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Phospholipases find versatile applications across industries, including detergent production, food modification, pharmaceuticals (especially in drug delivery systems), and cell signaling research. In this study, we present a strain of Bacillus paranthracis for the first time, demonstrating significant potential in the production of phosphatidylcholine-specific phospholipase C (PC-PLC). The investigation thoroughly examines the B. paranthracis PUMB_17 strain, focusing on the activity of PC-PLC and its purification process. Notably, the PUMB_17 strain displays extracellular PC-PLC production with high specific activity during the late exponential growth phase. To unravel the genetic makeup of PUMB_17, we employed nanopore-based whole-genome sequencing and subsequently conducted a detailed genome annotation. The genome comprises a solitary circular chromosome spanning 5,250,970 bp, featuring a guanine-cytosine ratio of 35.49. Additionally, two plasmids of sizes 64,250 bp and 5845 bp were identified. The annotation analysis reveals the presence of 5328 genes, encompassing 5186 protein-coding sequences, and 142 RNA genes, including 39 rRNAs, 103 tRNAs, and 5 ncRNAs. The aim of this study was to make a comprehensive genomic exploration that promises to enhance our understanding of the previously understudied and recently documented capabilities of Bacillus paranthracis and to shed light on a potential use of the strain in the industrial production of PC-PLC.
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The endometrium, the inner mucosal lining of the uterus, undergoes complex molecular and cellular changes across the menstrual cycle in preparation for embryo implantation. Transcriptome-wide analyses have mainly been utilized to study endometrial receptivity, the prerequisite for successful implantation, with most studies, so far, comparing the endometrial transcriptomes between (i) secretory and proliferative endometrium or (ii) mid-secretory and early secretory endometrium. In the current study, we provide a complete transcriptome description of the endometrium across the entire menstrual cycle and, for the first time, comprehensively characterize the proliferative phase of the endometrium. Our temporal transcriptome analysis includes five time points including the mid-proliferative, late proliferative (peri-ovulatory phase), early secretory, mid-secretory, and late secretory phases. Thus, we unveil exhaustively the transitions between the consecutive proliferative and secretory phases, highlighting their unique gene expression profiles and possible distinct biological functions. The transcriptome analysis reveals many differentially expressed genes (DEGs) across the menstrual cycle, most of which are phase-specific. As an example of coordinated gene activity, the expression profile of histone-encoding genes within the HIST cluster on chromosome 6 shows an increase in cluster activity during the late proliferative and a decline during the mid-secretory phase. Moreover, numerous DEGs are shared among all phases. In conclusion, in the current study, we delineate the endometrial proliferative phase-centered view of transcriptome dynamics across the menstrual cycle. Our data analysis highlights significant transcriptomic and functional changes occurring during the late proliferative phase-an essential transition point from the proliferative phase to the secretory phase. Future studies should explore how the biology of the late proliferative phase endometrium impacts the achievement of mid-secretory endometrial receptivity or contributes to molecular aberrations leading to embryo implantation failure.
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Endométrio , Perfilação da Expressão Gênica , Ciclo Menstrual , Transcriptoma , Feminino , Humanos , Endométrio/metabolismo , Ciclo Menstrual/genética , AdultoRESUMO
Non-coding RNAs (ncRNAs), such as microRNAs (miRNAs), short interfering RNAs (siRNAs), and long non-coding RNAs (lncRNAs), have emerged as pivotal regulators within the plant kingdom [...].
