An empirical analysis of mtSSRs: could microsatellite distribution patterns explain the evolution of mitogenomes in plants?

Microsatellites (SSRs) are tandem repeat sequences in eukaryote genomes, including plant cytoplasmic genomes. The mitochondrial genome (mtDNA) has been shown to vary in size, number, and distribution of SSRs among different plant groups. Thus, SSRs contribute with genomic diversity in mtDNAs. However, the abundance, distribution, and evolutionary significance of SSRs in mtDNA from a wide range of algae and plants have not been explored. In this study, the mtDNAs of 204 plant and algal species were investigated related to the presence of SSRs. The number of SSRs was positively correlated with genome size. Its distribution is dependent on plant and algal groups analyzed, although the cluster analysis indicates the conservation of some common motifs in algal and terrestrial plants that reflect common ancestry of groups. Many SSRs in coding and non-coding regions can be useful for molecular markers. Moreover, mitochondrial SSRs are highly abundant, representing an important source for natural or induced genetic variation, i.e., for biotechnological approaches that can modulate mtDNA gene regulation. Thus, this comparative study increases the understanding of the plant and algal SSR evolution and brings perspectives for further studies.

Insights into the transcriptional and post-transcriptional regulation of the rice SUMOylation machinery and into the role of two rice SUMO proteases.

BACKGROUND: SUMOylation is an essential eukaryotic post-translation modification that, in plants, regulates numerous cellular processes, ranging from seed development to stress response. Using rice as a model crop plant, we searched for potential regulatory points that may influence the activity of the rice SUMOylation machinery genes. RESULTS: We analyzed the presence of putative cis-acting regulatory elements (CREs) within the promoter regions of the rice SUMOylation machinery genes and found CREs related to different cellular processes, including hormone signaling. We confirmed that the transcript levels of genes involved in target-SUMOylation, containing ABA- and GA-related CREs, are responsive to treatments with these hormones. Transcriptional analysis in Nipponbare (spp. japonica) and LC-93-4 (spp. indica), showed that the transcript levels of all studied genes are maintained in the two subspecies, under normal growth. OsSUMO3 is an exceptional case since it is expressed at low levels or is not detectable at all in LC-93-4 roots and shoots, respectively. We revealed post-transcriptional regulation by alternative splicing (AS) for all genes studied, except for SUMO coding genes, OsSIZ2, OsOTS3, and OsELS2. Some AS forms have the potential to alter protein domains and catalytic centers. We also performed the molecular and phenotypic characterization of T-DNA insertion lines of some of the genes under study. Knockouts of OsFUG1 and OsELS1 showed increased SUMOylation levels and non-overlapping phenotypes. The fug1 line showed a dwarf phenotype, and significant defects in fertility, seed weight, and panicle architecture, while the els1 line showed early flowering and decreased plant height. We suggest that OsELS1 is an ortholog of AtEsd4, which was also supported by our phylogenetic analysis. CONCLUSIONS: Overall, we provide a comprehensive analysis of the rice SUMOylation machinery and discuss possible effects of the regulation of these genes at the transcriptional and post-transcriptional level. We also contribute to the characterization of two rice SUMO proteases, OsELS1 and OsFUG1.

Molecular responses to recurrent drought in two contrasting rice genotypes.

