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1.
BMC Genomics ; 22(1): 638, 2021 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-34479506

RESUMO

BACKGROUND: LncRNAs are extensively involved in plant biological processes. However, the lack of a comprehensive lncRNA landscape in moso bamboo has hindered the molecular study of lncRNAs. Moreover, the role of lncRNAs in secondary cell wall (SCW) biosynthesis of moso bamboo is elusive. RESULTS: For comprehensively identifying lncRNA throughout moso bamboo genome, we collected 231 RNA-Seq datasets, 1 Iso-Seq dataset, and 1 full-length cDNA dataset. We used a machine learning approach to improve the pipeline of lncRNA identification and functional annotation based on previous studies and identified 37,009 lncRNAs in moso bamboo. Then, we established a network of potential lncRNA-coding gene for SCW biosynthesis and identified SCW-related lncRNAs. We also proposed that a mechanism exists in bamboo to direct phenylpropanoid intermediates to lignin or flavonoids biosynthesis through the PAL/4CL/C4H genes. In addition, we identified 4 flavonoids and 1 lignin-preferred genes in the PAL/4CL/C4H gene families, which gained implications in molecular breeding. CONCLUSIONS: We provided a comprehensive landscape of lncRNAs in moso bamboo. Through analyses, we identified SCW-related lncRNAs and improved our understanding of lignin and flavonoids biosynthesis.


Assuntos
Parede Celular , Redes Reguladoras de Genes , Poaceae , RNA Longo não Codificante , Parede Celular/genética , Regulação da Expressão Gênica de Plantas , Poaceae/genética , RNA Longo não Codificante/genética , RNA de Plantas/genética
2.
BMC Plant Biol ; 21(1): 419, 2021 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-34517842

RESUMO

BACKGROUND: A key issue for implementation of CRISPR-Cas9 genome editing for plant trait improvement and gene function analysis is to efficiently deliver the components, including guide RNAs (gRNAs) and Cas9, into plants. Plant virus-based gRNA delivery strategy has proven to be an important tool for genome editing. However, its application in soybean which is an important crop has not been reported yet. ALSV (apple latent spherical virus) is highly infectious virus and could be explored for delivering elements for genome editing. RESULTS: To develop a ALSV-based gRNA delivery system, the Cas9-based Csy4-processed ALSV Carry (CCAC) system was developed. In this system, we engineered the soybean-infecting ALSV to carry and deliver gRNA(s). The endoribonuclease Csy4 effectively releases gRNAs that function efficiently in Cas9-mediated genome editing. Genome editing of endogenous phytoene desaturase (PDS) loci and exogenous 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) sequence in Nicotiana. benthamiana (N. benthamiana) through CCAC was confirmed using Sanger sequencing. Furthermore, CCAC-induced mutagenesis in two soybean endogenous GW2 paralogs was detected. CONCLUSIONS: With the aid of the CCAC system, the target-specific gRNA(s) can be easily manipulated and efficiently delivered into soybean plant cells by viral infection. This is the first virus-based gRNA delivery system for soybean for genome editing and can be used for gene function study and trait improvement.


Assuntos
Sistemas CRISPR-Cas/genética , Edição de Genes/métodos , Interações Hospedeiro-Patógeno/genética , Vírus de Plantas/genética , Soja/genética , Soja/virologia , Viroses/genética , Produtos Agrícolas/genética , Produtos Agrícolas/virologia , Regulação da Expressão Gênica de Plantas , Regulação Viral da Expressão Gênica , Genoma de Planta , Mutagênese , RNA Guia , RNA de Plantas , RNA Viral
3.
BMC Plant Biol ; 21(1): 423, 2021 Sep 17.
Artigo em Inglês | MEDLINE | ID: mdl-34535087

RESUMO

BACKGROUND: The GRAS gene family plays crucial roles in multiple biological processes of plant growth, including seed development, which is related to seedless traits of litchi (Litchi chinensis Sonn.). However, it hasn't been fully identified and analyzed in litchi, an economic fruit tree cultivated in subtropical regions. RESULTS: In this study, 48 LcGRAS proteins were identified and termed according to their chromosomal location. LcGRAS proteins can be categorized into 14 subfamilies through phylogenetic analysis. Gene structure and conserved domain analysis revealed that different subfamilies harbored various motif patterns, suggesting their functional diversity. Synteny analysis revealed that the expansion of the GRAS family in litchi may be driven by their tandem and segmental duplication. After comprehensively analysing degradome data, we found that four LcGRAS genes belong to HAM subfamily were regulated via miR171-mediated degradation. The various expression patterns of LcGRAS genes in different tissues uncovered they were involved in different biological processes. Moreover, the different temporal expression profiles of LcGRAS genes between abortive and bold seed indicated some of them were involved in maintaining the normal development of the seed. CONCLUSION: Our study provides comprehensive analyses on GRAS family members in litchi, insight into a better understanding of the roles of GRAS in litchi development, and lays the foundation for further investigations on litchi seed development.


