrin and chlorophyll anabolism, such as tetrapyrrole-binding protein ( GUN4 ), magnesium chelatases ( CHLI1 , CHLI2 , CHLD , CHLH1 ), light-dependent protochlorophyllide reductase ( POR1 ), copper target 1 protein ( CTH1 ), and coproporphyrinogen III oxidase ( CPX1 ). Down-regulation of this pathway has also been shown in β-cyclocitral-treated A. thaliana , indicating conservation of this signalling mechanism in plants. However, in contrast to A. thaliana , a very limited overlap in differential gene expression was found in β-cyclocitral-treated and 1 O 2 -treated C. reinhardtii . Furthermore, exogenous treatment with β-cyclocitral did not induce tolerance to 1 O 2 . We conclude that while β-cyclocitral may down-regulate chlorophyll synthesis, it does not seem to contribute to 1 O 2 -mediated high light stress signalling in algae.

Immune homeostasis is a constant balancing act between effector T cells and regulatory T cells defined by Foxp3 expression, the transcription factor that drives their differentiation and immunosuppressive activity. Immune homeostasis is altered when Treg cells are not generated or maintained in sufficient numbers. Treg cells rendered unstable by loss of Foxp3 expression, known as ex-Treg cells, gain pro-inflammatory functions. Treg cells may also become dysfunctional and lose their suppressive capabilities. These alterations can cause an imbalance between effector and regulatory subsets, which may ultimately lead to autoimmunity. This review discusses recent studies that identified genetic factors that maintain Treg cell stability as well as preserve their suppressive function. We focus on studies associated with systemic lupus erythematosus and highlight their findings in the context of potential therapeutic gene targeting in Treg cells to reverse the phenotypic changes and functional dysregulation inducing autoimmunity., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Roach and Morel.)

During desiccation, the cytoplasm of orthodox seeds solidifies into an intracellular glass with highly restricted diffusion and molecular mobility. Temperature and water content govern seed ageing rates, while oxygen (O2) can promote deteriorative reactions. However, whether the cytoplasmic physical state affects involvement of O2 in seed ageing remains unresolved. We aged Pinus densiflora seeds by controlled deterioration (CD) at 45 °C and distinct relative humidity (RH), resulting in cells with a glassy (11% and 30% RH) or fluid (60% and 80% RH) cytoplasm. Hypoxic conditions (0.4% O2) during CD delayed seed deterioration, lipid peroxidation, and decline of antioxidants (glutathione, α-tocopherol, and γ-tocopherol), but only when the cytoplasm was glassy. In contrast, when the cytoplasm was fluid, seeds deteriorated at the same rate regardless of O2 availability, while being associated with limited lipid peroxidation, detoxification of lipid peroxide products, substantial loss of glutathione, and resumption of glutathione synthesis. Changes in metabolite profiles provided evidence of other O2-independent enzymatic reactions in a fluid cytoplasm, including aldo-keto reductase and glutamate decarboxylase activities. Biochemical profiles of seeds stored under seed bank conditions resembled those obtained after CD regimes that maintained a glassy cytoplasm. Overall, O2 contributed more to seed ageing when the cytoplasm was glassy, rather than fluid., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Objective: To identify radiomic and clinical features associated with post-ablation recurrence of AF, given that cardiac morphologic changes are associated with persistent atrial fibrillation (AF), and initiating triggers of AF often arise from the pulmonary veins which are targeted in ablation. Methods: Subjects with pre-ablation contrast CT scans prior to first-time catheter ablation for AF between 2014-2016 were retrospectively identified. A training dataset (D 1 ) was constructed from left atrial and pulmonary vein morphometric features extracted from equal numbers of consecutively included subjects with and without AF recurrence determined at 1 year. The top-performing combination of feature selection and classifier methods based on C-statistic was evaluated on a validation dataset (D 2 ), composed of subjects retrospectively identified between 2005-2010. Clinical models ([Formula: see text]) were similarly evaluated and compared to radiomic ([Formula: see text]) and radiomic-clinical models ([Formula: see text]), each independently validated on D 2 . Results: Of 150 subjects in D 1 , 108 received radiofrequency ablation and 42 received cryoballoon. Radiomic features of recurrence included greater right carina angle, reduced anterior-posterior atrial diameter, greater atrial volume normalized to height, and steeper right inferior pulmonary vein angle. Clinical features predicting recurrence included older age, greater BMI, hypertension, and warfarin use; apixaban use was associated with reduced recurrence. AF recurrence was predicted with radio-frequency ablation models on D 2 subjects with C-statistics of 0.68, 0.63, and 0.70 for radiomic, clinical, and combined feature models, though these were not prognostic in patients treated with cryoballoon. Conclusions: Pulmonary vein morphology associated with increased likelihood of AF recurrence within 1 year of catheter ablation was identified on cardiac CT. Significance: Radiomic and clinical features-based predictive models may assist in identifying atrial fibrillation ablation candidates with greatest likelihood of successful outcome.