MAIN CONCLUSION: The set of variables analyzed as integrated by multivariate analysis of principal components consistently showed a memory effect induced by the drought pre-treatment in AN Cambará plants. The effects of drought can vary ddepending on many factors. Among these the occurrence of a previous water stress may leave a residual effect (memory), influencing the future performance of a plant in response to a new drought event. This study tested the hypothesis that plants experiencing recurrent drought would show more active mechanisms of water deficit tolerance, mainly plants of the genotype that is cultivated often experiencing water shortages periods. Additionally, all the plants subjected to water deficit were rehydrated by 24 h and the expression of transcription factors related to drought responses was re-evaluated. To this end, the water status of two rice genotypes, BRS Querência (flooded) and AN Cambará (dryland), was evaluated to identify molecular alterations likely underpinning drought-memory. In growth stage V5, some plants were exposed to water stress (10% VWC soil moisture-pre-treatment). Thereafter, the pots were rehydrated at the same level as the control pots and maintained under this condition until drought was reapplied (10% VWC) at the reproductive stage (R1-R2). Then, the plants were rehydrated and maintained at pot capacity for 24 h. Overall, the set of variables analyzed integrally by multivariate analysis of principal components consistently showed a memory effect induced by the drought pre-treatment in AN Cambará plants (the dryland genotype). This conclusion, based on data of the biochemical and molecular analyses, was supported by the greater capacity of maintenance of the water status by stomatal regulation of the pre-treated and rehydrated plants after the second drought stimulus.

Comparative transcriptomics of rice plants under cold, iron, and salt stresses.

Abiotic stresses such as salinity, iron toxicity, and low temperatures are the main limiting factors of rice (Oryza sativa L.) yield. The elucidation of the genes involved in responses to these stresses is extremely important to understand the mechanisms that confer tolerance, as well as for the development of cultivars adapted to these conditions. In this study, the RNA-seq technique was used to compare the transcriptional profile of rice leaves (cv. BRS Querência) in stage V3, exposed to cold, iron, and salt stresses for 24 h. A range of 41 to 51 million reads was aligned, in which a total range of 88.47 to 89.21 % was mapped in the reference genome. For cold stress, 7905 differentially expressed genes (DEGs) were observed, 2092 for salt and 681 for iron stress; 370 of these were common to the three DEG stresses. Functional annotation by software MapMan demonstrated that cold stress usually promoted the greatest changes in the overall metabolism, and an enrichment analysis of overrepresented gene ontology (GO) terms showed that most of them are contained in plastids, ribosome, and chloroplasts. Saline stress induced a more complex interaction network of upregulated overrepresented GO terms with a relatively low number of genes compared with cold stress. Our study demonstrated a high number of differentially expressed genes under cold stress and a greater relationship between salt and iron stress levels. The physiological process most affected at the molecular level by the three stresses seems to be photosynthesis.

Phylogenetic relationships and selective pressure on gene families related to iron homeostasis in land plants.

Iron is involved in many metabolic processes, such as respiration and photosynthesis, and therefore an essential element for plant development. Comparative analysis of gene copies between crops and lower plant groups can shed light on the evolution of genes important to iron homeostasis. A phylogenetic analysis of five metal homeostasis gene families (NAS, NRAMP, YSL, FRO, and IRT) selected in monocots, dicots, gymnosperms, and bryophytes was performed. The homologous genes were found using known iron homeostasis gene sequences of Oryza sativa, Arabidopsis thaliana, and Physcomitrella patens as queries. The phylogeny was constructed using bioinfomatics tools. A total of 243 gene sequences for 30 plant species were found. The evolutionary fingerprint analysis suggested a purifying selective pressure of iron homeostasis genes for most of the plant gene homologues. The NAS and YSL genes appear to accumulate more negative selection sites, suggesting a strong selective pressure on these two gene families. The divergence time analysis indicates IRT as the most ancient gene family and FRO as the most recent. NRAMP and YSL genes appear to share a close relationship in the evolution of iron homeostasis gene families.

Genetic changes in the genus Vitis and the domestication of vine.