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MicroRNAs , RNA Longo não Codificante , MicroRNAs/genética , RNA Longo não Codificante/genética , RNA Interferente Pequeno/genéticaRESUMO
Research background: Sourdough is a spontaneously formed, complex microbial ecosystem of various lactic acid bacteria (LAB) and yeast which, by producing specific metabolites, determines the quality of the baked products. In order to design and control the sourdough with preferred nutritional characteristics, it is crucial that the LAB diversity of the product of interest be elucidated. Experimental approach: Using the opportunities of next-generation sequencing (NGS) of the V1-V3 hypervariable gene region of 16S rRNA, we studied the microbial ecosystem of a whole grain sourdough made of Triticum monococcum, originating from Southwestern Bulgaria. Since the DNA extraction method is considered crucial for the accuracy of the sequencing results, as it can introduce significant differences in the examined microbiota, we used three different commercial kits for DNA isolation and analyzed their impact on the observed bacterial diversity. Results and conclusions: All three DNA extraction kits provided bacterial DNA which passed quality control and was successfully sequenced on Illumina MiSeq platform. The results received from the different DNA protocols showed variations in the microbial profiles. Alpha diversity indices (ACE, Chao1, Shannon, and Simpson) were also different among the three groups of results. Nevertheless, a strong dominance of phylum Firmicutes, class Bacilli, order Lactobacillales, represented mostly by family Lactobacillaceae, genus Lactobacillus (relative abundance of 63.11-82.28%) and family Leuconostocaceae, genus Weissella (relative abundance of 3.67-36.31%) was observed. Lactiplantibacillus plantarum and Levilactobacillus brevis with relative abundance of 16.15-31.24% and 6.21-16.29% respectively, were the two dominant species identified in all three DNA isolates. Novelty and scientific contribution: The presented results give insight into the taxonomic composition of bacterial community of a specific Bulgarian sourdough. Having in mind that the sourdough is a difficult matrix for DNA isolation on the one hand, and that there is no standardized DNA extraction protocol for this matrix on the other hand, this pilot study aims to give a small contribution to the future establishment and validation of such a protocol, which will allow accurate assessment of the specific microbiota of sourdough samples.
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Plant small RNAs (sRNAs) are a heterogeneous group of noncoding RNAs with a length of 20-24 nucleotides that are widely studied due to their importance as major regulators in various biological processes. sRNAs are divided into two main classes-microRNAs (miRNAs) and small interfering RNAs (siRNAs)-which differ in their biogenesis and functional pathways. Their identification and enrichment with new structural variants would not be possible without the use of various high-throughput sequencing (NGS) techniques, allowing for the detection of the total population of sRNAs in plants. Classifying sRNAs and predicting their functional role based on such high-performance datasets is a nontrivial bioinformatics task, as plants can generate millions of sRNAs from a variety of biosynthetic pathways. Over the years, many computing tools have been developed to meet this challenge. Here, we review more than 35 tools developed specifically for plant sRNAs over the past few years and explore some of their basic algorithms for performing tasks related to predicting, identifying, categorizing, and quantifying individual sRNAs in plant samples, as well as visualizing the results of these analyzes. We believe that this review will be practical for biologists who want to analyze their plant sRNA datasets but are overwhelmed by the number of tools available, thus answering the basic question of how to choose the right one for a particular study.
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Biologia Computacional , MicroRNAs , Biologia Computacional/métodos , Regulação da Expressão Gênica de Plantas , Sequenciamento de Nucleotídeos em Larga Escala , MicroRNAs/metabolismo , Plantas/genética , Plantas/metabolismo , RNA de Plantas/metabolismo , RNA Interferente Pequeno/metabolismoRESUMO
Microorganisms inhabiting freshwater environments are an integral part of the aquatic ecosystems. Very few data are available regarding the profiles of the microbial communities in the reservoirs in Bulgaria, despite their key role in the biogeochemical processes. In the present study, we provide the first comprehensive metagenomic analysis on the planktonic bacterial diversity of two large and economically important Bulgarian reservoirs (Batak and Tsankov Kamak) using next-generation sequencing of 16S ribosomal RNA gene (16S rRNA). Analysis of the metagenomic amplicon datasets, including quality filtering, clustering of Operational Taxonomic Units and taxonomy assignment revealed that 78.45% of the microbial communities between the two reservoirs were overlapping. The diversity (H) and Pielou's evenness (J) indices declined along the longitudinal axis of both reservoirs. The estimated values for the Shannon diversity index are typically observed in oligotrophic lakes. The microbial communities of both reservoirs were dominated by Proteobacteria, followed by Actinobacteria and Bacteroidetes all comprised over 95% of the relative abundance, regardless of the reservoir's large hydrogeological differences. The bacterioplankton was characterized by high phylogenetic heterogeneity in the taxonomic structure, being distributed among 211 genera. The genera Limnohabitans and Rhodoferax held the absolute predominance, implying their significance in the aquatic food webs. The obtained data can contribute to the better systematic understanding of the microbial diversity of freshwater environments.