Assuntos
Litchi/genética , Proteínas de Plantas/genética , Sementes/crescimento & desenvolvimento , Mapeamento Cromossômico , Regulação da Expressão Gênica de Plantas , Litchi/crescimento & desenvolvimento , MicroRNAs , Família Multigênica , Filogenia , RNA de Plantas , Sementes/genética , Sintenia , Fatores de Transcrição/genética
4.
BMC Plant Biol ; 21(1): 409, 2021 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-34493224

RESUMO

BACKGROUND: The periderm is a protective barrier crucial for land plant survival, but little is known about genetic factors involved in its development and regulation. Using a transcriptomic approach in the cork oak (Q. suber) periderm, we previously identified an RS2-INTERACTING KH PROTEIN (RIK) homologue of unknown function containing a K homology (KH)-domain RNA-binding protein, as a regulatory candidate gene in the periderm. RESULTS: To gain insight into the function of RIK in the periderm, potato (S. tuberosum) tuber periderm was used as a model: the full-length coding sequence of RIK, hereafter referred to as StRIK, was isolated, the transcript profile analyzed and gene silencing in potato performed to analyze the silencing effects on periderm anatomy and transcriptome. The StRIK transcript accumulated in all vegetative tissues studied, including periderm and other suberized tissues such as root and also in wounded tissues. Downregulation of StRIK in potato by RNA interference (StRIK-RNAi) did not show any obvious effects on tuber periderm anatomy but, unlike Wild type, transgenic plants flowered. Global transcript profiling of the StRIK-RNAi periderm did show altered expression of genes associated with RNA metabolism, stress and signaling, mirroring the biological processes found enriched within the in silico co-expression network of the Arabidopsis orthologue. CONCLUSIONS: The ubiquitous expression of StRIK transcript, the flower associated phenotype and the differential expression of StRIK-RNAi periderm point out to a general regulatory role of StRIK in diverse plant developmental processes. The transcriptome analysis suggests that StRIK might play roles in RNA maturation and stress response in the periderm.


Assuntos
Proteínas de Plantas/genética , Tubérculos/genética , RNA de Plantas/metabolismo , Solanum tuberosum/genética , Estresse Fisiológico/genética , Arabidopsis/genética , Núcleo Celular/genética , Núcleo Celular/metabolismo , Elementos de DNA Transponíveis , Flores/genética , Flores/fisiologia , Regulação da Expressão Gênica de Plantas , Redes Reguladoras de Genes , Inativação Gênica , Proteínas de Plantas/metabolismo , Tubérculos/anatomia & histologia , Tubérculos/citologia , Plantas Geneticamente Modificadas , Solanum tuberosum/citologia
5.
BMC Plant Biol ; 21(1): 410, 2021 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-34493227

RESUMO

BACKGROUND: Water deficit is an abiotic stress that retards plant growth and destabilizes crop production. Long non coding RNAs (lncRNAs) are a class of non-coding endogenous RNAs that participate in diverse cellular processes and stress responses in plants. lncRNAs could function as competing endogenous RNAs (ceRNA) and represent a novel layer of gene regulation. However, the regulatory mechanism of lncRNAs as ceRNA in drought stress response is yet unclear. RESULTS: In this study, we performed transcriptome-wide identification of drought-responsive lncRNAs in rice. Thereafter, we constructed a lncRNA-mediated ceRNA network by analyzing competing relationships between mRNAs and lncRNAs based on ceRNA hypothesis. A drought responsive ceRNA network with 40 lncRNAs, 23 miRNAs and 103 mRNAs was obtained. Network analysis revealed TCONS_00021861/miR528-3p/YUCCA7 regulatory axis as a hub involved in drought response. The miRNA-target expression and interaction were validated by RT-qPCR and RLM-5'RACE. TCONS_00021861 showed significant positive correlation (r = 0.7102) with YUCCA7 and negative correlation with miR528-3p (r = -0.7483). Overexpression of TCONS_00021861 attenuated the repression of miR528-3p on YUCCA7, leading to increased IAA (Indole-3-acetic acid) content and auxin overproduction phenotypes. CONCLUSIONS: TCONS_00021861 could regulate YUCCA7 by sponging miR528-3p, which in turn activates IAA biosynthetic pathway and confer resistance to drought stress. Our findings provide a new perspective of the regulatory roles of lncRNAs as ceRNAs in drought resistance of rice.