Chitin, a major component of fungal cell walls, has been associated with allergic disorders such as asthma. However, it is unclear how mammals recognize chitin and the principal receptor(s) on epithelial cells that sense chitin remain to be determined. In this study, we show that LYSMD3 is expressed on the surface of human airway epithelial cells and demonstrate that LYSMD3 is able to bind chitin, as well as β-glucan, on the cell walls of fungi. Knockdown or knockout of LYSMD3 also sharply blunts the production of inflammatory cytokines by epithelial cells in response to chitin and fungal spores. Competitive inhibition of the LYSMD3 ectodomain by soluble LYSMD3 protein, multiple ligands, or antibody against LYSMD3 also blocks chitin signaling. Our study reveals LYSMD3 as a mammalian pattern recognition receptor (PRR) for chitin and establishes its role in epithelial cell inflammatory responses to chitin and fungi., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Cutin and suberin are lipid polyesters deposited in specific apoplastic compartments. Their fundamental roles in plant biology include controlling the movement of gases, water and solutes, and conferring pathogen resistance. Both cutin and suberin have been shown to be present in the Arabidopsis seed coat where they regulate seed dormancy and longevity. In this study, we use accelerated and natural ageing seed assays, glutathione redox potential measures, optical and transmission electron microscopy and gas chromatography-mass spectrometry to demonstrate that increasing the accumulation of lipid polyesters in the seed coat is the mechanism by which the AtHB25 transcription factor regulates seed permeability and longevity. Chromatin immunoprecipitation during seed maturation revealed that the lipid polyester biosynthetic gene long-chain acyl-CoA synthetase 2 (LACS2) is a direct AtHB25 binding target. Gene transfer of this transcription factor to wheat and tomato demonstrated the importance of apoplastic lipid polyesters for the maintenance of seed viability. Our work establishes AtHB25 as a trans-species regulator of seed longevity and has identified the deposition of apoplastic lipid barriers as a key parameter to improve seed longevity in multiple plant species., (© 2021 The Authors New Phytologist © 2021 New Phytologist Foundation.)

The interplay between the phytohormone abscisic acid (ABA) and the gasotransmitter nitric oxide (NO) regulates seed germination and post-germinative seedling growth. We show that GAP1 (germination in ABA and cPTIO 1) encodes the transcription factor ANAC089 with a critical membrane-bound domain and extranuclear localization. ANAC089 mutants lacking the membrane-tethered domain display insensitivity to ABA, salt, and osmotic and cold stresses, revealing a repressor function. Whole-genome transcriptional profiling and DNA-binding specificity reveals that ANAC089 regulates ABA- and redox-related genes. ANAC089 truncated mutants exhibit higher NO and lower ROS and ABA endogenous levels, alongside an altered thiol and disulfide homeostasis. Consistently, translocation of ANAC089 to the nucleus is directed by changes in cellular redox status after treatments with NO scavengers and redox-related compounds. Our results reveal ANAC089 to be a master regulator modulating redox homeostasis and NO levels, able to repress ABA synthesis and signaling during Arabidopsis seed germination and abiotic stress., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Xanthophyll cycles are broadly important in photoprotection, and the reversible de-epoxidation of xanthophylls typically occurs in excess light conditions. However, as presented in this review, compiling evidence in a wide range of photosynthetic eukaryotes shows that xanthophyll de-epoxidation also occurs under diverse abiotic stress conditions in darkness. Light-driven photochemistry usually leads to the pH changes that activate de-epoxidases (e.g. violaxanthin de-epoxidase), but in darkness alternative electron transport pathways and luminal domains enriched in monogalactosyl diacyl glycerol (which enhance de-epoxidase activity) likely enable de-epoxidation. Another 'dark side' to sustaining xanthophyll de-epoxidation is inactivation and/or degradation of epoxidases (e.g. zeaxanthin epoxidase). There are obvious benefits of such activity regarding stress tolerance, and indeed this phenomenon has only been reported in stressful conditions. However, more research is required to unravel the mechanisms and understand the physiological roles of dark-induced formation of zeaxanthin. Notably, the de-epoxidation of violaxanthin to antheraxanthin and zeaxanthin in darkness is still a frequently ignored process, perhaps because it questions a previous paradigm. With that in mind, this review seeks to shed some light on the dark side of xanthophyll de-epoxidation, and point out areas for future work., (© 2021 The Authors New Phytologist © 2021 New Phytologist Foundation.)

While most ferns avoid freezing as they have a tropical distribution or shed their fronds, wintergreen species in temperate and boreoalpine ecosystems have to deal with sub-zero temperatures. Increasing evidence has revealed overlapping mechanisms of desiccation and freezing tolerance in angiosperms, but the physiological mechanisms behind freezing tolerance in ferns are far from clear. We evaluated photochemical and hydraulic parameters in five wintergreen fern species differing in their ability to tolerate desiccation. We assessed frond freezing tolerance, ice nucleation temperature and propagation pattern, and xylem anatomical traits. Dynamics of photochemical performance and xanthophyll cycle were evaluated during freeze-thaw events under controlled conditions and, in selected species, in the field. Only desiccation-tolerant species, which possessed a greater fraction of narrow tracheids (<18 μm) than sensitive species, tolerated freezing. Frond freezing occurred in the field at -3.4 ± 0.9 °C (SD) irrespective of freezing tolerance, freezable water content, or tracheid properties. Even in complete darkness, maximal photochemical efficiency of photosystem II was down-regulated concomitantly with zeaxanthin accumulation in response to freezing. This was reversible upon re-warming only in tolerant species. Our results suggest that adaptation for freezing tolerance is associated with desiccation tolerance through complementary xylem properties (which may prevent risk of irreversible cavitation) and effective photoprotection mechanisms. The latter includes de-epoxidation of xanthophylls in darkness, a process evidenced for the first time directly in the field., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Autoimmune diseases are driven largely by a pathogenic cytokine milieu produced by aberrantly activated lymphocytes. Many cytokines, including interferon gamma (IFN-γ), utilize the JAK/STAT pathway for signal propagation. Suppressor of Cytokine Signaling-1 (SOCS1) is an inducible, intracellular protein that regulates IFN-γ signaling by dampening JAK/STAT signaling. Using Fas deficient, MRL/MpJ-Fas lpr /J (MRL/lpr) mice, which develop lupus-like disease spontaneously, we tested the hypothesis that a peptide mimic of the SOCS1 kinase inhibitory region (SOCS1-KIR) would inhibit lymphocyte activation and modulate lupus-associated pathologies. Consistent with in vitro studies, SOCS1-KIR intraperitoneal administration reduced the frequency, activation, and cytokine production of memory CD8 + and CD4 + T lymphocytes within the peripheral blood, spleen, and lymph nodes. In addition, SOCS1-KIR administration reduced lymphadenopathy, severity of skin lesions, autoantibody production, and modestly reduced kidney pathology. On a cellular level, peritoneal SOCS1-KIR administration enhanced Foxp3 expression in total splenic and follicular regulatory T cells, reduced the effector memory/naïve T lymphocyte ratio for both CD4 + and CD8 + cells, and reduced the frequency of GL7 + germinal center enriched B cells. Together, these data show that SOCS1-KIR treatment reduced auto-reactive lymphocyte effector functions and suggest that therapeutic targeting of the SOCS1 pathway through peptide administration may have efficacy in mitigating autoimmune pathologies.