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Bactérias/genética , Água Doce/microbiologia , Metagenômica , Consórcios Microbianos , Plâncton/genética , Actinobacteria/classificação , Actinobacteria/genética , Bactérias/classificação , Bacteroidetes/classificação , Bacteroidetes/genética , Biodiversidade , Bulgária , Ecossistema , Sequenciamento de Nucleotídeos em Larga Escala , Lagos/microbiologia , Filogenia , Plâncton/classificação , Proteobactérias/classificação , Proteobactérias/genética , RNA Ribossômico 16S/genética , Análise de Sequência de DNARESUMO
We conducted a comprehensive analysis of codon usage bias (CUB) based on the available non-redundant full-length cDNA (nrFLcDNA) and expressed sequence tags (ESTs) data of cultivar Micro-Tom and evaluated the associations of observed CUB and measurements of transcriptional and translational effectiveness. The analysis presented in our study suggests a correlation, which is negative but highly correlated between Axis 1 and GC3s (r=-0.827, P<0.01), indicating that mutational bias has a significant and dominant repressive role to the choices of GC3. We also observed a strong positive correlation between codon adaptation index (CAI) and translational adaptation index (tAIg) (0.407, P<0.01), which demonstrates the facilitation of efficient translation by the optimal codon usage patterns of the highly expressed genes. We believe that the complete set of optimal codon usage patterns detected in this study will serve as a model to enhance the transgenesis in the studied cultivar of Solanum lycopersicum.
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Códon , Biossíntese de Proteínas , Solanum lycopersicum/genética , Composição de Bases , Sequência de Bases , Evolução Molecular , Etiquetas de Sequências Expressas , Expressão Gênica , Modelos Genéticos , Anotação de Sequência Molecular , Análise Multivariada , Mutação , Fases de Leitura Aberta , Proteínas de Plantas/genética , Plantas Geneticamente ModificadasRESUMO
Mosses host diverse bacterial communities essential for their fitness, nutrient acquisition, stress tolerance, and pathogen defense. Understanding the microbiome's taxonomic composition is the first step, but unraveling their functional capabilities is crucial for grasping their ecological significance. Metagenomics characterizes microbial communities by composition, while metatranscriptomics explores gene expression, providing insights into microbiome functionality beyond the structure. Here, we present for the first time a metatranscriptomic study of two moss species, Hypnum cupressiforme (Hedw.) and Platyhypnidium riparioides (Hedw.) Dixon., renowned as key biomonitors of atmospheric and water pollution. Our investigation extends beyond taxonomic profiling and offers a profound exploration of moss bacterial communities. Pseudomonadota and Actinobacteria are the dominant bacterial phyla in both moss species, but their proportions differ. In H. cupressiforme, Actinobacteria make up 62.45% and Pseudomonadota 32.48%, while in P. riparioides, Actinobacteria account for only 25.67% and Pseudomonadota 69.08%. This phylum-level contrast is reflected in genus-level differences. Our study also shows the expression of most genes related to nitrogen cycling across both microbiomes. Additionally, functional annotation highlights disparities in pathway prevalence, including carbon dioxide fixation, photosynthesis, and fatty acid biosynthesis, among others. These findings hint at potential metabolic distinctions between microbial communities associated with different moss species, influenced by their specific genotypes and habitats. The integration of metatranscriptomic data holds promise for enhancing our understanding of bryophyte-microbe partnerships, opening avenues for novel applications in conservation, bioremediation, and sustainable agriculture.
