RESUMO
Intrinsically disordered proteins (IDPs) and proteins with intrinsically disordered regions (IDRs) possess low sequence complexity of amino acids and display non-globular tertiary structures. They can act as scaffolds, form regulatory hubs, or trigger biomolecular condensation to control diverse aspects of biology. Emerging evidence has recently implicated critical roles of IDPs and IDR-contained proteins in nuclear transcription and cytoplasmic post-transcriptional processes, among other molecular functions. We here summarize the concepts and organizing principles of IDPs. We then illustrate recent progress in understanding the roles of key IDPs in machineries that regulate transcriptional and post-transcriptional gene silencing (PTGS) in plants, aiming at highlighting new modes of action of IDPs in controlling biological processes.
Assuntos
Proteínas Intrinsicamente Desordenadas , Proteínas Intrinsicamente Desordenadas/genética , Proteínas Intrinsicamente Desordenadas/química , Proteínas Intrinsicamente Desordenadas/metabolismo , Plantas/genética , Plantas/metabolismo , Inativação Gênica , Conformação ProteicaRESUMO
The shoot apical meristem (SAM) comprises a group of undifferentiated cells that divide to maintain the plant meristem and also give rise to all shoot organs. SAM fate is specified by class III HOMEODOMAIN-LEUCINE ZIPPER (HD-ZIP III) transcription factors, which are targets of miR166/165. In Arabidopsis, AGO10 is a critical regulator of SAM maintenance, and here we demonstrate that AGO10 specifically interacts with miR166/165. The association is determined by a distinct structure of the miR166/165 duplex. Deficient loading of miR166 into AGO10 results in a defective SAM. Notably, the miRNA-binding ability of AGO10, but not its catalytic activity, is required for SAM development, and AGO10 has a higher binding affinity for miR166 than does AGO1, a principal contributor to miRNA-mediated silencing. We propose that AGO10 functions as a decoy for miR166/165 to maintain the SAM, preventing their incorporation into AGO1 complexes and the subsequent repression of HD-ZIP III gene expression.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Meristema/crescimento & desenvolvimento , MicroRNAs/genética , RNA de Plantas/genética , Arabidopsis/genética , Proteínas Argonautas , Brotos de PlantaRESUMO
Intrinsically disordered proteins (IDPs) SAID1/2 are hypothetic dentin sialophosphoprotein-like proteins, but their true functions are unknown. Here, we identified SAID1/2 as negative regulators of SERRATE (SE), a core factor in miRNA biogenesis complex (microprocessor). Loss-of-function double mutants of said1; said2 caused pleiotropic developmental defects and thousands of differentially expressed genes that partially overlapped with those in se. said1; said2 also displayed increased assembly of microprocessor and elevated accumulation of microRNAs (miRNAs). Mechanistically, SAID1/2 promote pre-mRNA processing 4 kinase A-mediated phosphorylation of SE, causing its degradation in vivo. Unexpectedly, SAID1/2 have strong binding affinity to hairpin-structured pri-miRNAs and can sequester them from SE. Moreover, SAID1/2 directly inhibit pri-miRNA processing by microprocessor in vitro. Whereas SAID1/2 did not impact SE subcellular compartmentation, the proteins themselves exhibited liquid-liquid phase condensation that is nucleated on SE. Thus, we propose that SAID1/2 reduce miRNA production through hijacking pri-miRNAs to prevent microprocessor activity while promoting SE phosphorylation and its destabilization in Arabidopsis.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Proteínas Intrinsicamente Desordenadas , MicroRNAs , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas Intrinsicamente Desordenadas/genética , Proteínas Intrinsicamente Desordenadas/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Ligação a RNA/metabolismo , Processamento Pós-Transcricional do RNA , MicroRNAs/metabolismo , Ribonuclease III/metabolismo , Regulação da Expressão Gênica de PlantasRESUMO