Fumarylacetoacetate hydrolase (FAH) proteins form a superfamily found in Archaea, Bacteria, and Eukaryota. However, few fumarylacetoacetate hydrolase domain (FAHD)-containing proteins have been studied in Metazoa and their role in plants remains elusive. Sequence alignments revealed high homology between two Arabidopsis thaliana FAHD-containing proteins and human FAHD1 (hFAHD1) implicated in mitochondrial dysfunction-associated senescence. Transcripts of the closest hFAHD1 orthologue in Arabidopsis (AtFAHD1a) peak during seed maturation drying, which influences seed longevity and dormancy. Here, a homology study was conducted to assess if AtFAHD1a contributes to seed longevity and vigour. We found that an A. thaliana T-DNA insertional line ( Atfahd1a-1 ) had extended seed longevity and shallower thermo-dormancy. Compared to the wild type, metabolite profiling of dry Atfahd1a-1 seeds showed that the concentrations of several amino acids, some reducing monosaccharides, and δ-tocopherol dropped, whereas the concentrations of dehydroascorbate, its catabolic intermediate threonic acid, and ascorbate accumulated. Furthermore, the redox state of the glutathione disulphide/glutathione couple shifted towards a more reducing state in dry mature Atfahd1a-1 seeds, suggesting that AtFAHD1a affects antioxidant redox poise during seed development. In summary, AtFAHD1a appears to be involved in seed redox regulation and to affect seed quality traits such as seed thermo-dormancy and longevity.

Leaf senescence is an important process for plants to remobilize a variety of metabolites and nutrients to sink tissues, such as developing leaves, fruits and seeds. It has been suggested that reactive oxygen species (ROS) play an important role in the initiation of leaf senescence. Flag leaves of two different barley varieties, cv. Lomerit and cv. Carina, showed differences in the loss of photosystems and in the production of ROS at a late stage of senescence after significant loss of chlorophyll (Krieger-Liszkay et al. 2015). Here, we investigated photosynthetic electron transport and ROS production in primary leaves of these two varieties at earlier stages of senescence. Comparisons were made between plants grown outside in natural light and temperatures and plants grown in temperature-controlled growth chambers under low light intensity. Alterations in the content of photoactive P700, ferredoxin and plastocyanin (PC) photosynthetic electron transport were analyzed using in vivo near-infrared absorbance changes and chlorophyll fluorescence, while ROS were measured with spin-trapping electron paramagnetic resonance spectroscopy. Differences in ROS production between the two varieties were only observed in outdoor plants, whereas a loss of PC was common in both barley varieties regardless of growth conditions. We conclude that the loss of PC is the earliest detectable photosynthetic parameter of leaf senescence while differences in the production of individual ROS species occur later and depend on environmental factors., (© The Author(s) 2020. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Natural light intensities can rise several orders of magnitude over subsecond time spans, posing a major challenge for photosynthesis. Fluctuating light tolerance in the green alga Chlamydomonas reinhardtii requires alternative electron pathways, but the role of nonphotochemical quenching (NPQ) is not known. Here, fluctuating light (10 min actinic light followed by 10 min darkness) led to significant increase in NPQ/qE-related proteins, LHCSR1 and LHCSR3, relative to constant light of the same subsaturating or saturating intensity. Elevated levels of LHCSR1/3 increased the ability of cells to safely dissipate excess light energy to heat (i.e., qE-type NPQ) during dark to light transition, as measured with chlorophyll fluorescence. The low qE phenotype of the npq4 mutant, which is unable to produce LHCSR3, was abolished under fluctuating light, showing that LHCSR1 alone enables very high levels of qE. Photosystem (PS) levels were also affected by light treatments; constant light led to lower PsbA levels and F v / F m values, while fluctuating light led to lower PsaA and maximum P700 + levels, indicating that constant and fluctuating light induced PSII and PSI photoinhibition, respectively. Under fluctuating light, npq4 suffered more PSI photoinhibition and significantly slower growth rates than parental wild type, whereas npq1 and npq2 mutants affected in xanthophyll carotenoid compositions had identical growth under fluctuating and constant light. Overall, LHCSR3 rather than total qE capacity or zeaxanthin is shown to be important in C. reinhardtii in tolerating fluctuating light, potentially via preventing PSI photoinhibition.