Assuntos
Oryza/genética , RNA Longo não Codificante/genética , Desidratação/genética , Secas , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Redes Reguladoras de Genes , Células do Mesofilo/ultraestrutura , MicroRNAs/genética , Folhas de Planta/genética , RNA Mensageiro/genética , RNA de Plantas , Espécies Reativas de Oxigênio/metabolismo
6.
Int J Mol Sci ; 22(16)2021 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-34445207

RESUMO

Recent studies show a crucial role of post-transcriptional processes in the regulation of gene expression. Our research has shown that mRNA retention in the nucleus plays a significant role in such regulation. We studied larch microsporocytes during meiotic prophase, characterized by pulsatile transcriptional activity. After each pulse, the transcriptional activity is silenced, but the transcripts synthesized at this time are not exported immediately to the cytoplasm but are retained in the cell nucleus and especially in Cajal bodies, where non-fully-spliced transcripts with retained introns are accumulated. Analysis of the transcriptome of these cells and detailed analysis of the nuclear retention and transport dynamics of several mRNAs revealed two main patterns of nuclear accumulation and transport. The majority of studied transcripts followed the first one, consisting of a more extended retention period and slow release to the cytoplasm. We have shown this in detail for the pre-mRNA and mRNA encoding RNA pol II subunit 10. In this pre-mRNA, a second (retained) intron is posttranscriptionally spliced at a precisely defined time. Fully mature mRNA is then released into the cytoplasm, where the RNA pol II complexes are produced. These proteins are necessary for transcription in the next pulse to occur.mRNAs encoding translation factors and SERRATE followed the second pattern, in which the retention period was shorter and transcripts were rapidly transferred to the cytoplasm. The presence of such a mechanism in various cell types from a diverse range of organisms suggests that it is an evolutionarily conserved mechanism of gene regulation.


Assuntos
Núcleo Celular/metabolismo , Larix/metabolismo , Pólen/metabolismo , Prófase , RNA Mensageiro/metabolismo , RNA de Plantas/metabolismo , Núcleo Celular/genética , Larix/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Pólen/genética , RNA Polimerase II/genética , RNA Polimerase II/metabolismo , RNA Mensageiro/genética , RNA de Plantas/genética
7.
BMC Plant Biol ; 21(1): 375, 2021 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-34388962

RESUMO

BACKGROUND: The lace plant (Aponogeton madagascariensis) is an aquatic monocot that develops leaves with uniquely formed perforations through the use of a developmentally regulated process called programmed cell death (PCD). The process of perforation formation in lace plant leaves is subdivided into several developmental stages: pre-perforation, window, perforation formation, perforation expansion and mature. The first three emerging "imperforate leaves" do not form perforations, while all subsequent leaves form perforations via developmentally regulated PCD. PCD is active in cells called "PCD cells" that do not retain the antioxidant anthocyanin in spaces called areoles framed by the leaf veins of window stage leaves. Cells near the veins called "NPCD cells" retain a red pigmentation from anthocyanin and do not undergo PCD. While the cellular changes that occur during PCD are well studied, the gene expression patterns underlying these changes and driving PCD during leaf morphogenesis are mostly unknown. We sought to characterize differentially expressed genes (DEGs) that mediate lace plant leaf remodelling and PCD. This was achieved performing gene expression analysis using transcriptomics and comparing DEGs among different stages of leaf development, and between NPCD and PCD cells isolated by laser capture microdissection. RESULTS: Transcriptomes were sequenced from imperforate, pre-perforation, window, and mature leaf stages, as well as PCD and NPCD cells isolated from window stage leaves. Differential expression analysis of the data revealed distinct gene expression profiles: pre-perforation and window stage leaves were characterized by higher expression of genes involved in anthocyanin biosynthesis, plant proteases, expansins, and autophagy-related genes. Mature and imperforate leaves upregulated genes associated with chlorophyll development, photosynthesis, and negative regulators of PCD. PCD cells were found to have a higher expression of genes involved with ethylene biosynthesis, brassinosteroid biosynthesis, and hydrolase activity whereas NPCD cells possessed higher expression of auxin transport, auxin signalling, aspartyl proteases, cysteine protease, Bag5, and anthocyanin biosynthesis enzymes. CONCLUSIONS: RNA sequencing was used to generate a de novo transcriptome for A. madagascariensis leaves and revealed numerous DEGs potentially involved in PCD and leaf remodelling. The data generated from this investigation will be useful for future experiments on lace plant leaf development and PCD in planta.