The genus Vitis belongs to the Vitaceae family and is divided into two subgenera: Muscadinia and Vitis, the main difference between these subgenera being the number of chromosomes. There are many hypotheses about the origin of the genus, which have been formed with archaeological studies and lately with molecular analyses. Even though there is no consensus on the place of origin, these studies have shown that grapes have been used by man since ancient times, starting later on its domestication. Most studies point to the Near East and Greece as the beginning of domestication, current research suggests it took place in parallel in different sites, but in all cases Vitis vinifera (L.) subsp. sylvestris [Vitis vinifera (L.) subsp. sylvestris (Gmelin) Hagi] seems to be the species chosen by our ancestors to give rise to the now known Vitis vinifera (L.) subsp. vinifera [=sativa (Hegi)= caucasica (Vavilov)]. Its evolution and expansion into other territories followed the formation of new empires and their expansion, and this is where the historical importance of this crop lies. In this process, plants with hermaphrodite flowers were preferentially selected, with firmer, sweeter, larger fruits of different colors, thus favoring the selection of genes associated with these traits, also resulting in a change in seed morphology. Currently, genetic improvement programs have made use of wild species for the introgression of disease resistance genes and tolerance to diverse soil and climate environments. In addition, the mapping of genes of interest, both linked to agronomic and fruit quality traits, has allowed the use of molecular markers for assisted selection. Information on the domestication process and genetic resources help to understand the gene pool available for the development of cultivars that respond to producer and consumer requirements.

Genetic Control of Reproductive Traits under Different Temperature Regimes in Inbred Line Populations Derived from Crosses between S. pimpinellifolium and S. lycopersicum Accessions.

In the present work, we study the genetic control of reproductive traits under different heat stress conditions in two populations of inbred lines derived from crosses between two S. pimpinellifolium accessions and two tomato cultivars (E9×L5 and E6203×LA1589). The temperature increase affected the reproductive traits, especially at extremely high temperatures, where only a few lines were able to set fruits. Even though a relative modest number of QTLs was identified, two clusters of QTLs involved in the responses of reproductive traits to heat stress were detected in both populations on chromosomes 1 and 2. Interestingly, several epistatic interactions were detected in the E9×L5 population, which were classified into three classes based on the allelic interaction: dominant (one locus suppressed the allelic effects of a second locus), co-adaptive (the double-homozygous alleles from the same parent alleles showed a higher phenotypic value than the combination of homozygous alleles from alternative parents) and transgressive (the combination of double-homozygous alleles from different parents showed better performance than double-homozygous alleles from the same parents). These results reinforce the important role of non-additive genetic variance in the response to heat stress and the potential of the new allelic combinations that arise after wide crosses.

Genetic diversity, linkage disequilibrium, and population structure in a panel of Brazilian rice accessions.

Narrowing of genetic diversity and the quantitative nature of most agronomic traits is a challenge for rice breeding. Genome-wide association studies have a great potential to identify important variation in loci underlying quantitative and complex traits; however, before performing the analysis, it is important to assess parameters of the genotypic data and population under study, to improve the accuracy of the genotype-phenotype associations. The aim of this study was to access the genetic diversity, linkage disequilibrium, and population structure of a working panel of Brazilian and several introduced rice accessions, which are currently being phenotyped for a vast number of traits to undergo association mapping. Ninety-four accessions were genotyped with 7098 SNPs, and after filtering for higher call rates and removing rare variants, 93 accessions and 4973 high-quality SNPs remained for subsequent analyses and association studies. The overall mean of the polymorphic information content, heterozygosity, and gene diversity of the SNPs was comparable to other rice panels. The r2 measure of linkage disequilibrium decayed to 0.25 in approximately 150 kb, a slow decay, explained by the autogamous nature of rice and the small size of the panel. Regarding population structure, eight groups were formed according to Bayesian clustering. Principle components and neighbor-joining analyses were able to distinguish part of the groups formed, mainly regarding the sub-species indica and japonica. Our results demonstrate that the population and SNPs are of high quality for association mapping.

Iron tolerance in rice: an efficient method for performing quick early genotype screening.