Non-photochemical quenching (NPQ) helps dissipate surplus light energy, preventing formation of reactive oxygen species (ROS). In Chlamydomonas reinhardtii, the thylakoid membrane protein LHCSR3 is involved in pH-dependent (qE-type) NPQ, lacking in the npq4 mutant. Preventing PSII repair revealed that npq4 lost PSII activity faster than the wild type (WT) in elevated O2, while no difference between strains was observed in O2-depleted conditions. Low Fv/Fm values remained 1.5 h after moving cells out of high light, and this qH-type quenching was independent of LHCSR3 and not accompanied by losses of maximum PSII activity. Culturing cells in historic O2 atmospheres (30-35%) increased the qE of cells, due to increased LHCSR1 and PsbS levels, and LHCSR3 in the WT, showing that atmospheric O2 tensions regulate qE capacity. Colony growth of npq4 was severely restricted at elevated O2, and npq4 accumulated more reactive electrophile species (RES) than the WT, which could damage PSI. Levels of PsaA (PSI) were lower in npq4 grown at 35% O2, while PsbA (PSII) levels remained stable. We conclude that even at high O2 concentrations, the PSII repair cycle is sufficient to maintain net levels of PSII. However, LHCSR3 has an important function in protecting PSI against O2-mediated damage, such as via RES., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

In endophytes, the abundance of genes coding for enzymes processing reactive oxygen species (ROS), including hydrogen peroxide (H 2 O 2 ), argues for a crucial role of ROS metabolism in plant-microbe interaction for plant colonization. Here, we studied H 2 O 2 metabolism of bread wheat ( Triticum aestivum L.) seeds and their microbiota during germination and early seedling growth, the most vulnerable stages in the plant life cycle. Treatment with hot steam diminished the seed microbiota, and these seeds produced less extracellular H 2 O 2 than untreated seeds. Using a culture-dependent approach, Pantoea and Pseudomonas genera were the most abundant epiphytes of dry untreated seeds. Incubating intact seedlings from hot steam-treated seeds with Pantoea strains triggered H 2 O 2 production, whereas Pseudomonas strains dampened H 2 O 2 levels, attributable to higher catalase activities. The genus Pantoea was much less represented among seedling endophytes than genus Pseudomonas , with other endophytic genera, including Bacillus and Paenibacillus , also possessing high catalase activities. Overall, our results show that certain bacteria of the seed microbiota are able to modulate the extracellular redox environment during germination and early seedling growth, and high catalase activity is proposed as a key trait of seed endophytes.

High seed quality is a key trait to achieve successful crop establishment required for optimum yield and sustainable production. Seed storage conditions greatly impact two key seed quality traits; seed viability (ability to germinate and produce normal seedlings) and vigour (germination performance). Accumulated oxidative damage accompanies the loss of seed vigour and viability during ageing, indicating that redox control is key to longevity. Here, we studied the effects of controlled deterioration at 40°C and 75% relative humidity (RH) ('ageing') under two different O 2 concentrations (21 and 78% O 2 ) in Brassica oleracea Two B. oleracea genotypes with allelic differences at two QTLs that result in differences in abscisic acid (ABA) signalling and seed vigour were compared. Ageing led to a similar loss in germination speed in both genotypes that was lost faster under elevated O 2 In both genotypes, an equal oxidative shift in the glutathione redox state and a minor loss of α-tocopherol progressively occurred before seed viability was lost. In contrast, ABA levels were not affected by ageing. In conclusion, both ABA signalling and seed ageing impact seed vigour but not necessarily through the same biochemical mechanisms., (© 2019 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

Fast and uniform germination is key to agricultural production and can be achieved by seed 'priming' techniques. Here, we characterised the responses of bread wheat ( Triticum aestivum L.) seeds to a hot steam treatment ('BioFlash'), which accelerated water uptake, resulting in faster germination and seedling growth, typical traits of primed seed. Before the completion of germination, metabolite profiling of seeds revealed advanced accumulation of several amino acids (especially cysteine and serine), sugars (ribose, glucose), and organic acids (glycerate, succinate) in hot steam-treated seeds, whereas sugar alcohols (e.g. arabitol, mannitol) and trehalose decreased in all seeds. Tocochromanols (the 'vitamin E family') rose independently of the hot steam treatment. We further assessed shifts in the half-cell reduction potentials of low-molecular-weight (LMW) thiol-disulfide redox couples [i.e. glutathione disulfide (GSSG)/glutathione (GSH) and cystine/cysteine], alongside the activities of the reactive oxygen species (ROS)-processing enzyme superoxide dismutase, catalase, ascorbate peroxidase, and glutathione reductase. Upon the first 4 h of imbibition, a rapid conversion of LMW disulfides to thiols occurred. Completion of germination was associated with a re-oxidation of the LMW thiol-disulfide cellular redox environment, before more reducing conditions were re-established during seedling growth, accompanied by an increase in all ROS-processing enzyme activities. Furthermore, changes in the thiol-disulfide cellular redox state were associated to specific stages of wheat seed germination. In conclusion, the priming effect of the hot steam treatment advanced the onset of seed metabolism, including redox shifts associated with germination and seedling growth., (© 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