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STUDY QUESTION: What changes occur in the endometrium during aging, and do they impact fertility? SUMMARY ANSWER: Both the transcriptome and cellular composition of endometrial samples from women of advanced maternal age (AMA) are significantly different from that of samples from young women, suggesting specific changes in epithelial cells that may affect endometrial receptivity. WHAT IS KNOWN ALREADY: Aging is associated with the accumulation of senescent cells in aging tissues. Reproductive aging is mostly attributed to the decline in ovarian reserve and oocyte quality, whereas the endometrium is a unique complex tissue that is monthly renewed under hormonal regulation. Several clinical studies have reported lower implantation and pregnancy rates in oocyte recipients of AMA during IVF. Molecular studies have indicated the presence of specific mutations within the epithelial cells of AMA endometrium, along with altered gene expression of bulk endometrial tissue. STUDY DESIGN SIZE DURATION: Endometrial transcriptome profiling was performed for 44 women undergoing HRT during the assessment of endometrial receptivity before IVF. Patients younger than 28 years were considered as the young maternal age (YMA) group (age 23-27 years) and women older than 45 years were considered as the AMA group (age 47-50 years). Endometrial biopsies were obtained on Day 5 of progesterone treatment and RNA was extracted. All endometrial samples were evaluated as being receptive based on the expression of 68 common endometrial receptivity markers. Endometrial samples from another 24 women classified into four age groups (YMA, intermediate age group 1 (IMA1, age 29-35), intermediate age group 2 (IMA2, age 36-44), and AMA) were obtained in the mid-secretory stage of a natural cycle (NC) and used for validation studies across the reproductive lifespan. PARTICIPANTS/MATERIALS SETTING METHODS: A total of 24 HRT samples (12 YMA and 12 AMA) were subject to RNA sequencing (RNA-seq) and differential gene expression analysis, 20 samples (10 YMA and 10 AMA) were used for qPCR validation, and 24 NC samples (6 YMA, 6 IMA1, 6 IMA2 and 6AMA) were used for RNA-seq validation of AMA genes across the woman's reproductive lifespan. Immunohistochemistry (IHC) was used to confirm some expression changes at the protein level. Computational deconvolution using six endometrial cell type-specific transcriptomic profiles was conducted to compare the cellular composition between the groups. MAIN RESULTS AND THE ROLE OF CHANCE: Comparisons between YMA and AMA samples identified a lower proportion of receptive endometria in the AMA group (P = 0.007). Gene expression profiling identified 491 differentially expressed age-sensitive genes (P adj < 0.05) that revealed the effects of age on endometrial epithelial growth and receptivity, likely contributing to decreased reproductive performance. Our results indicate that changes in the expression of the cellular senescence marker p16INK4a and genes associated with metabolism, inflammation, and hormone response are involved in endometrial aging. Importantly, we demonstrate that the proportion of multi-ciliated cells, as discovered based on RNA-seq data deconvolution and tissue IHC results, is affected by endometrial aging, and propose a putative onset of age-related changes. Furthermore, we propose that aging has an impact on the transcriptomic profile of endometrial tissue in the context of endometrial receptivity. LARGE SCALE DATA: The raw sequencing data reported in this article are deposited at the Gene Expression Omnibus under accession code GSE236128. LIMITATIONS REASONS FOR CAUTION: This retrospective study identified changes in the endometrium of patients undergoing hormonal replacement and validated these changes using samples obtained during a NC. However, future studies must clarify the importance of these findings on the clinical outcomes of assisted reproduction. WIDER IMPLICATIONS OF THE FINDINGS: The findings reported in this study have important implications for devising future strategies aimed at improving fertility management in women of advanced reproductive age. STUDY FUNDING/COMPETING INTERESTS: This research was funded by the Estonian Research Council (grant no. PRG1076), Horizon 2020 innovation grant (ERIN, grant no. EU952516), Enterprise Estonia (grant no. EU48695), MSCA-RISE-2020 project TRENDO (grant no. 101008193), EU 874867 project HUTER, the Horizon Europe NESTOR grant (grant no. 101120075) of the European Commission, the EVA specialty program (grant no. KP111513) of the Maastricht University Medical Center (MUMC+), MICIU/AEI/10.13039/501100011033 and FEDER, EU projects Endo-Map (grant no. PID2021-12728OB-100), ROSY (grant no. CNS2022-135999), and the National Science Fund of Bulgaria (grant no. KII-06 H31/2). The authors declare no competing interests.
