RESUMO
SARS-CoV-2 is a novel positive-sense single-stranded RNA virus from the Coronaviridae family (genus Betacoronavirus), which has been established as causing the COVID-19 pandemic. The genome of SARS-CoV-2 is one of the largest among known RNA viruses, comprising of at least 26 known protein-coding loci. Studies thus far have outlined the coding capacity of the positive-sense strand of the SARS-CoV-2 genome, which can be used directly for protein translation. However, it has been recently shown that transcribed negative-sense viral RNA intermediates that arise during viral genome replication from positive-sense viruses can also code for proteins. No studies have yet explored the potential for negative-sense SARS-CoV-2 RNA intermediates to contain protein-coding loci. Thus, using sequence and structure-based bioinformatics methodologies, we have investigated the presence and validity of putative negative-sense ORFs (nsORFs) in the SARS-CoV-2 genome. Nine nsORFs were discovered to contain strong eukaryotic translation initiation signals and high codon adaptability scores, and several of the nsORFs were predicted to interact with RNA-binding proteins. Evolutionary conservation analyses indicated that some of the nsORFs are deeply conserved among related coronaviruses. Three-dimensional protein modeling revealed the presence of higher order folding among all putative SARS-CoV-2 nsORFs, and subsequent structural mimicry analyses suggest similarity of the nsORFs to DNA/RNA-binding proteins and proteins involved in immune signaling pathways. Altogether, these results suggest the potential existence of still undescribed SARS-CoV-2 proteins, which may play an important role in the viral lifecycle and COVID-19 pathogenesis.
Assuntos
COVID-19 , SARS-CoV-2 , COVID-19/genética , Genoma Viral , Humanos , Pandemias , RNA Viral/química , RNA Viral/genética , Proteínas de Ligação a RNA/genética , SARS-CoV-2/genéticaRESUMO
Nucleic acids are not only static carriers of genetic information but also play vital roles in controlling cellular lifecycles through their fascinating structural diversity [...].
Assuntos
Biologia Computacional , DNA , Conformação de Ácido Nucleico , RNA , RNA/química , RNA/metabolismo , DNA/química , DNA/metabolismo , Biologia Computacional/métodos , HumanosRESUMO
In a recently published paper, we have found that SARS-CoV-2 hot-spot mutations are significantly associated with inverted repeat loci and CG dinucleotides. However, fast-spreading strains with new mutations (so-called mink farm mutations, England mutations and Japan mutations) have been recently described. We used the new datasets to check the positioning of mutation sites in genomes of the new SARS-CoV-2 strains. Using an open-access Palindrome analyzer tool, we found mutations in these new strains to be significantly enriched in inverted repeat loci.
Assuntos
Mutação , SARS-CoV-2/genética , COVID-19/virologia , Genoma Viral , HumanosRESUMO
SARS-CoV-2 is an intensively investigated virus from the order Nidovirales (Coronaviridae family) that causes COVID-19 disease in humans. Through enormous scientific effort, thousands of viral strains have been sequenced to date, thereby creating a strong background for deep bioinformatics studies of the SARS-CoV-2 genome. In this study, we inspected high-frequency mutations of SARS-CoV-2 and carried out systematic analyses of their overlay with inverted repeat (IR) loci and CpG islands. The main conclusion of our study is that SARS-CoV-2 hot-spot mutations are significantly enriched within both IRs and CpG island loci. This points to their role in genomic instability and may predict further mutational drive of the SARS-CoV-2 genome. Moreover, CpG islands are strongly enriched upstream from viral ORFs and thus could play important roles in transcription and the viral life cycle. We hypothesize that hypermethylation of these loci will decrease the transcription of viral ORFs and could therefore limit the progression of the disease.
