The pollen quality of woody and herbaceous plants from the Chernobyl exclusion zone.

Pollen abnormalities frequency of Scots pine and pollen sterility of white clover, common columbine, and greater celandine growing in the Bryansk and Gomel' regions radioactively contaminated after the accident at the Chernobyl nuclear power plant (NPP) were assessed. The frequency of abnormal pine pollen at the most contaminated plots significantly exceeded the control level and positively correlated with the total absorbed dose and the absorbed dose from ß-radiation. No sustainable significant changes in pollen sterility of the herbaceous plants studied were found in the exclusion zone of the Chernobyl NPP.

Correction: Podlutskii et al. Arabidopsis thaliana Accessions from the Chernobyl Exclusion Zone Show Decreased Sensitivity to Additional Acute Irradiation. Plants 2022, 11, 3142.

In the original publication [...].

Comparative Analysis of the Effect of Gamma-, Electron, and Proton Irradiation on Transcriptomic Profile of Hordeum vulgare L. Seedlings: In Search for Molecular Contributors to Abiotic Stress Resilience.

The development of adaptation strategies for crops under ever-changing climate conditions is a critically important food security issue. Studies of barley responses to ionising radiation showed that this evolutionarily ancient stress factor can be successfully used to identify molecular pathways involved in adaptation to a range of abiotic stressors. In order to identify potential molecular contributors to abiotic stress resilience, we examined the transcriptomic profiles of barley seedlings after exposure to γ-rays, electrons, and protons. A total of 553 unique differentially expressed genes with increased expression and 124 with decreased expression were detected. Among all types of radiation, the highest number of differentially expressed genes was observed in electron-irradiated samples (428 upregulated and 56 downregulated genes). Significant upregulation after exposure to the three types of radiation was shown by a set of ROS-responsive genes, genes involved in DNA repair, cell wall metabolism, auxin biosynthesis and signalling, as well as photosynthesis-related genes. Most of these genes are known to be involved in plant ROS-mediated responses to other abiotic stressors, especially with genotoxic components, such as heavy metals and drought. Ultimately, the modulation of molecular pathways of plant responses to ionising radiation may be a prospective tool for stress tolerance programmes.

Radiation Hormesis in Barley Manifests as Changes in Growth Dynamics Coordinated with the Expression of PM19L-like, CML31-like, and AOS2-like.

The stimulation of growth and development of crops using ionising radiation (radiation hormesis) has been reported by many research groups. However, specific genes contributing to the radiation stimulation of plant growth are largely unknown. In this work, we studied the impact of the low-dose γ-irradiation of barley seeds on the growth dynamics and gene expression of eight barley cultivars in a greenhouse experiment. Our findings confirmed that candidate genes of the radiation growth stimulation, previously established in barley seedlings (PM19L-like, CML31-like, and AOS2-like), are significant in radiation hormesis throughout ontogeny. In γ-stimulated cultivars, the expression of these genes was aligned with the growth dynamics, yield parameters, and physiological conditions of plants. We identified contrasting cultivars for future gene editing and found that the γ-stimulated cultivar possessed some specific abiotic stress-responsive elements in the promotors of candidate genes, possibly revealing a new level of radiation hormesis effect execution. These results can be used in creating new productive barley cultivars, ecological toxicology of radionuclides, and eustress biology studies.

Multi-Omics Analysis of Vicia cracca Responses to Chronic Radiation Exposure in the Chernobyl Exclusion Zone.

Our understanding of the long-term consequences of chronic ionising radiation for living organisms remains scarce. Modern molecular biology techniques are helpful tools for researching pollutant effects on biota. To reveal the molecular phenotype of plants growing under chronic radiation exposure, we sampled Vicia cracca L. plants in the Chernobyl exclusion zone and areas with normal radiation backgrounds. We performed a detailed analysis of soil and gene expression patterns and conducted coordinated multi-omics analyses of plant samples, including transcriptomics, proteomics, and metabolomics. Plants growing under chronic radiation exposure showed complex and multidirectional biological effects, including significant alterations in the metabolism and gene expression patterns of irradiated plants. We revealed profound changes in carbon metabolism, nitrogen reallocation, and photosynthesis. These plants showed signs of DNA damage, redox imbalance, and stress responses. The upregulation of histones, chaperones, peroxidases, and secondary metabolism was noted.