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MicroRNAs (miRNAs) are a class of small non-coding RNAs, which are negative regulators of gene expression. Many genes in human uterine leiomyoma (ULM) are aberrantly expressed and in some cases this can be due to dysregulation of miRNAs. Here we present the first study to determine genome-wide miRNA expression patterns in uterine leiomyoma and myometrium using Solexa high-throughput sequencing. We found more than 50 miRNAs, which were differentially expressed, and furthermore we extend the list of putative new miRNA genes. The top five significantly de-regulated miRNAs in ULMs that we found in our libraries were miR-363, miR-490, miR-137, miR-217 and miR-4792. We also observed "isomiRs" with higher copy number than referenced mature miRNA specific for the leiomyoma libraries, which have a potential role in tumorigenesis. The microRNA transcriptomes obtained in this study deliver insights and further expand our understanding the role of small RNAs in uterine leiomyoma development.
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Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Leiomioma/genética , MicroRNAs/genética , Neoplasias Uterinas/genética , Sequência de Bases , DNA Complementar/química , DNA Complementar/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Biblioteca Gênica , Humanos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA/métodos , Homologia de Sequência do Ácido Nucleico , TranscriptomaRESUMO
Since the discovery of the Bence Jones protein in the middle to late 1800s and the subsequent identification of the carcinoembryonic antigen and alpha-fetoprotein in the 1970s, it has been demonstrated that the analysis of biofluids is essential to the diagnostic and follow-up processes of cancer [...].
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Lactiplantibacillus plantarum stands out as a remarkably diverse species of lactic acid bacteria, occupying a myriad of ecological niches. Particularly noteworthy is its presence in human breast milk, which can serve as a reservoir of probiotic bacteria, contributing significantly to the establishment and constitution of infant gut microbiota. In light of this, our study attempted to conduct an initial investigation encompassing both genomic and phenotypic aspects of the L. plantarum PU3 strain, that holds potential as a probiotic agent. By employing the cutting-edge third-generation Nanopore sequencing technology, L. plantarum PU3 revealed a circular chromosome of 3,180,940 bp and nine plasmids of various lengths. The L. plantarum PU3 genome has a total of 2962 protein-coding and non-coding genes. Our in-depth investigations revealed more than 150 probiotic gene markers that unfold the genetic determinants for acid tolerance, bile resistance, adhesion, and oxidative and osmotic stress. The in vivo analysis showed the strain's proficiency in utilizing various carbohydrates as growth substrates, complementing the in silico analysis of the genes involved in metabolic pathways. Notably, the strain demonstrated a pronounced affinity for D-sorbitol, D-mannitol, and D-Gluconic acid, among other carbohydrate sources. The in vitro experimental verification of acid, osmotic and bile tolerance validated the robustness of the strain in challenging environments. Encouragingly, no virulence factors were detected in the genome of PU3, suggesting its safety profile. In search of beneficial properties, we found potential bacteriocin biosynthesis clusters, suggesting its capability for antimicrobial activity. The characteristics exhibited by L. plantarum PU3 pave the way for promising strain potential, warranting further investigations to unlock its full capacity and contributions to probiotic and therapeutic avenues.