Assuntos
COVID-19/virologia , Ilhas de CpG , Mutação , SARS-CoV-2/genética , Metilação de DNA , Genoma Viral , Humanos , Ligação ProteicaRESUMO
The dynamic processes operating on genomic DNA, such as gene expression and cellular division, lead inexorably to topological challenges in the form of entanglements, catenanes, knots, "bubbles", R-loops, and other outcomes of supercoiling and helical disruption. The resolution of toxic topological stress is the function attributed to DNA topoisomerases. A prominent example is the negative supercoiling (nsc) trailing processive enzymes such as DNA and RNA polymerases. The multiple equilibrium states that nscDNA can adopt by redistribution of helical twist and writhe include the left-handed double-helical conformation known as Z-DNA. Thirty years ago, one of our labs isolated a protein from Drosophila cells and embryos with a 100-fold greater affinity for Z-DNA than for B-DNA, and identified it as topoisomerase II (gene Top2, orthologous to the human UniProt proteins TOP2A and TOP2B). GTP increased the affinity and selectivity for Z-DNA even further and also led to inhibition of the isomerase enzymatic activity. An allosteric mechanism was proposed, in which topoII acts as a Z-DNA-binding protein (ZBP) to stabilize given states of topological (sub)domains and associated multiprotein complexes. We have now explored this possibility by comprehensive bioinformatic analyses of the available protein sequences of topoII representing organisms covering the whole tree of life. Multiple alignment of these sequences revealed an extremely high level of evolutionary conservation, including a winged-helix protein segment, here denoted as Zτ, constituting the putative structural homolog of Zα, the canonical Z-DNA/Z-RNA binding domain previously identified in the interferon-inducible RNA Adenosine-to-Inosine-editing deaminase, ADAR1p150. In contrast to Zα, which is separate from the protein segment responsible for catalysis, Zτ encompasses the active site tyrosine of topoII; a GTP-binding site and a GxxG sequence motif are in close proximity. Quantitative Zτ-Zα similarity comparisons and molecular docking with interaction scoring further supported the "B-Z-topoII hypothesis" and has led to an expanded mechanism for topoII function incorporating the recognition of Z-DNA segments ("Z-flipons") as an inherent and essential element. We further propose that the two Zτ domains of the topoII homodimer exhibit a single-turnover "conformase" activity on given G(ate) B-DNA segments ("Z-flipins"), inducing their transition to the left-handed Z-conformation. Inasmuch as the topoII-Z-DNA complexes are isomerase inactive, we infer that they fulfill important structural roles in key processes such as mitosis. Topoisomerases are preeminent targets of anti-cancer drug discovery, and we anticipate that detailed elucidation of their structural-functional interactions with Z-DNA and GTP will facilitate the design of novel, more potent and selective anti-cancer chemotherapeutic agents.
Assuntos
DNA de Forma B , DNA Forma Z , Humanos , Simulação de Acoplamento Molecular , DNA/química , DNA Topoisomerases Tipo II/genética , DNA Topoisomerases Tipo II/metabolismo , Guanosina Trifosfato , Adenosina Desaminase/metabolismoRESUMO
Plant miRNAs are powerful regulators of gene expression at the post-transcriptional level, which was repeatedly proved in several model plant species. miRNAs are considered to be key regulators of many developmental, homeostatic, and immune processes in plants. However, our understanding of plant miRNAs is still limited, despite the fact that an increasing number of studies have appeared. This systematic review aims to summarize our current knowledge about miRNAs in spring barley (Hordeum vulgare), which is an important agronomical crop worldwide and serves as a common monocot model for studying abiotic stress responses as well. This can help us to understand the connection between plant miRNAs and (not only) abiotic stresses in general. In the end, some future perspectives and open questions are summarized.