Assuntos
Alismatales/genética , Alismatales/fisiologia , Apoptose , Folhas de Planta/fisiologia , Alismatales/crescimento & desenvolvimento , Antocianinas/biossíntese , Apoptose/genética , Parede Celular/enzimologia , Regulação da Expressão Gênica de Plantas , Células Vegetais , Reguladores de Crescimento de Plantas/fisiologia , Folhas de Planta/genética , Proteínas de Plantas/metabolismo , RNA de Plantas , RNA-Seq , Fatores de Transcrição/fisiologia , Transcriptoma
8.
Nat Commun ; 12(1): 4941, 2021 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-34400639

RESUMO

Plant small RNAs are important regulatory elements that fine-tune gene expression and maintain genome integrity by silencing transposons. Reproductive organs of monocots produce abundant phased, small interfering RNAs (phasiRNAs). The 21-nt reproductive phasiRNAs triggered by miR2118 are highly enriched in pre-meiotic anthers, and have been found in multiple eudicot species, in contrast with prior reports of monocot specificity. The 24-nt reproductive phasiRNAs are triggered by miR2275, and are highly enriched during meiosis in many angiosperms. Here, we report the widespread presence of the 21-nt reproductive phasiRNA pathway in eudicots including canonical and non-canonical microRNA (miRNA) triggers of this pathway. In eudicots, these 21-nt phasiRNAs are enriched in pre-meiotic stages, a spatiotemporal distribution consistent with that of monocots and suggesting a role in anther development. Although this pathway is apparently absent in well-studied eudicot families including the Brassicaceae, Solanaceae and Fabaceae, our work in eudicots supports an earlier singular finding in spruce, a gymnosperm, indicating that the pathway of 21-nt reproductive phasiRNAs emerged in seed plants and was lost in some lineages.


Assuntos
Magnoliopsida/metabolismo , Nucleotídeos/metabolismo , RNA de Plantas/genética , RNA Interferente Pequeno/metabolismo , Sementes/metabolismo , Fragaria/genética , Fragaria/metabolismo , Regulação da Expressão Gênica de Plantas , Meiose , MicroRNAs/genética , Filogenia , Picea/genética , Proteínas de Plantas/genética , RNA de Cadeia Dupla/metabolismo , Solanaceae/metabolismo , Transcriptoma
9.
BMC Plant Biol ; 21(1): 366, 2021 Aug 11.
Artigo em Inglês | MEDLINE | ID: mdl-34380425

RESUMO

BACKGROUND: Small RNAs are short non-coding RNAs that are key gene regulators controlling various biological processes in eukaryotes. Plants may regulate discrete sets of sRNAs in response to pathogen attack. Sclerotinia sclerotiorum is an economically important pathogen affecting hundreds of plant species, including the economically important oilseed B. napus. However, there are limited studies on how regulation of sRNAs occurs in the S. sclerotiorum and B. napus pathosystem. RESULTS: We identified different classes of sRNAs from B. napus using high throughput sequencing of replicated mock and infected samples at 24 h post-inoculation (HPI). Overall, 3999 sRNA loci were highly expressed, of which 730 were significantly upregulated during infection. These 730 up-regulated sRNAs targeted 64 genes, including disease resistance proteins and transcriptional regulators. A total of 73 conserved miRNA families were identified in our dataset. Degradome sequencing identified 2124 cleaved mRNA products from these miRNAs from combined mock and infected samples. Among these, 50 genes were specific to infection. Altogether, 20 conserved miRNAs were differentially expressed and 8 transcripts were cleaved by the differentially expressed miRNAs miR159, miR5139, and miR390, suggesting they may have a role in the S. sclerotiorum response. A miR1885-triggered disease resistance gene-derived secondary sRNA locus was also identified and verified with degradome sequencing. We also found further evidence for silencing of a plant immunity related ethylene response factor gene by a novel sRNA using 5'-RACE and RT-qPCR. CONCLUSIONS: The findings in this study expand the framework for understanding the molecular mechanisms of the S. sclerotiorum and B. napus pathosystem at the sRNA level.