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Sourdoughs (SDs) are spontaneously formed microbial ecosystems composed of various species of lactic acid bacteria (LAB) and acid-tolerant yeasts in food matrices of cereal flours mixed with water. To date, more than 90 LAB species have been isolated, significantly impacting the organoleptic characteristics, shelf life, and health properties of bakery products. To learn more about the unique bacterial communities involved in creating regional Bulgarian sourdoughs, we examined the metacommunities of five sourdoughs produced by spontaneous fermentation and maintained by backslopping in bakeries from three geographic locations. The 16S rRNA gene amplicon sequencing showed that the former genus Lactobacillus was predominant in the studied sourdoughs (51.0-78.9%). Weissella (0.9-42.8%), Herbaspirillum (1.6-3.8%), Serratia (0.1-11.7%), Pediococcus (0.2-7.5%), Bacteroides (0.1-1.3%), and Sphingomonas (0.1-0.5%) were also found in all 5 samples. Genera Leuconostoc, Enterococcus, Bacillus, and Asaia were sample-specific. It is interesting to note that the genus Weissella was more abundant in wholegrain samples. The greatest diversity at the species level was found in the former genus Lactobacillus, presented in the sourdough samples with 13 species. The UPGMA cluster analysis clearly demonstrated similarity in species' relative abundance between samples from the same location. In addition, we can conclude that the presence of two main clusters-one including samples from mountainous places (the cities of Smolyan and Bansko) and the other including samples from the city of Ruse (the banks of the Danube River)-may indicate the impact of climate and geographic location (e.g., terrain, elevation, land use, and nearby water bodies and their streams) on the abundance of microbiome taxa. As the bacterial population is crucial for bread standardization, we expect the local bakery sector to be interested in the relationship between process variables and their effect on bacterial dynamics described in this research study.
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Plant microRNAs (miRNAs) are single-stranded 20-22 nt small RNAs (sRNA) that are produced from their own genes. We have developed a de novo genome-wide approach for the computational identification of novel plant miRNAs based on the integration of the complete genome sequence with sRNA libraries. It comprises three modules - the clustering module identifies genomic regions that have two closely-located unidirectional sRNA clusters, the mirplan module explores the secondary structure of the genomic regions, and the duplex module predicts miRNA/miRNA* duplexes. We applied our approach to the Brachypodium genome and publicly available sRNA libraries and predicted 102 miRNAs. Our results extend the list of known miRNAs with 58 novel miRNAs and define the genomic loci of all predicted miRNAs. Because this approach considers specific features of plant miRNAs, it can be employed for the analysis of the genome and sRNA libraries generated for plant species to achieve systematic miRNA discovery.
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Brachypodium/genética , Mapeamento Cromossômico/métodos , Biologia Computacional/métodos , Genoma de Planta/genética , MicroRNAs/genética , Sequência de Bases , Biblioteca Gênica , MicroRNAs/química , Dados de Sequência Molecular , Análise de Sequência de DNARESUMO
The human endometrium is a highly dynamic tissue. Increasing evidence has shown that microRNAs (miRs) play essential roles in human endometrium development. Our previous assay, based on small RNA-sequencing (sRNA-seq) indicated the complexity and dynamics of numerous sequence variants of miRs (isomiRs) that can act together to control genes of functional relevance to the receptive endometrium (RE). Here, we used a greater average depth of sRNA-seq to detect poorly expressed small RNAs. The sequencing data confirmed the up-regulation of miR-449c and uncovered other members of the miR-449 family up-regulated in RE-among them miR-449a, as well as several isoforms of both miR-449a and miR-449c, while the third family member, miR-449b, was not identified. Stem-looped RT-qPCR analysis of miR expression at four-time points of the endometrial cycle verified the increased expression of the miR-449a/c family members in RE, among which the 5' isoform of miR-449c-miR-449c.1 was the most strongly up-regulated. Moreover, we found in a case study that the expression of miR-449c.1 and its precursor correlated with the histological assessment of the endometrial phase and patient age. We believe this study will promote the clinical investigation and application of the miR-449 family in the diagnosis and prognosis of human reproductive diseases.