Photosynthetic organisms suffering from light stress have to cope with an increased formation of reactive short-chain aldehydes. Singlet oxygen generated from highly-charged reaction centres can peroxidise the poly-unsaturated fatty acid (PUFA)-rich thylakoid membranes they are embedded in. Lipid peroxides decay to release α,β-unsaturated aldehydes that are reactive electrophile species (RES). Acrolein is one of the most abundant and reactive RES produced in chloroplasts. Here, in the model chlorophyte alga Chlamydomonas reinhardtii, a clear concentration-dependent "distress" induced by acrolein intoxication was observed in conjunction with depletion of the glutathione pool. The glutathione redox state (E GSSG/2GSH ) strongly correlated (R 2 = 0.95) with decreasing F v /F m values of chlorophyll fluorescence. However, treatment of C. reinhardtii with sub-toxic acrolein concentrations increased glutathione concentrations and raised the protein levels of a glutathione-S-transferase (GSTS1), mimicking the response to excess light, indicating that at lower concentrations, acrolein may contribute to high light acclimation, which could be interpreted as "eustress". Furthermore, similar patterns of chloroplastic protein carbonylation occurred under light stress and in response to exogenous acrolein. Priming cells by low doses of acrolein increased the alga's resistance to singlet oxygen. A RNA seq. analysis showed a large overlap in gene regulation under singlet oxygen and acrolein stresses. Particularly enriched were transcripts of enzymes involved in thiol/disulphide exchanges. Some of the genes are regulated by the SOR1 transcription factor, but acrolein treatment still induced an increase in glutathione contents and enhanced singlet oxygen tolerance of the sor1 mutant. The results support a role for RES in chloroplast-to-nucleus retrograde signalling during high light acclimation, with involvement of SOR1 and other pathways., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: Loperamide is an over-the-counter, inexpensive, antidiarrheal opioid that can produce life-threatening toxicity at high concentrations. CASE REPORT 1: A 28-year-old man with a history of depression and substance abuse disorder (SUD) presented to the emergency department (ED) with shortness of breath and lightheadedness. He ingested large amounts of loperamide daily. The patient's initial electrocardiogram (ECG) demonstrated sinus rhythm, right axis deviation, undetectable PR interval, QRS 168 ms, and QTc 693 ms. He was administered intravenous sodium bicarbonate and magnesium sulfate and admitted to the intensive care unit, eventually developing Torsades de Pointes (TdP). He was given lidocaine and isoproterenol infusions, and an external pacemaker was placed. He was discharged in stable condition on hospital day (HD) 16. CASE REPORT 2: A 39-year-old woman with a history of hepatitis C, depression, and SUD was transported to the ED after reported seizure-like activity. The patient experienced TdP in the ED and admitted to ingesting large amount of loperamide daily. An ECG demonstrated sinus rhythm, right axis deviation, PR interval 208 ms, QRS interval 142 ms, and QTc 687 ms. She was administered intravenous magnesium, sodium bicarbonate, and isoproterenol. After intensive care unit admission, the patient experienced no further TdP and was discharged on HD 6. WHY SHOULD AN EMERGENCY PHYSICIAN BE AWARE OF THIS?: Emergency physicians should proceed with caution when treating patients with loperamide toxicity. Even in asymptomatic patients and drug discontinuance, obtain consultation with a medical toxicologist, promptly treat ECG abnormalities aggressively, and admit all patients for further monitoring., (Copyright © 2017 Elsevier Inc. All rights reserved.)

High light causes photosystem II to generate singlet oxygen ( 1 O 2 ), a reactive oxygen species (ROS) that can react with membrane lipids, releasing reactive electrophile species (RES), such as acrolein. To investigate how RES may contribute to light stress responses, Chlamydomonas reinhardtii was high light-treated in photoautotrophic and mixotrophic conditions and also in an oxygen-enriched atmosphere to elevate ROS production. The responses were compared to exogenous acrolein. Non-photochemical quenching (NPQ) was higher in photoautotrophic cells, as a consequence of a more de-epoxidized state of the xanthophyll cycle pool and more LHCSR3 protein, showing that photosynthesis was under more pressure than in mixotrophic cells. Photoautotrophic cells had lowered α-tocopherol and β-carotene contents and a higher level of protein carbonylation, indicators of elevated 1 O 2 production. Levels of glutathione, glutathione peroxidase (GPX5) and glutathione-S-transferase (GST1), important antioxidants against RES, were also increased in photoautotrophic cells. In parallel to the wild-type, the LHCSR3-deficient npq4 mutant was high light-treated, which in photoautotrophic conditions exhibited particular sensitivity under elevated oxygen, the treatment that induced the highest RES levels, including acrolein. The npq4 mutant had more GPX5 and GST1 alongside higher levels of carbonylated protein and a more oxidized glutathione redox state. In wild-type cells glutathione contents doubled after 4 h treatment, either with high light under elevated oxygen or with a non-critical dose (600 ppm) of acrolein. Exogenous acrolein also increased GST1 levels, but not GPX5. Overall, RES-associated oxidative damage and glutathione metabolism are prominently associated with light stress and potentially in signaling responses of C. reinhardtii., (© 2017 Scandinavian Plant Physiology Society.)