U-insertion/deletion (U-indel) RNA editing in trypanosome mitochondria is directed by guide RNAs (gRNAs). This editing may developmentally control respiration in bloodstream forms (BSF) and insect procyclic forms (PCF). Holo-editosomes include the accessory RNA Editing Substrate Binding Complex (RESC) and RNA Editing Helicase 2 Complex (REH2C), but the specific proteins controlling differential editing remain unknown. Also, RNA editing appears highly error prone because most U-indels do not match the canonical pattern. However, despite extensive non-canonical editing of unknown functions, accurate canonical editing is required for normal cell growth. In PCF, REH2C controls editing fidelity in RESC-bound mRNAs. Here, we report that KREH2, a REH2C-associated helicase, developmentally controls programmed non-canonical editing, including an abundant 3' element in ATPase subunit 6 (A6) mRNA. The 3' element sequence is directed by a proposed novel regulatory gRNA. In PCF, KREH2 RNAi-knockdown up-regulates the 3' element, which establishes a stable structure hindering element removal by canonical initiator-gRNA-directed editing. In BSF, KREH2-knockdown does not up-regulate the 3' element but reduces its high abundance. Thus, KREH2 differentially controls extensive non-canonical editing and associated RNA structure via a novel regulatory gRNA, potentially hijacking factors as a 'molecular sponge'. Furthermore, this gRNA is bifunctional, serving in canonical CR4 mRNA editing whilst installing a structural element in A6 mRNA.
Assuntos
Trypanosoma brucei brucei , Trypanosoma , RNA Mensageiro/metabolismo , RNA Helicases/genética , RNA Helicases/metabolismo , Trypanosoma brucei brucei/genética , Trypanosoma brucei brucei/metabolismo , Trypanosoma/genética , RNA/genética , RNA de Protozoário/genética , RNA de Protozoário/metabolismoRESUMO
Chromatin remodelling factors (CHRs) typically function to alter chromatin structure. CHRs also reside in ribonucleoprotein complexes, but little is known about their RNA-related functions. Here we show that CHR2 (also known as BRM), the ATPase subunit of the large switch/sucrose non-fermentable (SWI/SNF) complex, is a partner of the Microprocessor component Serrate (SE). CHR2 promotes the transcription of primary microRNA precursors (pri-miRNAs) while repressing miRNA accumulation in vivo. Direct interaction with SE is required for post-transcriptional inhibition of miRNA accumulation by CHR2 but not for its transcriptional activity. CHR2 can directly bind to and unwind pri-miRNAs and inhibit their processing, and this inhibition requires the remodelling and helicase activity of CHR2 in vitro and in vivo. Furthermore, the secondary structures of pri-miRNAs differed between wild-type Arabidopsis thaliana and chr2 mutants. We conclude that CHR2 accesses pri-miRNAs through SE and remodels their secondary structures, preventing downstream processing by DCL1 and HYL1. Our study uncovers pri-miRNAs as a substrate of CHR2, and an additional regulatory layer upstream of Microprocessor activity to control miRNA accumulation.