White clover from the exclusion zone of the Chernobyl NPP: Morphological, biochemical, and genetic characteristics.

A comprehensive study of the biological effects of chronic radiation exposure (8 µGy/h) in populations of white clover (Trifolium repens L.) from the Chernobyl exclusion zone was carried out. White clover is one of the most important pasture legumes, having many agricultural applications. Studies at two reference and three radioactively contaminated plots showed no stable morphological effects in white clover at this level of radiation exposure. Increased activities of catalase and peroxidases were found in some impacted plots. Auxin concentration was enhanced in the radioactively contaminated plots. Genes involved in the maintenance of water homeostasis and photosynthetic processes (TIP1 and CAB1) were upregulated at radioactively contaminated plots.

Chronic Ionizing Radiation of Plants: An Evolutionary Factor from Direct Damage to Non-Target Effects.

In present times, the levels of ionizing radiation (IR) on the surface of Earth are relatively low, posing no high challenges for the survival of contemporary life forms. IR derives from natural sources and naturally occurring radioactive materials (NORM), the nuclear industry, medical applications, and as a result of radiation disasters or nuclear tests. In the current review, we discuss modern sources of radioactivity, its direct and indirect effects on different plant species, and the scope of the radiation protection of plants. We present an overview of the molecular mechanisms of radiation responses in plants, which leads to a tempting conjecture of the evolutionary role of IR as a limiting factor for land colonization and plant diversification rates. The hypothesis-driven analysis of available plant genomic data suggests an overall DNA repair gene families' depletion in land plants compared to ancestral groups, which overlaps with a decrease in levels of radiation exposure on the surface of Earth millions of years ago. The potential contribution of chronic IR as an evolutionary factor in combination with other environmental factors is discussed.

Differential gene expression in chronically irradiated herbaceous species from the Chernobyl exclusion zone.

PURPOSE: Transcriptional activity of genes related to ionizing radiation responses in chronically irradiated plant populations at radioactively contaminated territories can be a cost-effective and precise approach for stress response evaluation. However, there are limits to studying non-model plants in field conditions. The work studies the transcriptional activity of candidate genes of adaptation to chronic radiation exposure in plant populations from radioactively contaminated territories of the Chernobyl. MATERIALS AND METHODS: In this work, we studied plant species with different sensitivity to acute irradiation: Trifolium repens L., Taraxacum officinale Wigg., and Dactylis glomerata L., sampled in the Chernobyl exclusion zone. The differential expression of several candidate genes of adaptation to chronic radiation exposure in the leaves of these species was analyzed, including homologs of Arabidopsis thaliana genes SLAC1, APX1, GPX2, CAB1, NTRB, PP2-B11, RBOH-F, HY5, SnRK2.4, PDS1, CIPK20, SIP1, PIP1, TIP1. RESULTS AND CONCLUSIONS: All studied species were characterized by upregulation of the CAB1 homolog, encoding chlorophyll a/b binding protein, at radioactively contaminated plots. An increase in the expression of genes associated with water and hydrogen peroxide transport, intensity of photosynthesis, and stress responses (homolog of aquaporin TIP1 for T. repens; homologs of aquaporin PIP1 and transcription factor HY5 for D. glomerata; homolog of CBL-interacting serine/threonine protein kinase CIPK20 for T. officinale) was revealed. The methodological approach for studying gene expression in non-model plant species is described, which may allow large-scale screening studies of candidate genes in various plant species abundant in radioactively contaminated areas.