The tripeptide antioxidant glutathione (γ-l-glutamyl-l-cysteinyl-glycine; GSH) essentially contributes to thiol-disulphide conversions, which are involved in the control of seed development, germination, and seedling establishment. However, the relative contribution of GSH metabolism in different seed structures is not fully understood. We studied the GSH/glutathione disulphide (GSSG) redox couple and associated low-molecular-weight (LMW) thiols and disulphides related to GSH metabolism in bread wheat (Triticum aestivum L.) seeds, focussing on redox changes in the embryo and endosperm during germination. In dry seeds, GSH was the predominant LMW thiol and, 15 h after the onset of imbibition, embryos of non-germinated seeds contained 12 times more LMW thiols than the endosperm. In germinated seeds, the embryo contained 17 and 11 times more LMW thiols than the endosperm after 15 and 48 h, respectively. This resulted in the embryo having significantly more reducing half-cell reduction potentials of GSH/GSSG and cysteine (Cys)/cystine (CySS) redox couples (E GSSG/2GSH and E CySS/2Cys , respectively). Upon seed germination and early seedling growth, Cys and CySS concentrations significantly increased in both embryo and endosperm, progressively contributing to the cellular LMW thiol-disulphide redox environment (E thiol-disulphide ). The changes in E CySS/2Cys could be related to the mobilisation of storage proteins in the endosperm during early seedling growth. We suggest that E GSSG/2GSH and E CySS/2Cys can be used as markers of the physiological and developmental stage of embryo and endosperm. We also present a model of interaction between LMW thiols and disulphides with hydrogen peroxide (H 2 O 2 ) in redox regulation of bread wheat germination and early seedling growth.

Background: Ocular abnormalities present in microcephalic infants with presumed Zika virus (ZIKV) congenital disease includes focal pigment mottling of the retina, chorioretinal atrophy, optic nerve abnormalities, and lens dislocation. Target cells in the ocular compartment for ZIKV infectivity are unknown. The cellular response of ocular cells to ZIKV infection has not been described. Mechanisms for viral dissemination in the ocular compartment of ZIKV-infected infants and adults have not been reported. Here, we identify target cells for ZIKV infectivity in both the inner and outer blood-retinal barriers (IBRB and OBRB), describe the cytokine expression profile in the IBRB after ZIKV exposure, and propose a mechanism for viral dissemination in the retina., Methods: We expose primary cellular components of the IBRB including human retinal microvascular endothelial cells, retinal pericytes, and Müller cells as well as retinal pigmented epithelial cells of the OBRB to the PRVABC56 strain of ZIKV. Viral infectivity was analyzed by microscopy, immunofluorescence, and reverse transcription polymerase chain reaction (RT-PCR and qRT-PCR). Angiogenic and proinflammatory cytokines were measured by Luminex assays., Results: We find by immunofluorescent staining using the Flavivirus 4G2 monoclonal antibody that retinal endothelial cells and pericytes of the IBRB and retinal pigmented epithelial cells of the OBRB are fully permissive for ZIKV infection but not Müller cells when compared to mock-infected controls. We confirmed ZIKV infectivity in retinal endothelial cells, retinal pericytes, and retinal pigmented epithelial cells by RT-PCR and qRT-PCR using ZIKV-specific oligonucleotide primers. Expression profiles by Luminex assays in retinal endothelial cells infected with ZIKV revealed a marginal increase in levels of beta-2 microglobulin (β2-m), granulocyte macrophage colony-stimulating factor (GMCSF), intercellular adhesion molecule 1 (ICAM-1), interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP1), and vascular cell adhesion molecule 1 (VCAM-1) and higher levels of regulated upon activation, normal T cell expressed and presumably secreted (RANTES) but lower levels of interleukin-4 (IL-4) compared to controls., Conclusions: Retinal endothelial cells, retinal pericytes, and retinal pigmented epithelial cells are fully permissive for ZIKV lytic replication and are primary target cells in the retinal barriers for infection. ZIKV infection of retinal endothelial cells and retinal pericytes induces significantly higher levels of RANTES that likely contributes to ocular inflammation.

Photosynthetic organisms have to tolerate rapid changes in light intensity, which is facilitated by non-photochemical quenching (NPQ) and involves modification of energy transfer from light-harvesting complexes (LHC) to the photosystem reaction centres. NPQ includes dissipating excess light energy to heat (qE) and the reversible coupling of LHCII to photosystems (state transitions/qT), which are considered separate NPQ mechanisms. In the model alga Chlamydomonas reinhardtii the LHCSR3 protein has a well characterised role in qE. Here, it is shown in the npq4 mutant, deficient in LHCSR3, that energy coupling to photosystem II (PSII) more akin to qT is also disrupted, but no major differences in LHC phosphorylation or LHC compositions were found in comparison to wild-type cells. The qT of wild-type cells possessed two kinetically distinguishable phases, with LHCSR3 participating in the more rapid (<2 min) phase. This LHCSR3-mediated qT was sensitive to physiological levels of H 2 O 2 , which accelerated qE induction, revealing a way that may help C. reinhardtii tolerate a sudden increase in light intensity. Overall, a clear mechanistic overlap between qE and qT is shown.

Chloroplast protrusions (CPs) have frequently been observed in plants, but their significance to plant metabolism remains largely unknown. We investigated in the alpine plant Ranunculus glacialis L. treated under various CO2 concentrations if CP formation is related to photorespiration, specifically focusing on hydrogen peroxide (H2O2) metabolism. Immediately after exposure to different CO2 concentrations, the formation of CPs in leaf mesophyll cells was assessed and correlated to catalase (CAT) and ascorbate peroxidase (APX) activities. Under natural irradiation, the relative proportion of chloroplasts with protrusions (rCP) was highest (58.7 %) after exposure to low CO2 (38 ppm) and was lowest (3.0 %) at high CO2 (10,000 ppm). The same relationship was found for CAT activity, which decreased from 34.7 nkat mg(-1) DW under low CO2 to 18.4 nkat mg(-1) DW under high CO2, while APX activity did not change significantly. When exposed to natural CO2 concentration (380 ppm) in darkness, CP formation was significantly lower (18.2 %) compared to natural solar irradiation (41.3 %). In summary, CP formation and CAT activity are significantly increased under conditions that favour photorespiration, while in darkness or at high CO2 concentration under light, CP formation is significantly lower, providing evidence for an association between CPs and photorespiration.

