Mitogenomic Research of Silverleaf Sunflower (Helianthus argophyllus) and Its Interspecific Hybrids.

Interspecific hybridization is widespread for sunflowers, both in wild populations and commercial breeding. One of the most common species that can efficiently cross with Helianthus annuus is the Silverleaf sunflower-Helianthus argophyllus. The current study carried out structural and functional organization analyses of mitochondrial DNA in H. argophyllus and the interspecific hybrid, H. annuus (VIR114A line) × H. argophyllus. The complete mitogenome of H. argophyllus counts 300,843 bp, has a similar organization to the mitogenome of cultivated sunflower, and holds SNPs typical for wild sunflowers. RNA editing analysis predicted 484 sites in H. argophyllus mitochondrial CDS. The mitochondrial genome of the H. annuus × H. argophyllus hybrid is identical to the maternal line (VIR114A). We expected that significant rearrangements in the mitochondrial DNA of the hybrid would occur, due to the frequent recombination. However, the hybrid mitogenome lacks rearrangements, presumably due to the preservation of nuclear-cytoplasmic interaction paths.

Effects of bulk and nano-ZnO particles on functioning of photosynthetic apparatus in barley (Hordeum vulgare L.).

The functioning of the photosynthetic apparatus in barley (Hordeum vulgare L.) after 7-days of exposure to bulk (b-ZnO) and nanosized ZnO (n-ZnO) (300, 2000, and 10,000 mg/l) has been investigated. An impact on the amount of chlorophylls, photosynthetic efficiency, as well as the zinc accumulation in chloroplasts was demonstrated. Violation of the chloroplast fine structure was revealed. These changes were generally more pronounced with n-ZnO exposure, especially at high concentrations. For instance, the chlorophyll deficiency under 10,000 mg/l b-ZnO treatment was 31% and with exposure to 10,000 mg/l n-ZnO, the chlorophyll deficiency was already 52%. The expression analysis of the photosynthetic genes revealed their different sensitivity to b-ZnO and n-ZnO exposure. The genes encoding subunits of photosystem II (PSII) and, to a slightly lesser extent, photosystem I (PSI) showed the highest suppression of transcriptional levels. The mRNA levels of the subunits of cytochrome-b6f, NADH dehydrogenase, ribulose-1,5-bisphosphate carboxylase and ATP synthase, which, in addition to linear electron flow (LEF), participate in cyclic electron flow (CEF) and autotrophic CO2 fixation, were more stable or increased under b-ZnO and n-ZnO treatments. At the same time, CEF was increased. It was assumed that under the action of b-ZnO and n-ZnO, the processes of LEF are disrupted, and CEF is activated. This allows the plant to prevent photo-oxidation and compensate for the lack of ATP for the CO2 fixation process, thereby ensuring the stability of photosynthetic function in the initial stages of stress factor exposure. The study of photosynthetic structures of crops is important from the point of view of understanding the risks of reducing the production potential and the level of food security due to the growing use of nanoparticles in agriculture.

Dependence of maize yield on hydrothermal factors in various agro-climatic zones of the Rostov region of Russia in the context of climate change.

Trends in mean monthly temperature and precipitation during the growing season and their effects on the maize yield were analyzed at the Zimovnikovsky (Zim) and Rostov (Ros) state variety plots (SVPs), located in different agro-climatic zones of the Rostov region. For these two SVPs, in the period of 1975-2019, the Mann-Kendall test showed a statistically significant increase (p < 0.05) in mean temperature (0.70 and 0.52 °C/decade) and a trend of decreased total precipitation (- 14.81 and - 10.40 mm/decade) during the maize growing season. The dependence of the maize yield on hydrothermal factors was estimated for the period of 2011-2019 using the Pearson correlation coefficient (p < 0.05). The mean temperature in September at Zim negatively (r = - 0.78), and in June at Ros positively (r = 0.77) correlated with yield, which explained, according to the value of the coefficient of determination (R2), up to 60.7% and 58.7%, respectively, of the interannual variability of the maize yield. The precipitation in July at the Zim and Ros positively correlated (r = 0.75 and r = 0.71) with yield and explained up to 55.9% and 50.6%, respectively, of the interannual variability of the maize yield. The total amount of precipitation during the growing season at Zim was the dominant factor, explaining up to 75.7% of the interannual variability of maize yield. The continuation of the observed climatic trends during the growing season could lead in the next decade to both a decrease in the maize yield by an average of 0.25 t/ha at Zim and an increase in the maize yield by an average of 0.42 t/ha at Ros.