Assuntos
Adenosina Trifosfatases/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , MicroRNAs/biossíntese , Proteínas de Ligação a RNA/metabolismo , Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , MicroRNAs/genética , MicroRNAs/metabolismo , Ligação Proteica , Dobramento de RNA , Processamento Pós-Transcricional do RNA , Transcrição GênicaRESUMO
RNA helicases (RHs) are a family of ubiquitous enzymes that alter RNA structures and remodel ribonucleoprotein complexes typically using energy from the hydrolysis of ATP. RHs are involved in various aspects of RNA processing and metabolism, exemplified by transcriptional regulation, pre-mRNA splicing, miRNA biogenesis, liquid-liquid phase separation, and rRNA biogenesis, among other molecular processes. Through these mechanisms, RHs contribute to vegetative and reproductive growth, as well as abiotic and biotic stress responses throughout the life cycle in plants. In this review, we systematically characterize RH-featured domains and signature motifs in Arabidopsis. We also summarize the functions and mechanisms of RHs in various biological processes in plants with a focus on DEAD-box and DEAH-box RNA helicases, aiming to present the latest understanding of RHs in plant biology.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , RNA Helicases DEAD-box/genética , Plantas/genética , Plantas/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Splicing de RNARESUMO
Among many glycoproteins within the plant secretory system, KORRIGAN1 (KOR1), a membrane-anchored endo-ß-1,4-glucanase involved in cellulose biosynthesis, provides a link between N-glycosylation, cell wall biosynthesis, and abiotic stress tolerance. After insertion into the endoplasmic reticulum, KOR1 cycles between the trans-Golgi network (TGN) and the plasma membrane (PM). From the TGN, the protein is targeted to growing cell plates during cell division. These processes are governed by multiple sequence motifs and also host genotypes. Here, we investigated the interaction and hierarchy of known and newly identified sorting signals in KOR1 and how they affect KOR1 transport at various stages in the secretory pathway. Conventional steady-state localization showed that structurally compromised KOR1 variants were directed to tonoplasts. In addition, a tandem fluorescent timer technology allowed for differential visualization of young versus aged KOR1 proteins, enabling the analysis of single-pass transport through the secretory pathway. Observations suggest the presence of multiple checkpoints/branches during KOR1 trafficking, where the destination is determined based on KOR1's sequence motifs and folding status. Moreover, growth analyses of dominant PM-confined KOR1-L48L49âA48A49 variants revealed the importance of active removal of KOR1 from the PM during salt stress, which otherwise interfered with stress acclimation.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Celulase/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas de Membrana/metabolismo , Estresse Salino/fisiologia , Tolerância ao Sal/fisiologia , Rede trans-Golgi/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Membrana Celular/metabolismo , Parede Celular/metabolismo , Celulase/genética , Celulose/metabolismo , Regulação da Expressão Gênica de Plantas , Glicosilação , Complexo de Golgi/metabolismo , Proteínas de Membrana/genética , Mutação , Raízes de Plantas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas , Transporte Proteico , Controle de Qualidade , Estresse Salino/genética , Tolerância ao Sal/genética , Sais/metabolismo , Alinhamento de Sequência , TranscriptomaRESUMO
Trypanosome U-insertion/deletion RNA editing in mitochondrial mRNAs involves guide RNAs (gRNAs) and the auxiliary RNA editing substrate binding complex (RESC) and RNA editing helicase 2 complex (REH2C). RESC and REH2C stably copurify with editing mRNAs but the functional interplay between these complexes remains unclear. Most steady-state mRNAs are partially edited and include misedited "junction" regions that match neither pre-mRNA nor fully edited transcripts. Editing specificity is central to mitochondrial RNA maturation and function, but its basic control mechanisms remain unclear. Here we applied a novel nucleotide-resolution RNA-seq approach to examine ribosomal protein subunit 12 (RPS12) and ATPase subunit 6 (A6) mRNA transcripts. We directly compared transcripts associated with RESC and REH2C to those found in total mitochondrial RNA. RESC-associated transcripts exhibited site-preferential enrichments in total and accurate edits. REH2C loss-of-function induced similar substrate-specific and site-specific editing effects in total and RESC-associated RNA. It decreased total editing primarily at RPS12 5' positions but increased total editing at examined A6 3' positions. REH2C loss-of-function caused site-preferential loss of accurate editing in both transcripts. However, changes in total or accurate edits did not necessarily involve common sites. A few 5' nucleotides of the initiating gRNA (gRNA-1) directed accurate editing in both transcripts. However, in RPS12, two conserved 3'-terminal adenines in gRNA-1 could direct a noncanonical 2U-insertion that causes major pausing in 3'-5' progression. In A6, a noncanonical sequence element that depends on REH2C in a region normally targeted by the 3' half of gRNA-1 may hinder early editing progression. Overall, we defined transcript-specific effects of REH2C loss.