Arabidopsis thaliana Accessions from the Chernobyl Exclusion Zone Show Decreased Sensitivity to Additional Acute Irradiation.

Chronic ionising radiation exposure is a main consequence of radioactive pollution of the environment. The development of functional genomics approaches coupled with morphological and physiological studies allows new insights into plant adaptation to life under chronic irradiation. Using morphological, reproductive, physiological, and transcriptomic experiments, we evaluated the way in which Arabidopsis thaliana natural accessions from the Chernobyl exclusion zone recover from chronic low-dose and acute high-dose γ-irradiation of seeds. Plants from radioactively contaminated areas were characterized by lower germination efficiency, suppressed growth, decreased chlorophyll fluorescence, and phytohormonal changes. The transcriptomes of plants chronically exposed to low-dose radiation indicated the repression of mobile genetic elements and deregulation of genes related to abiotic stress tolerance. Furthermore, these chronically irradiated natural accessions showed higher tolerance to acute 150 Gy γ-irradiation of seeds, according to transcriptome and phytohormonal profiles. Overall, the lower sensitivity of the accessions from radioactively contaminated areas to acute high-dose irradiation may come at the cost of their growth performance under normal conditions.

A Pipeline for Non-model Organisms for de novo Transcriptome Assembly, Annotation, and Gene Ontology Analysis Using Open Tools: Case Study with Scots Pine.

RNA sequencing (RNA-seq) has opened up the possibility of studying virtually any organism at the whole transcriptome level. Nevertheless, the absence of a sequenced and accurately annotated reference genome may be an obstacle for applying this technique to non-model organisms, especially for those with a complex genome. While de novo transcriptome assembly can circumvent this problem, it is often computationally demanding. Furthermore, the transcriptome annotation and Gene Ontology enrichment analysis without an automatized system is often a laborious task. Here we describe step-by-step the pipeline that was used to perform the transcriptome assembly, annotation, and Gene Ontology analysis of Scots pine (Pinus sylvestris), a gymnosperm species with complex genome. Using only free software available for the scientific community and running on a standard personal computer, the pipeline intends to facilitate transcriptomic studies for non-model species, yet being flexible to be used with any organism.

Metabolic Profiling of γ-Irradiated Barley Plants Identifies Reallocation of Nitrogen Metabolism and Metabolic Stress Response.

The favorable responses of crop species to low-dose γ irradiation can help to develop cultivars with increased productivity and improved stress tolerance. In the present study, we tried to reveal the candidate metabolites involved in growth stimulation of barley seedlings after applying low-dose γ-radiation (60Co) to seeds. Stimulating doses (5-20 Gy) provided a significant increase in shoot length and biomass, while relatively high dose of 100 Gy led to significant inhibition of growth. Gas chromatography-mass spectrometry metabolomic analysis uncovered several compounds that may take part in radiation hormesis establishment in irradiated plants. This includes molecules involved in nitrogen redistribution (arginine, glutamine, asparagine, and γ-aminobutyric acid) and stress-responsive metabolites, such as ascorbate, myo-inositol and its derivates, and free amino acids (l-serine, ß-alanine, pipecolate, and GABA). These results contribute to the understanding of the molecular mechanisms of hormesis phenomenon.

Seed Gamma Irradiation of Arabidopsis thaliana ABA-Mutant Lines Alters Germination and Does Not Inhibit the Photosynthetic Efficiency of Juvenile Plants.