A point mutation in the photosystem I P700 chlorophyll a apoprotein A1 gene confers variegation in Helianthus annuus L.

KEY MESSAGE: Even a point mutation in the psaA gene mediates chlorophyll deficiency. The role of the plastid signal may perform the redox state of the compounds on the acceptor-side of PSI. Two extranuclear variegated mutants of sunflower, Var1 and Var33, were investigated. The yellow sectors of both mutants were characterized by an extremely low chlorophyll and carotenoid content, as well as poorly developed, unstacked thylakoid membranes. A full-genome sequencing of the cpDNA revealed mutations in the psaA gene in both Var1 and Var33. The cpDNA from the yellow sectors of Var1 differs from those in the wild type by only a single, non-synonymous substitution (Gly734Glu) in the psaA gene, which encodes a subunit of photosystem (PS) I. In the cpDNA from the yellow sectors of Var33, the single-nucleotide insertion in the psaA gene was revealed, leading to frameshift at the 580 amino acid position. Analysis of the photosynthetic electron transport demonstrated an inhibition of the PSI and PSII activities in the yellow tissues of the mutant plants. It has been suggested that mutations in the psaA gene of both Var1 and Var33 led to the disruption of PSI. Due to the non-functional PSI, photosynthetic electron transport is blocked, which, in turn, leads to photodamage of PSII. These data are confirmed by immunoblotting analysis, which showed a significant reduction in PsbA in the yellow leaf sectors, but not PsaA. The expression of chloroplast and nuclear genes encoding the PSI subunits (psaA, psaB, and PSAN), the PSII subunits (psbA, psbB, and PSBW), the antenna proteins (LHCA1, LHCB1, and LHCB4), the ribulose 1.5-bisphosphate carboxylase subunits (rbcL and RbcS), and enzymes of chlorophyll biosynthesis were down-regulated in the yellow leaf tissue. The extremely reduced transcriptional activity of the two protochlorophyllide oxidoreductase (POR) genes involved in chlorophyll biosynthesis is noteworthy. The disruption of NADPH synthesis, due to the non-functional PSI, probably led to a significant reduction in NADPH-protochlorophyllide oxidoreductase in the yellow sectors of Var1 and Var33. A dramatic decrease in chlorophyllide was shown in the yellow sectors. A reduction in NADPH-protochlorophyllide oxidoreductase, along with photodegradation, has been suggested as a result of chlorophyll deficiency.

Characterization of the mitochondrial genome of the MAX1 type of cytoplasmic male-sterile sunflower.

BACKGROUND: More than 70 cytoplasmic male sterility (CMS) types have been identified in Helianthus, but only for less than half of them, research of mitochondrial organization has been conducted. Moreover, complete mitochondrion sequences have only been published for two CMS sources - PET1 and PET2. It has been demonstrated that other sunflower CMS sources like MAX1, significantly differ from the PET1 and PET2 types. However, possible molecular causes for the CMS induction by MAX1 have not yet been proposed. In the present study, we have investigated structural changes in the mitochondrial genome of HA89 (MAX1) CMS sunflower line in comparison to the fertile mitochondrial genome. RESULTS: Eight significant major reorganization events have been determined in HA89 (MAX1) mtDNA: one 110 kb inverted region, four deletions of 439 bp, 978 bp, 3183 bp and 14,296 bp, respectively, and three insertions of 1999 bp, 5272 bp and 6583 bp. The rearrangements have led to functional changes in the mitochondrial genome of HA89 (MAX1) resulting in the complete elimination of orf777 and the appearance of new ORFs - orf306, orf480, orf645 and orf1287. Aligning the mtDNA of the CMS sources PET1 and PET2 with MAX1 we found some common reorganization features in their mitochondrial genome sequences. CONCLUSION: The new open reading frame orf1287, representing a chimeric atp6 gene, may play a key role in MAX1 CMS phenotype formation in sunflower, while the contribution of other mitochondrial reorganizations seems to appear negligible for the CMS development.

Origin of CMS-PET1 cytotype in cultivated sunflower: A new insight.