Assuntos
Proteínas de Protozoários/metabolismo , Edição de RNA , RNA Mensageiro/metabolismo , RNA Mitocondrial/metabolismo , RNA de Protozoário/metabolismo , Trypanosoma brucei brucei/metabolismo , Trypanosoma/metabolismo , Animais , Proteínas de Protozoários/genética , RNA Guia de Cinetoplastídeos , RNA Mensageiro/genética , RNA Mitocondrial/genética , RNA de Protozoário/genética , RNA-Seq , Especificidade por Substrato , Trypanosoma/genética , Trypanosoma brucei brucei/genéticaRESUMO
Plant viruses have evolved multiple strategies to overcome host defense to establish an infection. Here, we identified two components of a host mitogen-activated protein kinase (MAPK) cascade, MKK2 and MPK4, as bona fide targets of the ßC1 protein encoded by the betasatellite of tomato yellow leaf curl China virus (TYLCCNV). ßC1 interacts with the kinase domain of MKK2 and inhibits its activity. In vivo, ßC1 suppresses flagellin-induced MAPK activation and downstream responses by targeting MKK2. Furthermore, ßC1 also interacts with MPK4 and inhibits its kinase activity. TYLCCNV infection induces the activation of the MAPK cascade, mutation in MKK2 or MPK4 renders the plant more susceptible to TYLCCNV, and can complement the lack of ßC1. This work shows for the first time that a plant virus both activates and suppresses a MAPK cascade, and the discovery of the ability of ßC1 to selectively interfere with the host MAPK activation illustrates a novel virulence function and counter-host defense mechanism of geminiviruses.
Assuntos
Arabidopsis/imunologia , Geminiviridae/imunologia , Interações Hospedeiro-Patógeno/imunologia , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Nicotiana/imunologia , Proteínas Virais/metabolismo , Arabidopsis/metabolismo , Arabidopsis/virologia , Proteínas de Arabidopsis/antagonistas & inibidores , Geminiviridae/metabolismo , Geminiviridae/patogenicidade , Fosforilação , Nicotiana/metabolismo , Nicotiana/virologiaRESUMO
The majority of the genome is transcribed to RNA in living organisms. RNA transcripts can form astonishing arrays of secondary and tertiary structures via Watson-Crick, Hoogsteen, or wobble base pairing. In vivo, RNA folding is not a simple thermodynamic event of minimizing free energy. Instead, the process is constrained by transcription, RNA-binding proteins, steric factors, and the microenvironment. RNA secondary structure (RSS) plays myriad roles in numerous biological processes, such as RNA processing, stability, transportation, and translation in prokaryotes and eukaryotes. Emerging evidence has also implicated RSS in RNA trafficking, liquid-liquid phase separation, and plant responses to environmental variations such as temperature and salinity. At molecular level, RSS is correlated with splicing, polyadenylation, protein synthesis, and miRNA biogenesis and functions. In this review, we summarize newly reported methods for probing RSS in vivo and functions and mechanisms of RSS in plant physiology.
Assuntos
Processamento Pós-Transcricional do RNA , RNA , Pareamento de Bases , Biologia , Conformação de Ácido Nucleico , RNA/metabolismo , Splicing de RNA , RNA de Plantas/genética , RNA de Plantas/metabolismoRESUMO
The whitefly-transmitted geminiviruses induce severe developmental abnormalities in plants. Geminivirus-encoded C4 protein functions as one of viral symptom determinants that could induce abnormal cell division. However, the molecular mechanism by which C4 contributes to cell division induction remains unclear. Here we report that tomato leaf curl Yunnan virus (TLCYnV) C4 interacts with a glycogen synthase kinase 3 (GSK3)/SHAGGY-like kinase, designed NbSKη, in Nicotiana benthamiana. Pro32, Asn34 and Thr35 of TLCYnV C4 are critical for its interaction with NbSKη and required for C4-induced typical symptoms. Interestingly, TLCYnV C4 directs NbSKη to the membrane and reduces the nuclear-accumulation of NbSKη. The relocalization of NbSKη impairs phosphorylation dependent degradation on its substrate-Cyclin D1.1 (NbCycD1;1), thereby increasing the accumulation level of NbCycD1;1 and inducing the cell division. Moreover, NbSKη-RNAi, 35S::NbCycD1;1 transgenic N. benthamiana plants have the similar phenotype as 35S::C4 transgenic N. benthamiana plants on callus-like tissue formation resulted from abnormal cell division induction. Thus, this study provides new insights into mechanism of how a viral protein hijacks NbSKη to induce abnormal cell division in plants.