Assuntos
Hordeum , MicroRNAs , Hordeum/genética , Hordeum/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Estresse Fisiológico/genética , Plantas/metabolismo , Regulação da Expressão Gênica de PlantasRESUMO
Photosynthetically active radiation (PAR) is an important environmental cue inducing the production of many secondary metabolites involved in plant oxidative stress avoidance and tolerance. To examine the complex role of PAR irradiance and specific spectral components on the accumulation of phenolic compounds (PheCs), we acclimated spring barley (Hordeum vulgare) to different spectral qualities (white, blue, green, red) at three irradiances (100, 200, 400 µmol m-2 s-1). We confirmed that blue light irradiance is essential for the accumulation of PheCs in secondary barley leaves (in UV-lacking conditions), which underpins the importance of photoreceptor signals (especially cryptochrome). Increasing blue light irradiance most effectively induced the accumulation of B-dihydroxylated flavonoids, probably due to the significantly enhanced expression of the F3'H gene. These changes in PheC metabolism led to a steeper increase in antioxidant activity than epidermal UV-A shielding in leaf extracts containing PheCs. In addition, we examined the possible role of miRNAs in the complex regulation of gene expression related to PheC biosynthesis.
Assuntos
Hordeum , Raios Ultravioleta , Flavonoides/metabolismo , Hordeum/genética , Hordeum/metabolismo , Luz , Fenóis/metabolismo , Folhas de Planta/genética , Folhas de Planta/metabolismoRESUMO
Z-DNA and Z-RNA are functionally important left-handed structures of nucleic acids, which play a significant role in several molecular and biological processes including DNA replication, gene expression regulation and viral nucleic acid sensing. Most proteins that have been proven to interact with Z-DNA/Z-RNA contain the so-called Zα domain, which is structurally well conserved. To date, only eight proteins with Zα domain have been described within a few organisms (including human, mouse, Danio rerio, Trypanosoma brucei and some viruses). Therefore, this paper aimed to search for new Z-DNA/Z-RNA binding proteins in the complete PDB structures database and from the AlphaFold2 protein models. A structure-based similarity search found 14 proteins with highly similar Zα domain structure in experimentally-defined proteins and 185 proteins with a putative Zα domain using the AlphaFold2 models. Structure-based alignment and molecular docking confirmed high functional conservation of amino acids involved in Z-DNA/Z-RNA, suggesting that Z-DNA/Z-RNA recognition may play an important role in a variety of cellular processes.
Assuntos
DNA Forma Z/química , Proteínas de Ligação a DNA/química , Modelos Moleculares , Domínios e Motivos de Interação entre Proteínas , Proteínas de Ligação a RNA/química , RNA/química , Sequência de Aminoácidos , Sítios de Ligação , DNA Forma Z/metabolismo , Proteínas de Ligação a DNA/metabolismo , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Conformação de Ácido Nucleico , Ligação Proteica , Conformação Proteica , RNA/metabolismo , Proteínas de Ligação a RNA/metabolismo , Relação Estrutura-AtividadeRESUMO
Nucleic acid-binding proteins are traditionally divided into two categories: With the ability to bind DNA or RNA. In the light of new knowledge, such categorizing should be overcome because a large proportion of proteins can bind both DNA and RNA. Another even more important features of nucleic acid-binding proteins are so-called sequence or structure specificities. Proteins able to bind nucleic acids in a sequence-specific manner usually contain one or more of the well-defined structural motifs (zinc-fingers, leucine zipper, helix-turn-helix, or helix-loop-helix). In contrast, many proteins do not recognize nucleic acid sequence but rather local DNA or RNA structures (G-quadruplexes, i-motifs, triplexes, cruciforms, left-handed DNA/RNA form, and others). Finally, there are also proteins recognizing both sequence and local structural properties of nucleic acids (e.g., famous tumor suppressor p53). In this mini-review, we aim to summarize current knowledge about the amino acid composition of various types of nucleic acid-binding proteins with a special focus on significant enrichment and/or depletion in each category.