Plant growth response to γ-irradiation includes stimulating or inhibitory effects depending on plant species, dose applied, stage of ontogeny and other factors. Previous studies showed that responses to irradiation could depend on ABA accumulation and signaling. To elucidate the role of ABA in growth and photosynthetic responses to irradiation, lines Col-8, abi3-8 and aba3 -1 of Arabidopsis thaliana were used. Seeds were γ-irradiated using 60Co in the dose range 50-150 Gy. It was revealed that the dose of 150 Gy affected germination parameters of aba3 -1 and Col-8 lines, while abi3-8 line was the most resistant to the studied doses and even showed faster germination at early hours after γ-irradiation at 50 Gy. These results suggest that susceptibility to ABA is probably more important for growth response to γ-irradiation than ABA synthesis. The photosynthetic functioning of 16-day-old plants mainly was not disturbed by γ-irradiation of seeds, and no indication of photosystem II photoinhibition was noticed, revealing the robustness of the photosynthetic system of A. thaliana. Glutathione peroxidase activity and ABA concentrations in plant tissues were not affected in the studied dose range. These results contribute to the understanding of germination and photosynthesis fine-tuning and of mechanisms of plant tolerance to ionizing radiation.

Scots pine as a promising indicator organism for biomonitoring of the polluted environment: A case study on chronically irradiated populations.

In this paper the main results of long-term (2003-2016) observations on Scots pine populations inhabiting sites affected by the Chernobyl accident are presented. Populations growing for many years under chronic radiation exposure are characterized by the enhanced mutation rates, increased genetic diversity, changes in the gene expression and in the level of genome-wide methylation, alterations in the temporal dynamics of cytogenetic abnormalities and genetic structure of populations. However, significant changes at the genetic level had no effects on enzymatic activity, morphological abnormalities, and reproductive ability of pine trees. The results presented increase our understanding of the long-term effects of chronic radiation exposure on plant populations in the wild nature and provide important information for the management and monitoring of radioactively contaminated territories.

The response profile to chronic radiation exposure based on the transcriptome analysis of Scots pine from Chernobyl affected zone.

Radioactive contamination of the natural areas is one of the most long-lasting anthropogenic impacts on the environment. Scots pine (Pinus sylvestris L.) is a promising organism for radiation-related research because of its high radiosensitivity, but the genome size of Pinacea species has imposed obstacles for high-throughput studies so far. In this work, we conducted the analysis of the de novo assembled transcriptome of Scots pine populations growing in the Chernobyl-affected zone, which is still today contaminated with radionuclides because of the accident at the nuclear power plant in 1986. The transcriptome profiles indicate a clear pattern of adaptive stress response, which seems to be dose-dependent. The transcriptional response indicates a continuous modulation of the cellular redox system, enhanced expression of chaperones and histones, along with the control of ions balance. Interestingly, the activity of transposable element families is inversely correlated to the exposure levels to radiation. These adaptive responses, which are triggered by radiation doses 30 times lower than the one accepted as a safe for biota species by international regulations, suggest that the environmental management in radiation protection should be reviewed.

Evolutionary history of the Pasque-flowers (Pulsatilla, Ranunculaceae): Molecular phylogenetics, systematics and rDNA evolution.

Pulsatilla (Anemoneae, Ranunculaceae) is sister to Anemone s.s. and contains ca 40 perennial species of considerable horticultural and medical importance. We sequenced 31 of those species, plus nine subspecies, two cultivars and six outgroups, for two nuclear regions (high-copy nrITS and low-copy MLH1) and three plastid regions (rbcL, accD-psaI, trnL intron) in order to generate the first comprehensive species-level phylogeny of the genus. Phylogenetic trees were constructed using both concatenation-based (maximum likelihood and Bayesian inference) and coalescence methods. The better supported among the internal nodes were subjected to molecular clock dating and ancestral area reconstruction, and karyotypic characters identified by us using Fluorescence In Situ Hybridization were mapped across the tree. The preferred species tree from the coalescence analysis formed the basis of a new infrageneric classification based on monophyly plus degree of divergence. The earliest divergent of the three subgenera, Kostyczewianae, is represented by only a single species that is morphologically intermediate between Anemone s.s. and 'core' Pulsatilla. Subgenus Pulsatilla is considerably richer in species than its sister subgenus Preonanthus and contains three monophyletic sections. Species possessing nodding flowers and pectinately dissected leaves are phylogenetically derived compared with groups possessing erect flowers and palmately lobed leaves. Pulsatilla separated from Anemone s.s. at ca 25 Ma. Our results indicate a central Asian mountain origin of the genus and an initial diversification correlated with late Tertiary global cooling plus regional mountain uplift, aridification and consequent expansion of grasslands. The more rapid and extensive diversification within subgenus Pulsatilla began at ca 3 Ma and continued throughout the Quaternary, driven not only by major perturbations in global climate but also by well-documented polyploidy.