Assuntos
Ascomicetos/fisiologia , Brassica napus/genética , Brassica napus/microbiologia , Doenças das Plantas/microbiologia , RNA de Plantas , Pequeno RNA não Traduzido , Sequência Conservada , Resistência à Doença/genética , Regulação da Expressão Gênica de Plantas , Doenças das Plantas/genética , Análise de Sequência de RNA , Regulação para Cima
10.
Curr Opin Biotechnol ; 70: 204-212, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34217122

RESUMO

Small RNA (sRNA)-mediated RNA interference (RNAi) is a regulatory mechanism conserved in almost all eukaryotes. sRNAs play a critical role in host pathogen interactions either endogenously or by traveling between the interacting organisms and inducing 'cross-Kingdom RNAi' in the counterparty. Cross-kingdom RNAi is the mechanistic basis of host-induced gene silencing (HIGS), which relies on genetically expressing pathogen-gene targeting RNAs in crops, and has been successfully utilized against both microbial pathogens and pests. HIGS is limited by the need to produce genetically engineered crops. Recent studies have demonstrated that double-stranded RNAs and sRNAs can be efficiently taken up by many fungal pathogens, and induce gene silencing in fungal cells. This mechanism, termed 'environmental RNAi', allows direct application of pathogen-gene targeting RNAs onto crops to silence fungal virulence-related genes for plant protection. In this review, we will focus on how we can leverage cross-kingdom RNAi and environmental RNAi for crop disease control.


Assuntos
Proteção de Cultivos , Produtos Agrícolas , Produtos Agrícolas/genética , Interferência de RNA , RNA de Plantas , RNA Interferente Pequeno
11.
Comput Biol Med ; 136: 104662, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34311261

RESUMO

The coronavirus disease of 2019 (COVID-19) began as an outbreak and has taken a toll on human lives. The current pandemic requires scientific attention; hence we designed a systematic computational workflow to identify the cellular microRNAs (miRNAs) from human host possessing the capability to target and silence 3'UTR of SARS-CoV-2 genome. Based on this viewpoint, we extended our miRNA search to medicinal plants like Ocimum tenuiflorum, Zingiber officinale and Piper nigrum, which are well-known to possess antiviral properties, and are often consumed raw or as herbal decoctions. Such an approach, that makes use of miRNA of one species to interact and silence genes of another species including viruses is broadly categorized as cross-kingdom interactions. As a part of our genomics study on host-virus-plant interaction, we identified one unique 3'UTR conserved site 'GGAAGAG' amongst 5024 globally submitted SARS-CoV-2 complete genomes, which can be targeted by the human miRNA 'hsa-miR-1236-3p' and by Z. officinale miRNA 'zof-miR2673b'. Additionally, we also predicted that the members of miR477 family commonly found in these three plant genomes possess an inherent potential to silence viral genome RNA and facilitate antiviral defense against SARS-CoV-2 infection. In conclusion, this study reveals a universal site in the SARS-CoV-2 genome that may be crucial for targeted therapeutics to cure COVID-19.


Assuntos
COVID-19 , MicroRNAs , Plantas Medicinais , Regiões 3' não Traduzidas/genética , Biologia Computacional , Genômica , Humanos , MicroRNAs/genética , Plantas Medicinais/genética , RNA de Plantas , SARS-CoV-2
12.
Plant Mol Biol ; 106(6): 521-531, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34224063

RESUMO

KEY MESSAGE: We characterize a functional lincRNA, XH123 in cotton seedling in defense of cold stress. The silencing of XH123 leads to increased sensitivity to cold stress and the decay of chloroplast. Cotton, which originated from the arid mid-American region, is one of the most important cash crops worldwide. Cultivated cotton is now widely spread throughout high-altitude regions such as those in the far northwest of Asia. In such areas, spring temperatures below 12 ℃ impose cold stress on cotton seedlings, with concomitant threat of lost yield and productivity. It is documented that cold stress can induce differential expression of long noncoding RNAs (lncRNAs) in cotton; however, it is not yet clear if these cold-responsive lncRNAs are actively involved with tolerance of cold stress at the molecular level. Here, we select ten long intergenic non-coding RNAs as candidate genes and use virus-induced gene silencing and additional cold treatments to examine their roles in the response to cold stress during the cotton seedling stage. One such gene, XH123, was revealed to be involved in tolerance of cold stress. Specifically, XH123-silenced plants demonstrated sensitivity to cold stress, exhibiting chloroplast damage and increased endogenous levels of reactive oxygen species. The transcriptome profile of XH123-silenced seedlings was similar to that of cold-stressed seedlings having the known cold stress gene PIF3 silenced. These results imply that the lincRNA XH123 is actively involved with cold stress regulation in cotton during the seedling stage.