The vast majority of commercial sunflower hybrids worldwide are produced using cytoplasmic male sterility (CMS) of the PET1 type, resulting from the interspecific hybridization of Helianthus petiolaris with Helianthus annuus. Due to the fact that CMS-PET1 was not previously detected in wild sunflower, it was believed that this cytotype could arise during interspecific hybridization and is specific solely for cultivated sunflower. In this study, the open reading frame, orfH522, associated with the CMS-PET1 phenotype, was revealed for the first time in the 3'-flanking region of the mitochondrial atpA gene in wild H. annuus. An analysis of whole genome data from 1089 accessions showed that the frequency of occurrence of CMS-orfH522 in wild H. annuus populations is 3.58%, while in wild H. petiolaris populations, it is 1.26%. In general, the analysis demonstrated that PET1-CMS is a natural cytotype of H. annuus, and the appearance of the CMS phenotype in cultivated sunflowers is associated with the loss of stabilizing nuclear genes of fertility restorers, which occurred during interspecific hybridization. These data can explain the patterns of differential cytoplasmic and nuclear introgression occurring in wild sunflower and are useful for further evolutionary studies.

Effects of ZnO nanoparticles and its bulk form on growth, antioxidant defense system and expression of oxidative stress related genes in Hordeum vulgare L.

A comparative analysis of physio-biochemical indices and transcriptional activity of oxidative stress genes in barley (Hordeum vulgare L.) seedlings after 7-days exposure to bulk- and nano-ZnO (300 and 2000 mg/L) was carried out. A dose-dependent reduction in the length and weight of roots and shoots, as well as a significant accumulation of Zn in plant parts, was shown. Alterations in the shape and size of organelles, cytoplasmic vacuolization, and chloroplast and mitochondrial disorganization were also revealed. These processes are particularly pronounced when H. vulgare is exposed to the higher concentrations of nano-ZnO. The study of the antioxidant defense system revealed mainly an increase in the level of reduced glutathione and the activities of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), and glutathione S-transferase (GST). The increases in activity, by 4-fold and 3-fold, was found for glutathione transferase in the roots when exposed to 2000 mg/L bulk- and nano-ZnO, respectively. The study of transcriptional activity demonstrated that in the roots under the influence of bulk- and nano-ZnO, along with Mn-SOD, Fe-SOD is highly expressed, mainly associated with the protection of chloroplasts. Analysis of the Cat 1 and Cat 2 gene expression showed that the main contribution to the increase in catalase activity in treated H. vulgare is made by the CAT-1 isozyme. Generally, in response to the impact of the studied ZnO forms, the antioxidant defense system is activated in H. vulgare, which effectively prevents the progression of oxidative stress in early stages of plant ontogenesis. Nevertheless, with constant exposure to bulk- and nano-ZnO at high concentrations, such activation leads to a depletion of the plant's energy resources, which negatively affects its growth and development. The results obtained could be useful in predicting the risks associated with the further transfer of nano-ZnO to the environment.

Comparative analysis of chloroplast genomes of seven perennial Helianthus species.

Sequencing and a comparative analysis of the complete chloroplast genomes of seven perennial Helianthus species were carried out. The chloroplast genomes have a typical quadripartite structure, including large and small single regions and a pair of inverted repeats. Genome sizes were between 151,152 bp and 151,289 bp. The genome of H. grosseserratus was the smallest, while that of H. microcephalus was the largest. The size variation of the chloroplast genomes is substantially due to the change in the length of simple sequence repeats (SSRs) in non-coding regions. An analysis of these SSRs revealed 35 polymorphic loci (average PIC value > 0.5) that can be used to examine ecological and evolutionary processes in wild Helianthus species. Eight divergence hotspots, including five intergenic regions (petN-psbM, clpP intron, rps3-rpl16, ndhD-ccsA, and ndhF-rpl32) and three gene regions (rbcL, ycf1, and ndhF) were also identified in Helianthus chloroplast genomes. The evolutionary selection pressure analysis revealed a strong purifying selection. Only the rbcL gene experienced efficiency of positive selection at the annual/perennial transitions. The inverted repeat (IR)/single copy (SC) boundaries were identical in all of these (Helianthus) species. In general, the comparison of the genomes revealed low levels of sequence variability (Pi = 0.00051). This indicates that the chloroplast genomes of the studied perennial species of Helianthus, in addition to purifying selection, are closely related and have a recent divergence time.