Background and Aims: In photosynthetic organisms exposure to high light induces the production of reactive oxygen species (ROS), such as hydrogen peroxide (H2O2), which in part is prevented by non-photochemical quenching (NPQ). As one of the most stable and longest-lived ROS, H2O2 is involved in key signalling pathways in development and stress responses, although in excess it can induce damage. A ubiquitous response to high light is the induction of the xanthophyll cycle, but its role in algae is unclear as it is not always associated with NPQ induction. The aim of this study was to reveal how diurnal changes in the level of H2O2 are regulated in a freshwater algal community., Methods: A natural freshwater community of algae in a temporary rainwater pool was studied, comprising photosynthetic Euglena species, benthic Navicula diatoms, Chlamydomonas and Chlorella species. Diurnal measurements were made of photosynthetic performance, concentrations of photosynthetic pigments and H2O2. The frequently studied model organisms Chlamydomonas and Chlorella species were isolated to study photosynthesis-related H2O2 responses to high light., Key Results: NPQ was shown to prevent H2O2 release in Chlamydomonas and Chlorella species under high light; in addition, dissolved organic carbon excited by UV-B radiation was probably responsible for a part of the H2O2 produced in the water column. Concentrations of H2O2 peaked at 2 µm at midday and algae rapidly scavenged H2O2 rather than releasing it. A vertical H2O2 gradient was observed that was lowest next to diatom-rich benthic algal mats. The diurnal changes in photosynthetic pigments included the violaxanthin and diadinoxanthin cycles; the former was induced prior to the latter, but neither was strictly correlated with NPQ., Conclusions: The diurnal cycling of H2O2 was apparently modulated by the organisms in this freshwater algal community. Although the community showed flexibility in its levels of NPQ, the diurnal changes in xanthophylls correlated with H2O2 concentrations. Alternative NPQ mechanisms in algae involving proteins of the light-harvesting complex type and antioxidant protection of the thylakoid membrane by de-epoxidized carotenoids are discussed., (© The Author 2015. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Plant surfaces form the barrier between a plant and its environment. Upon damage, the wound healing process begins immediately and is accompanied by a rapid production of extracellular reactive oxygen species (ROS), essential in deterring pathogens, signalling responses and cell wall restructuring. Although many enzymes produce extracellular ROS, it is unclear if ROS-producing enzymes act synergistically. We characterised the oxidative burst of superoxide (O2(·-)) and hydrogen peroxide (H2O2) that follows wounding in pea (Pisum sativum L.) seedlings. Rates of ROS production were manipulated by exogenous application of enzyme substrates and inhibitors. The results indicate significant roles for di-amine oxidases (DAO) and peroxidases (Prx) rather than NADPH oxidase. The burst of O2(·-) was strongly dependent on the presence of H2O2 produced by DAO. Potential substrates released from wounded seedlings included linoleic acid that, upon exogenous application, strongly stimulated catalase-sensitive O2(·-) production. Moreover, a 65kD plasma membrane (PM) guaiacol Prx was found in the secretome of wounded seedlings and showed dependence on linoleic acid for O2(·-) production. Lipoxygenases are suggested to modulate O2(·-) production by consuming polyunsaturated fatty acids in the apoplast. Overall, a O2(·-)-producing mechanism involving H2O2-derived from DAO, linoleic acid and a PM-associated Prx is proposed., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

The production of reactive oxygen species (ROS) is an unavoidable part of photosynthesis. Stress that accompanies high light levels and low CO2 availability putatively includes enhanced ROS production in the so-called Mehler reaction. Such conditions are thought to encourage O2 to become an electron acceptor at photosystem I, producing the ROS superoxide anion radical (O2·-) and hydrogen peroxide (H2 O2 ). In contrast, here it is shown in Chlamydomonas reinhardtii that CO2 depletion under high light levels lowered cellular H2 O2 production, and that elevated CO2 levels increased H2 O2 production. Using various photosynthetic and mitochondrial mutants of C. reinhardtii, the chloroplast was identified as the main source of elevated H2 O2 production under high CO2 availability. High light levels under low CO2 availability induced photoprotective mechanisms called non-photochemical quenching, or NPQ, including state transitions (qT) and high energy state quenching (qE). The qE-deficient mutant npq4 produced more H2 O2 than wild-type cells under high light levels, although less so under high CO2 availability, whereas it demonstrated equal or greater enzymatic H2 O2 -degrading capacity. The qT-deficient mutant stt7-9 produced the same H2 O2 as wild-type cells under high CO2 availability. Physiological levels of H2 O2 were able to hinder qT and the induction of state 2, providing an explanation for why under high light levels and high CO2 availability wild-type cells behaved like stt7-9 cells stuck in state 1., (© 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.)

Photosynthetic organisms and isolated photosystems are of interest for technical applications. In nature, photosynthetic electron transport has to work efficiently in contrasting environments such as shade and full sunlight at noon. Photosynthetic electron transport is regulated on many levels, starting with the energy transfer processes in antenna and ending with how reducing power is ultimately partitioned. This review starts by explaining how light energy can be dissipated or distributed by the various mechanisms of non-photochemical quenching, including thermal dissipation and state transitions, and how these processes influence photoinhibition of photosystem II (PSII). Furthermore, we will highlight the importance of the various alternative electron transport pathways, including the use of oxygen as the terminal electron acceptor and cyclic flow around photosystem I (PSI), the latter which seem particularly relevant to preventing photoinhibition of photosystem I. The control of excitation pressure in combination with the partitioning of reducing power influences the light-dependent formation of reactive oxygen species in PSII and in PSI, which may be a very important consideration to any artificial photosynthetic system or technical device using photosynthetic organisms.