Assuntos
Proteínas de Ligação a DNA/genética , DNA/ultraestrutura , Conformação de Ácido Nucleico , RNA/ultraestrutura , Sequência de Aminoácidos/genética , Proteínas de Transporte/genética , Proteínas de Transporte/ultraestrutura , DNA/genética , DNA Forma Z , Quadruplex G , Humanos , Zíper de Leucina/genética , Nucleoproteínas/genética , Nucleoproteínas/ultraestrutura , RNA/química , Dedos de Zinco/genéticaRESUMO
The importance of gene expression regulation in viruses based upon G-quadruplex may point to its potential utilization in therapeutic targeting. Here, we present analyses as to the occurrence of putative G-quadruplex-forming sequences (PQS) in all reference viral dsDNA genomes and evaluate their dependence on PQS occurrence in host organisms using the G4Hunter tool. PQS frequencies differ across host taxa without regard to GC content. The overlay of PQS with annotated regions reveals the localization of PQS in specific regions. While abundance in some, such as repeat regions, is shared by all groups, others are unique. There is abundance within introns of Eukaryota-infecting viruses, but depletion of PQS in introns of bacteria-infecting viruses. We reveal a significant positive correlation between PQS frequencies in dsDNA viruses and corresponding hosts from archaea, bacteria, and eukaryotes. A strong relationship between PQS in a virus and its host indicates their close coevolution and evolutionarily reciprocal mimicking of genome organization.
Assuntos
Biologia Computacional/métodos , DNA/genética , Quadruplex G , Genoma Viral , Proteínas Virais/genética , Archaea/virologia , Bactérias/virologia , Regulação da Expressão Gênica , Genoma , Humanos , Vírus/genéticaRESUMO
G-quadruplexes have long been perceived as rare and physiologically unimportant nucleic acid structures. However, several studies have revealed their importance in molecular processes, suggesting their possible role in replication and gene expression regulation. Pathways involving G-quadruplexes are intensively studied, especially in the context of human diseases, while their involvement in gene expression regulation in plants remains largely unexplored. Here, we conducted a bioinformatic study and performed a complex circular dichroism measurement to identify a stable G-quadruplex in the gene RPB1, coding for the RNA polymerase II large subunit. We found that this G-quadruplex-forming locus is highly evolutionarily conserved amongst plants sensu lato (Archaeplastida) that share a common ancestor more than one billion years old. Finally, we discussed a new hypothesis regarding G-quadruplexes interacting with UV light in plants to potentially form an additional layer of the regulatory network.
Assuntos
Quadruplex G , Proteínas de Plantas/química , Plantas/química , RNA Polimerase II/química , Sequência de Aminoácidos , Arabidopsis/química , Arabidopsis/genética , Arabidopsis/efeitos da radiação , Dicroísmo Circular , Biologia Computacional , Evolução Molecular , Quadruplex G/efeitos da radiação , Regulação da Expressão Gênica de Plantas/genética , Glaucófitas/química , Glaucófitas/genética , Glaucófitas/efeitos da radiação , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/efeitos da radiação , Plantas/genética , Plantas/efeitos da radiação , RNA Polimerase II/genética , Rodófitas/química , Rodófitas/genética , Rodófitas/efeitos da radiação , Alinhamento de Sequência , Raios UltravioletaRESUMO
Recently, the quest for the mythical fountain of youth has produced extensive research programs that aim to extend the healthy lifespan of humans. Despite advances in our understanding of the aging process, the surprisingly extended lifespan and cancer resistance of some animal species remain unexplained. The p53 protein plays a crucial role in tumor suppression, tissue homeostasis, and aging. Long-lived, cancer-free African elephants have 20 copies of the TP53 gene, including 19 retrogenes (38 alleles), which are partially active, whereas humans possess only one copy of TP53 and have an estimated cancer mortality rate of 11-25%. The mechanism through which p53 contributes to the resolution of the Peto's paradox in Animalia remains vague. Thus, in this work, we took advantage of the available datasets and inspected the p53 amino acid sequence of phylogenetically related organisms that show variations in their lifespans. We discovered new correlations between specific amino acid deviations in p53 and the lifespans across different animal species. We found that species with extended lifespans have certain characteristic amino acid substitutions in the p53 DNA-binding domain that alter its function, as depicted from the Phenotypic Annotation of p53 Mutations, using the PROVEAN tool or SWISS-MODEL workflow. In addition, the loop 2 region of the human p53 DNA-binding domain was identified as the longest region that was associated with longevity. The 3D model revealed variations in the loop 2 structure in long-lived species when compared with human p53. Our findings show a direct association between specific amino acid residues in p53 protein, changes in p53 functionality, and the extended animal lifespan, and further highlight the importance of p53 protein in aging.