Data on the polymorphic sites in the chloroplast genomes of seven perennial Helianthus species.

Data present the chloroplast genome sequences of seven wild perennial Helianthus species obtained by using the Illumina HiSeq and NextSeq platforms. Datasets not included in the primary publication [1] are a source for further evolutionary studies. In particular, the annotated chloroplast genomes and datasets of single nucleotide polymorphisms (SNP), simple sequence repeats (SSR), insertion and deletion polymorphisms (INDEL) for H. tuberosus, H. salicifolius, H. pauciflorus, H. microcephalis, H. hirsutus, H. strumosus, and H. grosseserratus are presented. The raw reads are available in Figshare (https://doi.org/10.6084/m9.figshare.12600155). The complete chloroplast genome sequences for the seven perennial Helianthus species are available on GenBank NCBI under the accessions MT302562.1 - MT302568.1; the remaining data are provided in this article.

The Investigation of Perennial Sunflower Species (Helianthus L.) Mitochondrial Genomes.

The genus Helianthus is a diverse taxonomic group with approximately 50 species. Most sunflower genomic investigations are devoted to economically valuable species, e.g., H. annuus, while other Helianthus species, especially perennial, are predominantly a blind spot. In the current study, we have assembled the complete mitogenomes of two perennial species: H. grosseserratus (273,543 bp) and H. strumosus (281,055 bp). We analyzed their sequences and gene profiles in comparison to the available complete mitogenomes of H. annuus. Except for sdh4 and trnA-UGC, both perennial sunflower species had the same gene content and almost identical protein-coding sequences when compared with each other and with annual sunflowers (H. annuus). Common mitochondrial open reading frames (ORFs) (orf117, orf139, and orf334) in sunflowers and unique ORFs for H. grosseserratus (orf633) and H. strumosus (orf126, orf184, orf207) were identified. The maintenance of plastid-derived coding sequences in the mitogenomes of both annual and perennial sunflowers and the low frequency of nonsynonymous mutations point at an extremely low variability of mitochondrial DNA (mtDNA) coding sequences in the Helianthus genus.

Rice Breeding in Russia Using Genetic Markers.

The article concentrates on studying tolerance to soil salinization, water flooding, and blast in Russian and Asian rice varieties, as well as hybrids of the second and third generations from their crossing in order to obtain sustainable paddy crops based on domestic varieties using DNA markers. Samples IR 52713-2B-8-2B-1-2, IR 74099-3R-3-3, and NSIC Rc 106 were used as donors of the SalTol tolerance gene. Varieties with the Sub1A locus were used as donors of the flood resistance gene: Br-11, CR-1009, Inbara-3, TDK-1, and Khan Dan. The lines C101-A-51 (Pi-2), C101-Lac (Pi-1, Pi-33), IR-58 (Pi-ta), and Moroberekan (Pi-b) were used to transfer blast resistance genes. Hybridization of the stress-sensitive domestic varieties Novator, Flagman, Virazh, and Boyarin with donor lines of the genes of interest was carried out. As a result of the studies carried out using molecular marking based on PCR in combination with traditional breeding, early-maturing rice lines with genes for resistance to salinity (SalTol) and flooding (Sub1A), suitable for cultivation in southern Russia, were obtained. Introgression and pyramiding of the blast resistance genes Pi-1, Pi-2, Pi-33, Pi-ta, and Pi-b into the genotypes of domestic rice varieties were carried out. DNA marker analysis revealed disease-resistant rice samples carrying 5 target genes in a homozygous state. The created rice varieties that carry the genes for blast resistance (Pentagen, Magnate, Pirouette, Argamac, Kapitan, and Lenaris) were submitted for state variety testing. The introduction of such varieties into production will allow us to avoid epiphytotic development of the disease, preserving the biological productivity of rice and obtaining environmentally friendly agricultural products.

Data on the polymorphic sites in the chloroplast genomes of the sunflower alloplasmic CMS lines.