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Tools for microRNA (miR) sequencing data analyses are broadly used in biomedical research. However, the complexity of computational approaches still remains a challenge for biologists with scarce experience in data analytics and bioinformatics. Here, we present miRGalaxy, a Galaxy-based framework for comprehensive analysis of miRs and their sequence variants-miR isoforms (isomiRs). Though isomiRs are commonly reported in deep-sequencing experiments, their detailed structure complexity and specific differential expression (DE) remain not fully examined by the majority of the available analysis tools. miRGalaxy encompasses biologist-user-friendly tools and workflows dedicated to the analysis of the isomiR-ome and its complex behavior in various biological samples. miRGalaxy is developed as a modular, accessible, redistributable, shareable, and user-friendly framework for scientists working with small RNA (sRNA)-seq data. Due to its modular workflow, advanced users can customize the steps and tools for their needs. In addition, the framework provides an analysis report where the significant output results are summarized in charts and visualizations. miRGalaxy can be accessed via preconfigured Docker image flavor and a Toolshed installation if the user already has a running Galaxy instance. Over the last decade, studies on the expression of miRs and isomiRs in normal and deregulated tissues have led to the discovery of their potential as diagnostic biomarkers. The detection of miRs in biofluids further expanded the exploration of the miR repertoire as a source of liquid biopsy biomarkers. Here we show the miRGalaxy framework application for in-depth analysis of the sRNA-seq data from two different biofluids, milk and plasma, to identify, annotate, and discover specific differentially expressed miRs and isomiRs.
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Potato spindle tuber viroid (PSTVd) infects various plants. PSTVd pathogenesis is associated with interference with the cellular metabolism and defense signaling pathways via direct interaction with host factors or via the transcriptional or post-transcriptional modulation of gene expression. To better understand host defense mechanisms to PSTVd infection, we analyzed the gene expression in two pepper cultivars, Capsicum annuum Kurtovska kapia (KK) and Djulunska shipka (DS), which exhibit mild symptoms of PSTVd infection. Deep sequencing-based transcriptome analysis revealed differential gene expression upon infection, with some genes displaying contrasting expression patterns in KK and DS plants. More genes were downregulated in DS plants upon infection than in KK plants, which could underlie the more severe symptoms seen in DS plants. Gene ontology enrichment analysis revealed that most of the downregulated differentially expressed genes in both cultivars were enriched in the gene ontology term photosynthesis. The genes upregulated in DS plants fell in the biological process of gene ontology term defense response. We validated the expression of six overlapping differentially expressed genes that are involved in photosynthesis, plant hormone signaling, and defense pathways by quantitative polymerase chain reaction. The observed differences in the responses of the two cultivars to PSTVd infection expand the understanding of the fine-tuning of plant gene expression that is needed to overcome the infection.
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Haberlea rhodopensis is a paleolithic tertiary relict species that belongs to the unique group of resurrection plants sharing remarkable tolerance to desiccation. When exposed to severe drought stress, this species shows an ability to maintain structural integrity of its deactivated photosynthetic apparatus, which easily reactivates upon rehydration. In addition to its homoiochlorophyllous nature, the resurrection capability of H. rhodopensis is of particular importance to the global climate change mitigation. In this study, we sequenced, assembled, and analyzed the mitochondrial (mt) genome of H. rhodopensis for the first time. The master circle has a typical circular structure of 484 138 bp in length with a 44.1% GC content in total. The mt genome of H. rhodopensis contains 59 genes in total, including 35 protein-coding, 21 tRNAs, and 3 rRNAs genes. 7 tandem repeats and 85 simple sequence repeats (SSRs) are distributed throughout the mt genome. The alignment of 20 plant mt genomes confirms the phylogenetic position of H. rhodopensis in the Lamiales order. Our comprehensive analysis of the complete mt genome of H. rhodopensis is a significant addition to the limited database of organelle genomes of resurrection species. Comparative and phylogenetic analysis provides valuable information for a better understanding of mitochondrial molecular evolution in plants.