Assuntos
Bases de Dados Genéticas , Dosagem de Genes , Longevidade , Modelos Moleculares , Animais , Domínios Proteicos , Estrutura Secundária de Proteína , Especificidade da Espécie , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
The p53 family of transcription factors plays key roles in development, genome stability, senescence and tumor development, and p53 is the most important tumor suppressor protein in humans. Although intensively investigated for many years, its initial evolutionary history is not yet fully elucidated. Using bioinformatic and structure prediction methods on current databases containing newly-sequenced genomes and transcriptomes, we present a detailed characterization of p53 family homologs in remote members of the Holozoa group, in the unicellular clades Filasterea, Ichthyosporea and Corallochytrea. Moreover, we show that these newly characterized homologous sequences contain domains that can form structures with high similarity to the human p53 family DNA-binding domain, and some also show similarities to the oligomerization and SAM domains. The presence of these remote homologs demonstrates an ancient origin of the p53 protein family.
Assuntos
Eucariotos/genética , Evolução Molecular , Família Multigênica , Homologia de Sequência de Aminoácidos , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/genética , Sequência de Aminoácidos , Bases de Dados Genéticas , Eucariotos/classificação , Éxons , Íntrons , Modelos Moleculares , Filogenia , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Proteína Supressora de Tumor p53/metabolismoRESUMO
The role of local DNA structures in the regulation of basic cellular processes is an emerging field of research. Amongst local non-B DNA structures, the significance of G-quadruplexes was demonstrated in the last decade, and their presence and functional relevance has been demonstrated in many genomes, including humans. In this study, we analyzed the presence and locations of G-quadruplex-forming sequences by G4Hunter in all complete bacterial genomes available in the NCBI database. G-quadruplex-forming sequences were identified in all species, however the frequency differed significantly across evolutionary groups. The highest frequency of G-quadruplex forming sequences was detected in the subgroup Deinococcus-Thermus, and the lowest frequency in Thermotogae. G-quadruplex forming sequences are non-randomly distributed and are favored in various evolutionary groups. G-quadruplex-forming sequences are enriched in ncRNA segments followed by mRNAs. Analyses of surrounding sequences showed G-quadruplex-forming sequences around tRNA and regulatory sequences. These data point to the unique and non-random localization of G-quadruplex-forming sequences in bacterial genomes.
Assuntos
Bactérias/genética , DNA Bacteriano/química , Quadruplex G , Genoma Bacteriano , Humanos , Conformação de Ácido Nucleico , FilogeniaRESUMO
Expansions of trinucleotide repeats (TNRs) are associated with genetic disorders such as Friedreich's ataxia. The tumor suppressor p53 is a central regulator of cell fate in response to different types of insults. Sequence and structure-selective modes of DNA recognition are among the main attributes of p53 protein. The focus of this work was analysis of the p53 structure-selective recognition of TNRs associated with human neurodegenerative diseases. Here, we studied binding of full length p53 and several deletion variants to TNRs folded into DNA hairpins or loops. We demonstrate that p53 binds to all studied non-B DNA structures, with a preference for non-B DNA structures formed by pyrimidine (Py) rich strands. Using deletion mutants, we determined the C-terminal DNA binding domain of p53 to be crucial for recognition of such non-B DNA structures. We also observed that p53 in vitro prefers binding to the Py-rich strand over the purine (Pu) rich strand in non-B DNA substrates formed by sequence derived from the first intron of the frataxin gene. The binding of p53 to this region was confirmed using chromatin immunoprecipitation in human Friedreich's ataxia fibroblast and adenocarcinoma cells. Altogether these observations provide further evidence that p53 binds to TNRs' non-B DNA structures.