Assuntos
Resposta ao Choque Frio/genética , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas , Gossypium/genética , RNA Longo não Codificante/genética , RNA de Plantas/genética , Adaptação Fisiológica/genética , Cloroplastos/genética , Cloroplastos/metabolismo , Cloroplastos/ultraestrutura , Temperatura Baixa , Inativação Gênica , Gossypium/crescimento & desenvolvimento , Microscopia Eletrônica de Transmissão , Folhas de Planta/genética , Folhas de Planta/metabolismo , Folhas de Planta/ultraestrutura , RNA-Seq/métodos , Plântula/genética , Plântula/crescimento & desenvolvimento
13.
Int J Mol Sci ; 22(13)2021 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-34209014

RESUMO

Elevated concentrations of heavy metals such as cadmium (Cd) have a negative impact on staple crop production due to their ability to elicit cytotoxic and genotoxic effects on plants. In order to understand the relationship between Cd stress and plants in an effort to improve Cd tolerance, studies have identified genetic mechanisms which could be important for conferring stress tolerance. In recent years epigenetic studies have garnered much attention and hold great potential in both improving the understanding of Cd stress in plants as well as revealing candidate mechanisms for future work. This review describes some of the main epigenetic mechanisms involved in Cd stress responses. We summarize recent literature and data pertaining to chromatin remodeling, DNA methylation, histone acetylation and miRNAs in order to understand the role these epigenetic traits play in cadmium tolerance. The review aims to provide the framework for future studies where these epigenetic traits may be used in plant breeding and molecular studies in order to improve Cd tolerance.


Assuntos
Cádmio/toxicidade , Produtos Agrícolas/crescimento & desenvolvimento , Resistência a Medicamentos , Epigênese Genética/efeitos dos fármacos , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Produtos Agrícolas/efeitos dos fármacos , Produtos Agrícolas/genética , Metilação de DNA/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Código das Histonas/efeitos dos fármacos , MicroRNAs/efeitos dos fármacos , MicroRNAs/genética , RNA de Plantas/efeitos dos fármacos , RNA de Plantas/genética
14.
Int J Mol Sci ; 22(14)2021 Jul 06.
Artigo em Inglês | MEDLINE | ID: mdl-34298866

RESUMO

MicroRNA (miRNA) expression analysis is very important for investigating its functions. To date, no research on reference genes (RGs) for miRNAs in gymnosperms, including Cryptomeria fortunei, has been reported. Here, ten miRNAs (i.e., pab-miR159a, cln-miR162, cas-miR166d, pab-miR395b, ppt-miR894, cln-miR6725, novel1, novel6, novel14 and novel16) and three common RGs (U6, 5S and 18S) were selected as candidate RGs. qRT-PCR was used to analyse their expressions in C. fortunei under various experimental conditions, including multiple stresses (cold, heat, drought, salt, abscisic acid and gibberellin) and in various tissues (roots, stems, tender needles, cones and seeds). Four algorithms (delta Ct, geNorm, NormFinder and BestKeeper) were employed to assess the stability of candidate RG expression; the geometric mean and RefFinder program were used to comprehensively evaluate RG stability. According to the results, novel16, cln-miR6725, novel1 and U6 were the most stable RGs for studying C. fortunei miRNA expression. In addition, the expression of three target miRNAs (aly-miR164c-5p, aly-miR168a-5p and smo-miR396) was examined to verify that the selected RGs are suitable for miRNA expression normalisation. This study may aid further investigations of miRNA expression/function in the response of C. fortunei to abiotic stress and provides an important basis for the standardisation of miRNA expression in other gymnosperm species.


Assuntos
Cryptomeria/genética , Regulação da Expressão Gênica de Plantas/genética , MicroRNAs/genética , Estresse Fisiológico/genética , China , Secas , Perfilação da Expressão Gênica/métodos , Genes de Plantas/genética , RNA de Plantas/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Padrões de Referência , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos
15.
Int J Mol Sci ; 22(13)2021 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-34209661

RESUMO

Self-incompatibility (SI) is conserved among members of the Brassicaceae plant family. This trait is controlled epigenetically by the dominance hierarchy of the male determinant alleles. We previously demonstrated that a single small RNA (sRNA) gene is sufficient to control the linear dominance hierarchy in Brassica rapa and proposed a model in which a homology-based interaction between sRNAs and target sites controls the complicated dominance hierarchy of male SI determinants. In Arabidopsis halleri, male dominance hierarchy is reported to have arisen from multiple networks of sRNA target gains and losses. Despite these findings, it remains unknown whether the molecular mechanism underlying the dominance hierarchy is conserved among Brassicaceae. Here, we identified sRNAs and their target sites that can explain the linear dominance hierarchy of Arabidopsis lyrata, a species closely related to A. halleri. We tested the model that we established in Brassica to explain the linear dominance hierarchy in A. lyrata. Our results suggest that the dominance hierarchy of A. lyrata is also controlled by a homology-based interaction between sRNAs and their targets.