Assuntos
Begomovirus/metabolismo , Ciclina D1/metabolismo , Quinase 3 da Glicogênio Sintase/metabolismo , Nicotiana/metabolismo , Proteínas de Plantas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Virais/metabolismo , Agrobacterium tumefaciens/fisiologia , Begomovirus/patogenicidade , Divisão Celular , Ciclina D1/química , Deleção de Genes , Proteínas Luminescentes/química , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Fosforilação , Filogenia , Folhas de Planta/genética , Folhas de Planta/metabolismo , Folhas de Planta/microbiologia , Folhas de Planta/ultraestrutura , Proteínas de Plantas/antagonistas & inibidores , Proteínas de Plantas/química , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Plantas Geneticamente Modificadas/microbiologia , Plantas Geneticamente Modificadas/ultraestrutura , Mutação Puntual , Complexo de Endopeptidases do Proteassoma/metabolismo , Complexo de Endopeptidases do Proteassoma/ultraestrutura , Domínios e Motivos de Interação entre Proteínas , Processamento de Proteína Pós-Traducional , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/genética , Estabilidade Proteica , Transporte Proteico , Proteólise , Interferência de RNA , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/metabolismo , Nicotiana/genética , Nicotiana/microbiologia , Nicotiana/ultraestrutura , Proteínas Virais/químicaRESUMO
Phosphorylation of the RNA polymerase II (Pol II) C-terminal domain (CTD) regulates transcription of protein-coding mRNAs and noncoding RNAs. CTD function in transcription of protein-coding RNAs has been studied extensively, but its role in plant noncoding RNA transcription remains obscure. Here, using Arabidopsis thaliana CTD PHOSPHATASE-LIKE4 knockdown lines (CPL4RNAi ), we showed that CPL4 functions in genome-wide, conditional production of 3'-extensions of small nuclear RNAs (snRNAs) and biogenesis of novel transcripts from protein-coding genes downstream of the snRNAs (snRNA-downstream protein-coding genes [snR-DPGs]). Production of snR-DPGs required the Pol II snRNA promoter (PIIsnR), and CPL4RNAi plants showed increased read-through of the snRNA 3'-end processing signal, leading to continuation of transcription downstream of the snRNA gene. We also discovered an unstable, intermediate-length RNA from the SMALL SCP1-LIKE PHOSPHATASE14 locus (imRNASSP14 ), whose expression originated from the 5' region of a protein-coding gene. Expression of the imRNASSP14 was driven by a PIIsnR and was conditionally 3'-extended to produce an mRNA. In the wild type, salt stress induced the snRNA-to-snR-DPG switch, which was associated with alterations of Pol II-CTD phosphorylation at the target loci. The snR-DPG transcripts occur widely in plants, suggesting that the transcriptional snRNA-to-snR-DPG switch may be a ubiquitous mechanism to regulate plant gene expression in response to environmental stresses.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Fosfoproteínas Fosfatases/metabolismo , RNA Mensageiro/biossíntese , RNA Nuclear Pequeno/biossíntese , Estresse Salino/fisiologia , Arabidopsis/genética , Elementos de DNA Transponíveis/genética , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genes de Plantas , Loci Gênicos , Luciferases/metabolismo , Modelos Biológicos , Mutação/genética , Motivos de Nucleotídeos/genética , Fases de Leitura Aberta/genética , Fosforilação , Plantas Geneticamente Modificadas , RNA Polimerase II/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA de Plantas/metabolismo , RNA Nuclear Pequeno/genética , Fatores de Transcrição/metabolismo , Regulação para Cima/genéticaRESUMO
Transcripts have intrinsic propensity to form stable secondary structure that is fundamental to regulate RNA transcription, splicing, translation, RNA localization and turnover. Numerous methods that integrate chemical reactions with next-generation sequencing (NGS) have been applied to study in vivo RNA structure, providing new insights into RNA biology. Dimethyl sulfate (DMS) probing coupled with mutational profiling through NGS (DMS-MaPseq) is a newly developed method for revealing genome-wide or target-specific RNA structure. Herein, we present our experimental protocol of a modified DMS-MaPseq method for plant materials. The DMS treatment condition was optimized, and library preparation procedures were simplified. We also provided custom scripts for bioinformatic analysis of genome-wide DMS-MaPseq data. Bioinformatic results showed that our method could generate high-quality and reproducible data. Further, we assessed sequencing depth and coverage for genome-wide RNA structure profiling in Arabidopsis, and provided two examples of in vivo structure of mobile RNAs. We hope that our modified DMS-MaPseq method will serve as a powerful tool for analyzing in vivo RNA structurome in plants.