Assuntos
DNA/química , DNA/metabolismo , Ataxia de Friedreich/genética , Ataxia de Friedreich/metabolismo , Conformação de Ácido Nucleico , Expansão das Repetições de Trinucleotídeos , Repetições de Trinucleotídeos , Proteína Supressora de Tumor p53/metabolismo , Expressão Gênica , Humanos , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Pirimidinas , Proteínas Recombinantes , Proteína Supressora de Tumor p53/químicaRESUMO
The importance of local DNA structures in the regulation of basic cellular processes is an emerging field of research. Amongst local non-B DNA structures, G-quadruplexes are perhaps the most well-characterized to date, and their presence has been demonstrated in many genomes, including that of humans. G-quadruplexes are selectively bound by many regulatory proteins. In this paper, we have analyzed the amino acid composition of all seventy-seven described G-quadruplex binding proteins of Homo sapiens. Our comparison with amino acid frequencies in all human proteins and specific protein subsets (e.g., all nucleic acid binding) revealed unique features of quadruplex binding proteins, with prominent enrichment for glycine (G) and arginine (R). Cluster analysis with bootstrap resampling shows similarities and differences in amino acid composition of particular quadruplex binding proteins. Interestingly, we found that all characterized G-quadruplex binding proteins share a 20 amino acid long motif/domain (RGRGR GRGGG SGGSG GRGRG) which is similar to the previously described RG-rich domain (RRGDG RRRGG GGRGQ GGRGR GGGFKG) of the FRM1 G-quadruplex binding protein. Based on this protein fingerprint, we have predicted a new set of potential G-quadruplex binding proteins sharing this interesting domain rich in glycine and arginine residues.
Assuntos
Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , DNA/química , Motivos de Aminoácidos , DNA/metabolismo , Quadruplex G , Humanos , Conformação de Ácido Nucleico , Mapas de Interação de ProteínasRESUMO
Site-specific DNA recognition and binding activity belong to common attributes of all three members of tumor suppressor p53 family proteins: p53, p63 and p73. It was previously shown that heavy metals can affect p53 conformation, sequence-specific binding and suppress p53 response to DNA damage. Here we report for the first time that cadmium, nickel and cobalt, which have already been shown to disturb various DNA repair mechanisms, can also influence p63 and p73 sequence-specific DNA binding activity and transactivation of p53 family target genes. Based on results of electrophoretic mobility shift assay and luciferase reporter assay, we conclude that cadmium inhibits sequence-specific binding of all three core domains to p53 consensus sequences and abolishes transactivation of several promoters (e.g. BAX and MDM2) by 50µM concentrations. In the presence of specific DNA, all p53 family core domains were partially protected against loss of DNA binding activity due to cadmium treatment. Effective cadmium concentration to abolish DNA-protein interactions was about two times higher for p63 and p73 proteins than for p53. Furthermore, we detected partial reversibility of cadmium inhibition for all p53 family members by EDTA. DTT was able to reverse cadmium inhibition only for p53 and p73. Nickel and cobalt abolished DNA-p53 interaction at sub-millimolar concentrations while inhibition of p63 and p73 DNA binding was observed at millimolar concentrations. In summary, cadmium strongly inhibits p53, p63 and p73 DNA binding in vitro and in cells in comparison to nickel and cobalt. The role of cadmium inhibition of p53 tumor suppressor family in carcinogenesis is discussed.