'Omic' technologies as a helpful tool in radioecological research.

This article presents a brief review of the modern 'omic' technologies, namely genomics, epigenomics, transcriptomics, proteomics, and metabolomics, as well as the examples of their possible use in radioecology. For each technology, a short description of advances, limitations, and instrumental applications is given. In addition, the review contains examples of successful use of 'omic' technologies in the assessment of biological effects of pollutants in the field conditions.

Radiation exposure of barley seeds can modify the early stages of plants' development.

The reactions of barley seeds (Nur and Grace varieties) in terms of the root and sprout lengths, germination and root mass were studied after γ-irradiation with doses in the range of 2-50 Gy. The dose range in which plants' growth stimulation occurs (16-20 Gy) was identified. It was shown that increased size of seedlings after irradiation with stimulating doses was due to the enhancing pace of development rather than an earlier germination. The activity of the majority of the enzymes studied increased in the range of doses that cause stimulation of seedlings development. The influences of the dose rate, the quality of seeds, their moisture and time interval between irradiation and initiation of germination on the manifestation of the effects of radiation were investigated. The experimental data on the effect of γ-irradiation on seedlings development were significantly better explained by mathematical models that take into account the hormetic effect.

Influence of long-term chronic exposure and weather conditions on Scots pine populations.

Over a period of 8 years (2007-2014), we were evaluating seed quality and morphological abnormalities in Scots pine trees affected as a result of the Chernobyl accident. The calculated dose rates for the trees at the study sites varied from background values at the reference sites to 40 mGy/year at the most contaminated site. We investigated whether radioactive contamination and/or weather factors could decrease the reproductive capacity or increase the frequency of morphological abnormalities of needles in pine trees. Scots pine seeds are characterized by high interannual variability of viability, which is largely determined by weather conditions. No consistent differences in reproductive capacity were detected between the impacted and reference populations. Brachyblasts with three needles were found only in the affected populations; however, their frequency was very low and only at the very border of significance at the p < 0.10 level.

Radiation exposure in the remote period after the Chernobyl accident caused oxidative stress and genetic effects in Scots pine populations.

Even 30 years after the Chernobyl accident, biological effects of irradiation are observed in the chronically exposed Scots pine populations. Chronic radiation exposure at dose rates above 50 mGy∙yr-1 caused oxidative stress and led to the increase of antioxidants concentrations in these populations. Genetic variability was examined for 6 enzymes and 14 enzymatic loci of 6 Scots pine populations. Dose rates over 10 mGy∙yr-1 caused the increased frequency of mutations and changes in genetic structure of Scots pine populations. However, the same dose rates had no effect on enzymatic activities. The results indicate that even relatively low dose rates of radiation can be considered as an ecological factor which should be taken into account for ecological management and radiation protection of biota species.

Genetic diversity in Scots pine populations along a radiation exposure gradient.

Polymorphisms of antioxidant enzymes were studied in the endosperm and embryos of seeds from Scots pine populations inhabiting sites in the Bryansk region of Russia radioactively contaminated as a result of the Chernobyl accident. Chronic radiation exposure at dose rates from 0.8 µGy/h led to a significant increase in the rate of enzymatic loci mutations. The main parameters of genetic variability of the affected Scots pine populations had considerably higher values than those from the reference site. Changes in the genetic makeup of Scots pine populations were observed at dose rates greater than 10.4 µGy/h. However, the higher mutation rate had no effect on the activities of antioxidant enzymes.