Assuntos
Arabidopsis/genética , Biologia Computacional/métodos , Genoma de Planta , Sequenciamento de Nucleotídeos em Larga Escala/métodos , RNA Mensageiro/química , RNA de Plantas/química , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Pareamento de Bases , Sequência de Bases , Complexos de Proteínas Captadores de Luz/genética , Complexos de Proteínas Captadores de Luz/metabolismo , Mutação , Conformação de Ácido Nucleico , Complexo de Proteína do Fotossistema II/genética , Complexo de Proteína do Fotossistema II/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA de Plantas/genética , RNA de Plantas/metabolismo , Ésteres do Ácido Sulfúrico/químicaRESUMO
MicroRNA (miRNA) is processed from primary transcripts with hairpin structures (pri-miRNAs) by microprocessors in the nucleus. How cytoplasmic-borne microprocessor components are transported into the nucleus to fulfill their functions remains poorly understood. Here, we report KETCH1 (karyopherin enabling the transport of the cytoplasmic HYL1) as a partner of hyponastic leaves 1 (HYL1) protein, a core component of microprocessor in Arabidopsis and functional counterpart of DGCR8/Pasha in animals. Null mutation of ketch1 is embryonic-lethal, whereas knockdown mutation of ketch1 caused morphological defects, reminiscent of mutants in the miRNA pathway. ketch1 knockdown mutation also substantially reduced miRNA accumulation, but did not alter nuclear-cytoplasmic shuttling of miRNAs. Rather, the mutation significantly reduced nuclear portion of HYL1 protein and correspondingly compromised the pri-miRNA processing in the nucleus. We propose that KETCH1 transports HYL1 from the cytoplasm to the nucleus to constitute functional microprocessor in Arabidopsis This study provides insight into the largely unknown nuclear-cytoplasmic trafficking process of miRNA biogenesis components through eukaryotes.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Núcleo Celular/metabolismo , MicroRNAs/metabolismo , Proteínas de Ligação a RNA/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Núcleo Celular/genética , Regulação da Expressão Gênica de Plantas , Carioferinas , MicroRNAs/genética , Mutação , Plantas Geneticamente Modificadas , Transporte Proteico , Processamento Pós-Transcricional do RNA , Proteínas de Ligação a RNA/genética , Nicotiana/genética , Nicotiana/metabolismoRESUMO
Like metazoans, plants use small regulatory RNAs (sRNAs) to direct gene expression. Several classes of sRNAs, which are distinguished by their origin and biogenesis, exist in plants. Among them, microRNAs (miRNAs) and trans-acting small interfering RNAs (ta-siRNAs) mainly inhibit gene expression at post-transcriptional levels. In the past decades, plant miRNAs and ta-siRNAs have been shown to be essential for numerous developmental processes, including growth and development of shoots, leaves, flowers, roots and seeds, among others. In addition, miRNAs and ta-siRNAs are also involved in the plant responses to abiotic and biotic stresses, such as drought, temperature, salinity, nutrient deprivation, bacteria, virus and others. This review summarizes the roles of miRNAs and ta-siRNAs in plant physiology and development.