Assuntos
Cádmio/química , Cobalto/química , Proteínas de Ligação a DNA/química , DNA/química , Proteínas de Membrana/química , Níquel/química , Proteínas Nucleares/química , Proteínas Supressoras de Tumor/química , Linhagem Celular Tumoral , Ditiotreitol/química , Ácido Edético/química , Humanos , Metais/química , Metais Pesados/química , Ligação Proteica , Estrutura Terciária de Proteína , Ativação Transcricional , Proteína Tumoral p73 , Proteína Supressora de Tumor p53/químicaRESUMO
Sequences of nucleic acids with the potential to form four-stranded G-quadruplex structures are intensively studied mainly in the context of human diseases, pathogens, or extremophile organisms; nonetheless, the knowledge about their occurrence and putative role in plants is still limited. This work is focused on G-quadruplex-forming sites in two gene sets of interest: drought stress-responsive genes, and genes related to the production/biosynthesis of phenolic compounds in the model plant organism Arabidopsis thaliana. In addition, 20 housekeeping genes were analyzed as well, where the constitutive gene expression was expected (with no need for precise regulation depending on internal or external factors). The results have shown that none of the tested gene sets differed significantly in the content of G-quadruplex-forming sites, however, the highest frequency of G-quadruplex-forming sites was found in the 5'-UTR regions of phenolic compounds' biosynthesis genes, which indicates the possibility of their regulation at the mRNA level. In addition, mainly within the introns and 1000 bp flanks downstream gene regions, G-quadruplex-forming sites were highly underrepresented. Finally, cluster analysis allowed us to observe similarities between particular genes in terms of their PQS characteristics. We believe that the original approach used in this study may become useful for further and more comprehensive bioinformatic studies in the field of G-quadruplex genomics.
RESUMO
Epigenetics deals with changes in gene expression that are not caused by modifications in the primary sequence of nucleic acids. These changes beyond primary structures of nucleic acids not only include DNA/RNA methylation, but also other reversible conversions, together with histone modifications or RNA interference. In addition, under particular conditions (such as specific ion concentrations or protein-induced stabilization), the right-handed double-stranded DNA helix (B-DNA) can form noncanonical structures commonly described as "non-B DNA" structures. These structures comprise, for example, cruciforms, i-motifs, triplexes, and G-quadruplexes. Their formation often leads to significant differences in replication and transcription rates. Noncanonical RNA structures have also been documented to play important roles in translation regulation and the biology of noncoding RNAs. In human and animal studies, the frequency and dynamics of noncanonical DNA and RNA structures are intensively investigated, especially in the field of cancer research and neurodegenerative diseases. In contrast, noncanonical DNA and RNA structures in plants have been on the fringes of interest for a long time and only a few studies deal with their formation, regulation, and physiological importance for plant stress responses. Herein, we present a review focused on the main fields of epigenetics in plants and their possible roles in stress responses and signaling, with special attention dedicated to noncanonical DNA and RNA structures.
Assuntos
Quadruplex G , Ácidos Nucleicos , Animais , Humanos , DNA/genética , DNA/química , Epigênese Genética , RNA/genética , RNA/química , Plantas/genéticaRESUMO
The opium poppy's ability to produce various alkaloids is both useful and problematic. Breeding of new varieties with varying alkaloid content is therefore an important task. In this paper, the breeding technology of new low morphine poppy genotypes, based on a combination of a TILLING approach and single-molecule real-time NGS sequencing, is presented. Verification of the mutants in the TILLING population was obtained using RT-PCR and HPLC methods. Only three of the single-copy genes of the morphine pathway among the eleven genes were used for the identification of mutant genotypes. Point mutations were obtained only in one gene (CNMT) while an insertion was obtained in the other (SalAT). Only a few expected transition SNPs from G:C to A:T were obtained. In the low morphine mutant genotype, the production of morphine was decreased to 0.1% from 1.4% in the original variety. A comprehensive description of the breeding process, a basic characterization of the main alkaloid content, and a gene expression profile for the main alkaloid-producing genes is provided. Difficulties with the TILLING approach are also described and discussed.