Assuntos
Arabidopsis/genética , Epistasia Genética , Regulação da Expressão Gênica de Plantas , Genes Dominantes , Recombinação Homóloga , RNA de Plantas , Predomínio Social , Alelos , Genótipo , Haplótipos , Conformação de Ácido Nucleico
16.
Int J Mol Sci ; 22(13)2021 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-34281265

RESUMO

The demonstration that spray-induced gene silencing (SIGS) can confer strong disease resistance, bypassing the laborious and time-consuming transgenic expression of double-stranded (ds)RNA to induce the gene silencing of pathogenic targets, was ground-breaking. However, future field applications will require fundamental mechanistic knowledge of dsRNA uptake, processing, and transfer. There is increasing evidence that extracellular vesicles (EVs) mediate the transfer of transgene-derived small interfering (si)RNAs in host-induced gene silencing (HIGS) applications. In this study, we establish a protocol for barley EV isolation and assess the possibilities for EVs regarding the translocation of sprayed dsRNA from barley (Hordeum vulgare) to its interacting fungal pathogens. We found barley EVs that were 156 nm in size, containing predominantly 21 and 19 nucleotide (nts) siRNAs, starting with a 5'-terminal Adenine. Although a direct comparison of the RNA cargo between HIGS and SIGS EV isolates is improper given their underlying mechanistic differences, we identified sequence-identical siRNAs in both systems. Overall, the number of siRNAs isolated from the EVs of dsRNA-sprayed barley plants with sequence complementarity to the sprayed dsRNA precursor was low. However, whether these few siRNAs are sufficient to induce the SIGS of pathogenic target genes requires further research. Taken together, our results raise the possibility that EVs may not be mandatory for the spray-delivered siRNA uptake and induction of SIGS.


Assuntos
Proteção de Cultivos/métodos , Hordeum/genética , Hordeum/microbiologia , RNA Interferente Pequeno/administração & dosagem , Família 3 do Citocromo P450/genética , Resistência à Doença/genética , Vesículas Extracelulares/genética , Vesículas Extracelulares/microbiologia , Inativação Gênica , Interações entre Hospedeiro e Microrganismos/genética , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controle , Interferência de RNA , RNA de Plantas/genética , RNA de Plantas/isolamento & purificação , RNA Interferente Pequeno/isolamento & purificação
17.
Planta ; 254(2): 31, 2021 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-34283297

RESUMO

MAIN CONCLUSION: Comparative analysis of miRNAs and their gene targets between the evergreen and yellowish-brown Cryptomeria fortunei phenotypes in cold winters suggests a possible role of miRNA-regulated pathways in needle color. Cryptomeria fortunei (Chinese cedar) is a conifer tree of considerable economic, ornamental and ecological importance. Despite the evergreen nature of C. fortunei, most needles turn yellowish- or reddish-brown in winter. The roles of microRNAs (miRNAs) in regulating pigment biosynthesis in color-leafed plants have been widely investigated. However, whether or not an miRNA-mediated staged discoloration mechanism exists in evergreen C. fortunei is currently unknown. In this study, we deciphered the microRNAs landscape in overwintering C. fortunei needles using high-throughput sequencing. A total of 517 known and 212 novel miRNA mature/star sequences, including 233 differentially expressed miRNAs, were identified. Based on integrated transcriptome and miRNA analysis, 2702 target unigenes of the miRNAs were predicted and these targets were significantly enriched in pigment-related biosynthesis pathways. A miRNA-target pigment biosynthesis regulatory network was then constructed, and its module miRNA (ath-miR858b, aly-miR858-3p, cme-miR828 and novel33_mature)-MYBs (v-myb avian myeloblastosis viral oncogene homolog) appeared to be a key factor regulating needle discoloration in C. fortunei. These miRNA-MYBs were further confirmed by degradome sequencing. Overall, these findings provide new insight into the posttranscriptional regulatory mechanism of leaf/needle discoloration in gymnosperms and may contribute to the miRNA-mediated genetic improvement of evergreen C. fortunei needles.