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Craterostigma/genética , Genoma Mitocondrial , Craterostigma/metabolismo , Desidratação/metabolismo , Secas , Genes de Plantas , Lamiales/genética , Lamiales/metabolismo , Fotossíntese , Filogenia , Folhas de Planta/metabolismo , Proteínas de Plantas/genética , Sequências de Repetição em Tandem , ÁguaRESUMO
Embryo implantation depends on endometrial receptivity (ER). To achieve ER, the preparation of the uterine lining requires controlled priming by ovarian hormones and the expression of numerous genes in the endometrial tissue. microRNAs (miRs) have emerged as critical genetic regulators of ER in fertility and of the diseases that are associated with infertility. With the rapid development of next-generation sequencing technologies, it has become clear that miR genes can produce canonical miRs and variants-isomiRs. Here, we describe miR/isomiR expression dynamics across the four time points of natural chorionic gonadotropin (hCG)-administered cycles. Sequencing of the small RNAs (sRNA-seq) revealed that the most significant expression changes during the transition from the pre-receptive to the receptive phase occurred in the isomiR families of miR-125a, miR-125b, miR-10a, miR-10b, miR-449c, miR-92a, miR-92b, and miR-99a. Pairing the analysis of the differentially expressed (DE) miRs/isomiRs and their predicted DE mRNA targets uncovered 280 negatively correlating pairs. In the receptive endometrium, the 5'3'-isomiRs of miR-449c, which were among the most highly up-regulated isomiRs, showed a negative correlation with their target, transcription factor (TF) MYCN, which was down-regulated. Joint analysis of the miR/isomiR and TF expression identified several regulatory interactions. Based on these data, a regulatory TF-miR/isomiR gene-target circuit including let7g-5p and miR-345; the isomiR families of miR-10a, miR-10b, miR-92a, and miR-449c; and MYCN and TWIST1 was proposed to play a key role in the establishment of ER. Our work uncovers the complexity and dynamics of the endometrial isomiRs that can act cooperatively with miRs to control the functionally important genes that are critical to ER. Further studies of miR/isomiR expression patterns that are paired with those of their target mRNAs may provide a more in-depth picture of the endometrial pathologies that are associated with implantation failure.
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Numerous studies on microRNAs (miRNA) in cancer and other diseases have been accompanied by diverse computational approaches and experimental methods to predict and validate miRNA biological and clinical significance as easily accessible disease biomarkers. In recent years, the application of the next-generation deep sequencing for the analysis and discovery of novel RNA biomarkers has clearly shown an expanding repertoire of diverse sequence variants of mature miRNAs, or isomiRs, resulting from alternative post-transcriptional processing events, and affected by (patho)physiological changes, population origin, individual's gender, and age. Here, we provide an in-depth overview of currently available bioinformatics approaches for the detection and visualization of both mature miRNA and cognate isomiR sequences. An attempt has been made to present in a systematic way the advantages and downsides of in silico approaches in terms of their sensitivity and accuracy performance, as well as used methods, workflows, and processing steps, and end output dataset overlapping issues. The focus is given to the challenges and pitfalls of isomiR expression analysis. Specifically, we address the availability of tools enabling research without extensive bioinformatics background to explore this fascinating corner of the small RNAome universe that may facilitate the discovery of new and more reliable disease biomarkers.