Chlamydomonas reinhardtii is a photoautotrophic green alga, which can be grown mixotrophically in acetate-supplemented media (Tris-acetate-phosphate). We show that acetate has a direct effect on photosystem II (PSII). As a consequence, Tris-acetate-phosphate-grown mixotrophic C. reinhardtii cultures are less susceptible to photoinhibition than photoautotrophic cultures when subjected to high light. Spin-trapping electron paramagnetic resonance spectroscopy showed that thylakoids from mixotrophic C. reinhardtii produced less (1)O2 than those from photoautotrophic cultures. The same was observed in vivo by measuring DanePy oxalate fluorescence quenching. Photoinhibition can be induced by the production of (1)O2 originating from charge recombination events in photosystem II, which are governed by the midpoint potentials (Em) of the quinone electron acceptors. Thermoluminescence indicated that the Em of the primary quinone acceptor (QA/QA(-)) of mixotrophic cells was stabilised while the Em of the secondary quinone acceptor (QB/QB(-)) was destabilised, therefore favouring direct non-radiative charge recombination events that do not lead to (1)O2 production. Acetate treatment of photosystem II-enriched membrane fragments from spinach led to the same thermoluminescence shifts as observed in C. reinhardtii, showing that acetate exhibits a direct effect on photosystem II independent from the metabolic state of a cell. A change in the environment of the non-heme iron of acetate-treated photosystem II particles was detected by low temperature electron paramagnetic resonance spectroscopy. We hypothesise that acetate replaces the bicarbonate associated to the non-heme iron and changes the environment of QA and QB affecting photosystem II charge recombination events and photoinhibition., (Copyright © 2013 Elsevier B.V. All rights reserved.)

In photosynthetic organisms, excess light is a stress that induces production of reactive oxygen species inside the chloroplasts. As a response, the capacity of antioxidative defence mechanisms increases. However, when cells of Chlamydomonas reinhardtii were shifted from dark to high light, a reversible partial inactivation of catalase activity was observed, which correlated with a transient increase in the level of H2 O2 in the 10 μm range. This concentration range seems to be necessary to activate H2 O2 -dependent signalling pathways stimulating the expression of H2 O2 responsive genes, such as the heat shock protein HSP22C. Catalase knock-down mutants had lost the transient accumulation of H2 O2 , suggesting that a decrease in catalase activity was the key element for establishing a transient H2 O2 burst. Catalase was inactivated by a one-electron event consistent with the reduction of a single cysteine. We propose that under high light intensity, the redox state of the photosynthetic electron transport chain is sensed and transmitted to the cytosol to regulate the catalase activity. This allows a transient accumulation of H2 O2 , inducing a signalling event that is transmitted to the nucleus to modulate the expression of chloroplast-directed protection enzymes., (© 2012 Blackwell Publishing Ltd.)

Cadmium (Cd) is highly toxic to plants causing growth reduction and chlorosis. It binds thiols and competes with essential transition metals. It affects major biochemical processes such as photosynthesis and the redox balance, but the connection between cadmium effects at the biochemical level and its deleterious effect on growth has seldom been established. In this study, two Cd hypersensitive mutants, cad1-3 impaired in phytochelatin synthase (PCS1), and nramp3nramp4 impaired in release of vacuolar metal stores, have been compared. The analysis combines genetics with measurements of photosynthetic and antioxidant functions. Loss of AtNRAMP3 and AtNRAMP4 function or of PCS1 function leads to comparable Cd sensitivity. Root Cd hypersensitivities conferred by cad1-3 and nramp3nramp4 are cumulative. The two mutants contrast in their tolerance to oxidative stress. In nramp3nramp4, the photosynthetic apparatus is severely affected by Cd, whereas it is much less affected in cad1-3. In agreement with chloroplast being a prime target for Cd toxicity in nramp3nramp4, the Cd hypersensitivity of this mutant is alleviated in the dark. The Cd hypersensitivity of nramp3nramp4 mutant highlights the critical role of vacuolar metal stores to supply essential metals to plastids and maintain photosynthetic function under Cd and oxidative stresses., (© 2012 Blackwell Publishing Ltd.)

Absorption of light in excess of the capacity for photosynthetic electron transport is damaging to photosynthetic organisms. Several mechanisms exist to avoid photodamage, which are collectively referred to as nonphotochemical quenching. This term comprises at least two major processes. State transitions (qT) represent changes in the relative antenna sizes of photosystems II and I. High energy quenching (qE) is the increased thermal dissipation of light energy triggered by lumen acidification. To investigate the respective roles of qE and qT in photoprotection, a mutant (npq4 stt7-9) was generated in Chlamydomonas reinhardtii by crossing the state transition-deficient mutant (stt7-9) with a strain having a largely reduced qE capacity (npq4). The comparative phenotypic analysis of the wild type, single mutants, and double mutants reveals that both state transitions and qE are induced by high light. Moreover, the double mutant exhibits an increased photosensitivity with respect to the single mutants and the wild type. Therefore, we suggest that besides qE, state transitions also play a photoprotective role during high light acclimation of the cells, most likely by decreasing hydrogen peroxide production. These results are discussed in terms of the relative photoprotective benefit related to thermal dissipation of excess light and/or to the physical displacement of antennas from photosystem II.