OBJECTIVES: This study was conducted to establish a method for early, quick and cheap screening of iron excess tolerance in rice (Oryza sativa L.) cultivars. RESULTS: Based on the experiments, iron excess leads to reduction in shoot length (SL) and this can be a useful characteristic for adequate screening of tolerant genotypes. The sensitive genotypes Nipponbare and BR-IRGA 409 indicated higher accumulation of iron in their tissues while BRS-Agrisul and Epagri 108 also accumulated iron, but at lower concentrations. BR-IRGA 410 displayed an intermediate phenotype regarding iron accumulation. No changes in shoot Cu content can be observed when comparing treatments. On the other hand, an increase was seen for Zn and Mn when shoots are subjected to Fe2+ excess. Fe stress at a lower concentration than 7 mM increased Zn but decreased Mn contents in shoots of BR-IRGA 409. Strong positive correlations were found here for Fe × Zn (0.93); Fe × Mn (0.97) and Zn × Mn (0.92), probably due to the Fe-induced activation of bivalent cation transporters. Results show that genotypes scored as sensitive present higher concentration of Fe in shoots and this is an efficient method to characterize rice cultivars regarding iron response.

Meta-Analysis of the QTLome of Fusarium Head Blight Resistance in Bread Wheat: Refining the Current Puzzle.

Background: Fusarium Head Blight (FHB) is a worldwide devastating disease of bread wheat (Triticum aestivum L.). Genetic resistance is the most effective way to control FHB and many QTL related to this trait have been mapped on the wheat genetic map. This information, however, must be refined to be more efficiently used in breeding programs and for the advance of the basic research. The objective of the present study was to in-depth analyze the QTLome of FHB resistance in bread wheat, further integrating genetic, genomic, and transcriptomic data, aiming to find candidate genes. Methods: An exhaustive bibliographic review on 76 scientific papers was carried out collecting information about QTL related to FHB resistance mapped on bread wheat. A dense genetic consensus map with 572,862 loci was generated for QTL projection. Meta-analysis could be performed on 323 QTL. Candidate gene mining was carried out within the most refined loci, containing genes that were cross-validated with publicly available transcriptional expression data of wheat under Fusarium infection. Most highlighted genes were investigated for protein evidence. Results: A total of 556 QTL were found in the literature, distributed on all sub-genomes and chromosomes of wheat. Meta-analysis generated 65 meta-QTL, and this refinement allows one to find markers more tightly linked to these regions. Candidate gene mining within the most refined meta-QTL, meta-QTL 1/chr. 3B, harvested 324 genes and transcriptional data cross-validated 10 of these genes, as responsive to FHB. One is of these genes encodes a Glycosiltransferase and the other encodes for a Cytochrome P450, and these such proteins have already been verified as being responsible for FHB resistance, but the remaining eight genes still have to be further studied, as promising loci for breeding. Conclusions: The QTLome of FHB resistance in wheat was successfully assembled and a refinement in terms of number and length of loci was obtained. The integration of the QTLome with genomic and transcriptomic data has allowed for the discovery of promising candidate genes for use in breeding programs.

Activation of rice WRKY transcription factors: an army of stress fighting soldiers?

Rice WRKYs comprise a large family of transcription factors and present remarkable structure features and a unique DNA binding site. Their importance in plants goes beyond the response to stressful stimuli, since they participate in hormonal pathways and developmental processes. Indeed, the majority of WRKYs present an independent activation since they are able to perform self-transcriptional regulation. However, some WRKY activation depends on epigenetic and transcript regulation by micro RNAs. Their protein function depends, almost always, on the posttranslational changes. Taking to account its properties of auto-activation, all these regulators process are extremely important for complete WRKY regulation. In this sense, here we provide an overview of transcriptional activation and posttranscriptional and posttranslational regulation of rice WRKY genes under stresses.

Publisher Correction: Genomes of 13 domesticated and wild rice relatives highlight genetic conservation, turnover and innovation across the genus Oryza.

This article was not made open access when initially published online, which was corrected before print publication. In addition, ORCID links were missing for 12 authors and have been added to the HTML and PDF versions of the article.

Genomes of 13 domesticated and wild rice relatives highlight genetic conservation, turnover and innovation across the genus Oryza.