Assuntos
Cryptomeria , MicroRNAs , China , Regulação da Expressão Gênica de Plantas , Sequenciamento de Nucleotídeos em Larga Escala , MicroRNAs/genética , Agulhas , RNA de Plantas/genética , Árvores/genética
18.
Int J Mol Sci ; 22(14)2021 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-34299307

RESUMO

Crop yield is severely affected by biotic and abiotic stresses. Plants adapt to these stresses mainly through gene expression reprogramming at the transcriptional and post-transcriptional levels. Recently, the exogenous application of double-stranded RNAs (dsRNAs) and RNA interference (RNAi) technology has emerged as a sustainable and publicly acceptable alternative to genetic transformation, hence, small RNAs (micro-RNAs and small interfering RNAs) have an important role in combating biotic and abiotic stresses in plants. RNAi limits the transcript level by either suppressing transcription (transcriptional gene silencing) or activating sequence-specific RNA degradation (post-transcriptional gene silencing). Using RNAi tools and their respective targets in abiotic stress responses in many crops is well documented. Many miRNAs families are reported in plant tolerance response or adaptation to drought, salinity, and temperature stresses. In biotic stress, the spray-induced gene silencing (SIGS) provides an intelligent method of using dsRNA as a trigger to silence target genes in pests and pathogens without producing side effects such as those caused by chemical pesticides. In this review, we focus on the potential of SIGS as the most recent application of RNAi in agriculture and point out the trends, challenges, and risks of production technologies. Additionally, we provide insights into the potential applications of exogenous RNAi against biotic stresses. We also review the current status of RNAi/miRNA tools and their respective targets on abiotic stress and the most common responsive miRNA families triggered by stress conditions in different crop species.


Assuntos
Produtos Agrícolas/genética , Interferência de RNA , Animais , Produção Agrícola/métodos , Proteção de Cultivos/métodos , Inativação Gênica , Controle de Insetos , Insetos/genética , Insetos/patogenicidade , MicroRNAs/genética , Defesa das Plantas contra Herbivoria/genética , RNA de Cadeia Dupla/genética , RNA de Plantas/genética , RNA Interferente Pequeno/genética , Estresse Fisiológico/genética
19.
Annu Rev Phytopathol ; 59: 265-288, 2021 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-34077241

RESUMO

Gene silencing guided by small RNAs governs a broad range of cellular processes in eukaryotes. Small RNAs are important components of plant immunity because they contribute to pathogen-triggered transcription reprogramming and directly target pathogen RNAs. Recent research suggests that silencing of pathogen genes by plant small RNAs occurs not only during viral infection but also in nonviral pathogens through a process termed host-induced gene silencing, which involves trans-species small RNA trafficking. Similarly, small RNAs are also produced by eukaryotic pathogens and regulate virulence. This review summarizes the small RNA pathways in both plants and filamentous pathogens, including fungi and oomycetes, and discusses their role in host-pathogen interactions. We highlight secondarysmall interfering RNAs of plants as regulators of immune receptor gene expression and executors of host-induced gene silencing in invading pathogens. The current status and prospects of trans-species gene silencing at the host-pathogen interface are discussed.


Assuntos
Doenças das Plantas , Imunidade Vegetal , Interações Hospedeiro-Patógeno/genética , Imunidade Vegetal/genética , Interferência de RNA , RNA de Plantas , Virulência
20.
Methods Mol Biol ; 2323: 75-97, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34086275

RESUMO

Preparative synthesis of RNA is a challenging task that is usually accomplished by either chemical or enzymatic polymerization of ribonucleotides in vitro. Herein, we describe an alternative approach in which RNAs of interest are expressed as a fusion with a 5S rRNA-derived scaffold. The scaffold provides protection against cellular ribonucleases resulting in cellular accumulations comparable to those of regular ribosomal RNAs. After isolation of the chimeric RNA from the cells, the scaffold can be removed, if necessary, by deoxyribozyme-catalyzed cleavage followed by preparative electrophoretic separation of the reaction products. The protocol is designed for sustained production of high quality RNA on the milligram scale.


Assuntos
Clonagem Molecular/métodos , RNA Ribossômico 5S , RNA/biossíntese , Sequência de Bases , DNA Catalítico/metabolismo , Eletroforese em Gel de Ágar , Eletroforese em Gel de Poliacrilamida , Escherichia coli/isolamento & purificação , Escherichia coli/metabolismo , Conformação de Ácido Nucleico , Pennisetum/genética , Plasmídeos/genética , Plasmídeos/isolamento & purificação , RNA/genética , RNA/isolamento & purificação , RNA de Plantas/genética , RNA Ribossômico 5S/genética , Transformação Bacteriana , Vibrio/genética
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