Data presents the chloroplast genome sequences of the five sunflower alloplasmic cytoplasmic male sterility (CMS) lines obtained with using the Illumina MiSeq, HiSeq and NextSeq platforms. The sunflower alloplasmic CMS lines has the same nuclear genome from line HA89, but they differ in cytoplasmic genomes, inherited from annual (PET1, PET2 - H. petiolaris, ANN2 - H. annuus) and perennial (MAX1 - H. maximilliani) species of the genus Helianthus L. The chloroplast genomes were annotated. Also presented is a dataset of variable sites such as single nucleotide polymorphism (SNP), simple sequence repeat (SSR), insertion and deletion (INDEL) in the chloroplast genome of the sequenced alloplasmic lines. The raw reads are available in FIGSHARE (https://doi.org/10.6084/m9.figshare.7520183). The complete chloroplast genome sequences for the sunflower alloplasmic lines are available in GenBank NCBI under the accessions MK341448.1-MK341452.1; the remaining data are provided with this article.

Organization Features of the Mitochondrial Genome of Sunflower (Helianthus annuus L.) with ANN2-Type Male-Sterile Cytoplasm.

This study provides insights into the flexibility of the mitochondrial genome in sunflower (Helianthus annuus L.) as well as into the causes of ANN2-type cytoplasmic male sterility (CMS). De novo assembly of the mitochondrial genome of male-sterile HA89(ANN2) sunflower line was performed using high-throughput sequencing technologies. Analysis of CMS ANN2 mitochondrial DNA sequence revealed the following reorganization events: twelve rearrangements, seven insertions, and nine deletions. Comparisons of coding sequences from the male-sterile line with the male-fertile line identified a deletion of orf777 and seven new transcriptionally active open reading frames (ORFs): orf324, orf327, orf345, orf558, orf891, orf933, orf1197. Three of these ORFs represent chimeric genes involving atp6 (orf1197), cox2 (orf558), and nad6 (orf891). In addition, orf558, orf891, orf1197, as well as orf933, encode proteins containing membrane domain(s), making them the most likely candidate genes for CMS development in ANN2. Although the investigated CMS phenotype may be caused by simultaneous action of several candidate genes, we assume that orf1197 plays a major role in developing male sterility in ANN2. Comparative analysis of mitogenome organization in sunflower lines representing different CMS sources also allowed identification of reorganization hot spots in the mitochondrial genome of sunflower.

Mitochondrial genomes organization in alloplasmic lines of sunflower (Helianthus annuus L.) with various types of cytoplasmic male sterility.

BACKGROUND: Cytoplasmic male sterility (CMS) is a common phenotype in higher plants, that is often associated with rearrangements in mitochondrial DNA (mtDNA), and is widely used to produce hybrid seeds in a variety of valuable crop species. Investigation of the CMS phenomenon promotes understanding of fundamental issues of nuclear-cytoplasmic interactions in the ontogeny of higher plants. In the present study, we analyzed the structural changes in mitochondrial genomes of three alloplasmic lines of sunflower (Helianthus annuus L.). The investigation was focused on CMS line PET2, as there are very few reports about its mtDNA organization. METHODS: The NGS sequencing, de novo assembly, and annotation of sunflower mitochondrial genomes were performed. The comparative analysis of mtDNA of HA89 fertile line and two HA89 CMS lines (PET1, PET2) occurred. RESULTS: The mtDNA of the HA89 fertile line was almost identical to the HA412 line (NC_023337). The comparative analysis of HA89 fertile and CMS (PET1) analog mitochondrial genomes revealed 11,852 bp inversion, 4,732 bp insertion, 451 bp deletion and 18 variant sites. In the mtDNA of HA89 (PET2) CMS line we determined 27.5 kb and 106.5 kb translocations, 711 bp and 3,780 bp deletions, as well as, 5,050 bp and 15,885 bp insertions. There are also 83 polymorphic sites in the PET2 mitochondrial genome, as compared with the fertile line. DISCUSSION: The observed mitochondrial reorganizations in PET1 resulted in only one new open reading frame formation (orfH522), and PET2 mtDNA rearrangements led to the elimination of orf777, duplication of atp6 gene and appearance of four new ORFs with transcription activity specific for the HA89 (PET2) CMS line-orf645, orf2565, orf228 and orf285. Orf228 and orf285 are the atp9 chimeric ORFs, containing transmembrane domains and possibly may impact on mitochondrial membrane potential. So orf228 and orf285 may be the cause for the appearance of the PET2 CMS phenotype, while the contribution of other mtDNA reorganizations in CMS formation is negligible.