The genus Oryza is a model system for the study of molecular evolution over time scales ranging from a few thousand to 15 million years. Using 13 reference genomes spanning the Oryza species tree, we show that despite few large-scale chromosomal rearrangements rapid species diversification is mirrored by lineage-specific emergence and turnover of many novel elements, including transposons, and potential new coding and noncoding genes. Our study resolves controversial areas of the Oryza phylogeny, showing a complex history of introgression among different chromosomes in the young 'AA' subclade containing the two domesticated species. This study highlights the prevalence of functionally coupled disease resistance genes and identifies many new haplotypes of potential use for future crop protection. Finally, this study marks a milestone in modern rice research with the release of a complete long-read assembly of IR 8 'Miracle Rice', which relieved famine and drove the Green Revolution in Asia 50 years ago.

Evaluation of stability and validation of reference genes for RT-qPCR expression studies in rice plants under water deficit.

Many studies use strategies that allow for the identification of a large number of genes expressed in response to different stress conditions to which the plant is subjected throughout its cycle. In order to obtain accurate and reliable results in gene expression studies, it is necessary to use reference genes, which must have uniform expression in the majority of cells in the organism studied. RNA isolation of leaves and expression analysis in real-time quantitative polymerase chain reaction (RT-qPCR) were carried out. In this study, nine candidate reference genes were tested, actin 11 (ACT11), ubiquitin conjugated to E2 enzyme (UBC-E2), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), beta tubulin (ß-tubulin), eukaryotic initiation factor 4α (eIF-4α), ubiquitin 10 (UBQ10), ubiquitin 5 (UBQ5), aquaporin TIP41 (TIP41-Like) and cyclophilin, in two genotypes of rice, AN Cambará and BRS Querência, with different levels of soil moisture (20%, 10% and recovery) in the vegetative (V5) and reproductive stages (period preceding flowering). Currently, there are different softwares that perform stability analyses and define the most suitable reference genes for a particular study. In this study, we used five different methods: geNorm, BestKeeper, ΔCt method, NormFinder and RefFinder. The results indicate that UBC-E2 and UBQ5 can be used as reference genes in all samples and softwares evaluated. The genes ß-tubulin and eIF-4α, traditionally used as reference genes, along with GAPDH, presented lower stability values. The gene expression of basic leucine zipper (bZIP23 and bZIP72) was used to validate the selected reference genes, demonstrating that the use of an inappropriate reference can induce erroneous results.

Tandem repeat distribution of gene transcripts in three plant families.

Tandem repeats (microsatellites or SSRs) are molecular markers with great potential for plant genetic studies. Modern strategies include the transfer of these markers among widely studied and orphan species. In silico analyses allow for studying distribution patterns of microsatellites and predicting which motifs would be more amenable to interspecies transfer. Transcribed sequences (Unigene) from ten species of three plant families were surveyed for the occurrence of micro and minisatellites. Transcripts from different species displayed different rates of tandem repeat occurrence, ranging from 1.47% to 11.28%. Both similar and different patterns were found within and among plant families. The results also indicate a lack of association between genome size and tandem repeat fractions in expressed regions. The conservation of motifs among species and its implication on genome evolution and dynamics are discussed.

SSR Locator: Tool for Simple Sequence Repeat Discovery Integrated with Primer Design and PCR Simulation.

Microsatellites or SSRs (simple sequence repeats) are ubiquitous short tandem duplications occurring in eukaryotic organisms. These sequences are among the best marker technologies applied in plant genetics and breeding. The abundant genomic, BAC, and EST sequences available in databases allow the survey regarding presence and location of SSR loci. Additional information concerning primer sequences is also the target of plant geneticists and breeders. In this paper, we describe a utility that integrates SSR searches, frequency of occurrence of motifs and arrangements, primer design, and PCR simulation against other databases. This simulation allows the performance of global alignments and identity and homology searches between different amplified sequences, that is, amplicons. In order to validate the tool functions, SSR discovery searches were performed in a database containing 28 469 nonredundant rice cDNA sequences.