Assuntos
MicroRNAs/genética , RNA de Plantas/genética , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , RNA Interferente Pequeno/genética , Estresse Fisiológico/genética , Estresse Fisiológico/fisiologiaRESUMO
Global climate change, increasingly erratic weather and a burgeoning global population are significant threats to the sustainability of future crop production. There is an urgent need for the development of robust measures that enable crops to withstand the uncertainty of climate change whilst still producing maximum yields. Resurrection plants possess the unique ability to withstand desiccation for prolonged periods, can be restored upon watering and represent great potential for the development of stress tolerant crops. Here, we describe the remarkable stress characteristics of Tripogon loliiformis, an uncharacterised resurrection grass and close relative of the economically important cereals, rice, sorghum, and maize. We show that T. loliiformis survives extreme environmental stress by implementing autophagy to prevent Programmed Cell Death. Notably, we identified a novel role for trehalose in the regulation of autophagy in T.loliiformis. Transcriptome, Gas Chromatography Mass Spectrometry, immunoblotting and confocal microscopy analyses directly linked the accumulation of trehalose with the onset of autophagy in dehydrating and desiccated T. loliiformis shoots. These results were supported in vitro with the observation of autophagosomes in trehalose treated T. loliiformis leaves; autophagosomes were not detected in untreated samples. Presumably, once induced, autophagy promotes desiccation tolerance in T.loliiformis, by removal of cellular toxins to suppress programmed cell death and the recycling of nutrients to delay the onset of senescence. These findings illustrate how resurrection plants manipulate sugar metabolism to promote desiccation tolerance and may provide candidate genes that are potentially useful for the development of stress tolerant crops.
Assuntos
Autofagia/genética , Craterostigma/crescimento & desenvolvimento , Transcriptoma/genética , Trealose/metabolismo , Mudança Climática , Craterostigma/genética , Dessecação , Oryza , Folhas de Planta/genética , Folhas de Planta/metabolismo , Poaceae/genética , Estresse Fisiológico/genética , Trealose/genética , ÁguaRESUMO
BACKGROUND: RNA secondary structure (RSS) can influence the regulation of transcription, RNA processing, and protein synthesis, among other processes. 3' untranslated regions (3' UTRs) of mRNA also hold the key for many aspects of gene regulation. However, there are often contradictory results regarding the roles of RSS in 3' UTRs in gene expression in different organisms and/or contexts. RESULTS: Here, we incidentally observe that the primary substrate of miR159a (pri-miR159a), when embedded in a 3' UTR, could promote mRNA accumulation. The enhanced expression is attributed to the earlier polyadenylation of the transcript within the hybrid pri-miR159a-3' UTR and, resultantly, a poorly structured 3' UTR. RNA decay assays indicate that poorly structured 3' UTRs could promote mRNA stability, whereas highly structured 3' UTRs destabilize mRNA in vivo. Genome-wide DMS-MaPseq also reveals the prevailing inverse relationship between 3' UTRs' RSS and transcript accumulation in the transcriptomes of Arabidopsis, rice, and even human. Mechanistically, transcripts with highly structured 3' UTRs are preferentially degraded by 3'-5' exoribonuclease SOV and 5'-3' exoribonuclease XRN4, leading to decreased expression in Arabidopsis. Finally, we engineer different structured 3' UTRs to an endogenous FT gene and alter the FT-regulated flowering time in Arabidopsis. CONCLUSIONS: We conclude that highly structured 3' UTRs typically cause reduced accumulation of the harbored transcripts in Arabidopsis. This pattern extends to rice and even mammals. Furthermore, our study provides a new strategy of engineering the 3' UTRs' RSS to modify plant traits in agricultural production and mRNA stability in biotechnology.
Assuntos
Arabidopsis , Exorribonucleases , Animais , Humanos , Regiões 3' não Traduzidas , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Exorribonucleases/genética , Exorribonucleases/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Regulação da Expressão Gênica , Mamíferos/genéticaRESUMO
MicroRNAs (miRNAs) are produced from highly structured primary transcripts (pri-miRNAs) and regulate numerous biological processes in eukaryotes. Due to the extreme heterogeneity of these structures, the initial processing sites of plant pri-miRNAs and the structural rules that determine their processing have been predicted for many miRNAs but remain elusive for others. Here we used semi-active DCL1 mutants and advanced degradome-sequencing strategies to accurately identify the initial processing sites for 147 of 326 previously annotated Arabidopsis miRNAs and to illustrate their associated pri-miRNA cleavage patterns. Elucidating the in vivo RNA secondary structures of 73 pri-miRNAs revealed that about 95% of them differ from in silico predictions, and that the revised structures offer clearer interpretation of the processing sites and patterns. Finally, DCL1 partners Serrate and HYL1 could synergistically and independently impact processing patterns and in vivo RNA secondary structures of pri-miRNAs. Together, our work sheds light on the precise processing mechanisms of plant pri-miRNAs.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , MicroRNAs , RNA de Plantas , Arabidopsis/genética , Arabidopsis/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , RNA de Plantas/genética , RNA de Plantas/metabolismo , Ribonuclease III/genética , Ribonuclease III/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Análise de Sequência de RNA , Processamento Pós-Transcricional do RNA , Regulação da Expressão Gênica de PlantasRESUMO
The H3 methyltransferases ATXR5 and ATXR6 deposit H3.1K27me1 to heterochromatin to prevent genomic instability and transposon re-activation. Here, we report that atxr5 atxr6 mutants display robust resistance to Geminivirus. The viral resistance is correlated with activation of DNA repair pathways, but not with transposon re-activation or heterochromatin amplification. We identify RAD51 and RPA1A as partners of virus-encoded Rep protein. The two DNA repair proteins show increased binding to heterochromatic regions and defense-related genes in atxr5 atxr6 vs wild-type plants. Consequently, the proteins have reduced binding to viral DNA in the mutant, thus hampering viral amplification. Additionally, RAD51 recruitment to the host genome arise via BRCA1, HOP2, and CYCB1;1, and this recruitment is essential for viral resistance in atxr5 atxr6. Thus, Geminiviruses adapt to healthy plants by hijacking DNA repair pathways, whereas the unstable genome, triggered by reduced H3.1K27me1, could retain DNA repairing proteins to suppress viral amplification in atxr5 atxr6.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Geminiviridae , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Heterocromatina/metabolismo , Geminiviridae/genética , Histonas/metabolismo , Replicação do DNA , Reparo do DNA/genética , Metiltransferases/metabolismoRESUMO
As important components of small RNA (smRNA) pathways, Argonaute (AGO) proteins mediate the interaction of incorporated smRNAs with their targets. Arabidopsis contains 10 AGO proteins with specialized or redundant functions. Among them, AGO1 mainly acts in microRNA (miRNA) and small-interfering RNA (siRNA) pathways for post-transcriptional gene silencing (PTGS), whereas AGO4 regulates transcriptional gene silencing (TGS) via endogenous 24-nucleotide (nt) smRNAs. To fully characterize smRNAs associated with AGO1 and AGO4, we developed a two-step protocol to purify AGO/smRNA complexes from flowers, leaves, roots and seedlings with enhanced purity, and sequenced the smRNAs by Illumina technology. Besides recovering most previously annotated smRNAs, we also identified some additional miRNAs, phased smRNA clusters and small-interfering RNAs derived from the overlapping region of natural antisense transcript pairs (NAT) (nat-siRNAs). We also identified a smRNA distribution feature on miRNA precursors which may help to identify authentic miRNAs. Organ-specific sequencing provided digital expression profiles of all obtained smRNAs, especially miRNAs. The presence and conservation of collateral miRNAs on known miRNA precursors were also investigated. Intriguingly, about 30% of AGO1-associated smRNAs were 24-nt long and unrelated to the 21-nt species. Further analysis showed that DNA-dependent RNA polymerase IV (Pol IV)-dependent smRNAs were mainly 24 nt and associated with AGO4, whereas the majority of the potential Pol V-dependent ones were 21-nt smRNAs and bound to AGO1, suggesting the potential involvement of AGO1 in Pol V-related pathways.