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1.
New Phytol ; 243(6): 2351-2367, 2024 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-39030826

RESUMEN

Viroids are pathogenic noncoding RNAs that completely rely on their host molecular machinery to accomplish their life cycle. Several interactions between viroids and their host molecular machinery have been identified, including interference with epigenetic mechanisms such as DNA methylation. Despite this, whether viroids influence changes in other epigenetic marks such as histone modifications remained unknown. Epigenetic regulation is particularly important during pathogenesis processes because it might be a key regulator of the dynamism of the defense response. Here we have analyzed the changes taking place in Cucumis sativus (cucumber) facultative and constitutive heterochromatin during hop stunt viroid (HSVd) infection using chromatin immunoprecipitation (ChIP) of the two main heterochromatic marks: H3K9me2 and H3K27me3. We find that HSVd infection is associated with changes in both H3K27me3 and H3K9me2, with a tendency to decrease the levels of repressive epigenetic marks through infection progression. These epigenetic changes are connected to the transcriptional regulation of their expected targets, genes, and transposable elements. Indeed, several genes related to the defense response are targets of both epigenetic marks. Our results highlight another host regulatory mechanism affected by viroid infection, providing further information about the complexity of the multiple layers of interactions between pathogens/viroids and hosts/plants.


Asunto(s)
Epigénesis Genética , Regulación de la Expresión Génica de las Plantas , Heterocromatina , Histonas , Enfermedades de las Plantas , Viroides , Heterocromatina/metabolismo , Heterocromatina/genética , Viroides/genética , Viroides/fisiología , Viroides/patogenicidad , Histonas/metabolismo , Enfermedades de las Plantas/virología , Enfermedades de las Plantas/genética , Cucumis sativus/virología , Cucumis sativus/genética , Virus de Plantas/fisiología , Virus de Plantas/patogenicidad , Elementos Transponibles de ADN/genética , Interacciones Huésped-Patógeno/genética
2.
Mol Plant Pathol ; 25(7): e13469, 2024 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-38956901

RESUMEN

Viroids, one of the smallest known infectious agents, induce symptoms of varying severity, ranging from latent to severe, based on the combination of viroid isolates and host plant species. Because viroids are transmissible between plant species, asymptomatic viroid-infected plants may serve as latent sources of infection for other species that could exhibit severe symptoms, occasionally leading to agricultural and economic losses. Therefore, predicting the symptoms induced by viroids in host plants without biological experiments could remarkably enhance control measures against viroid damage. Here, we developed an algorithm using unsupervised machine learning to predict the severity of disease symptoms caused by viroids (e.g., potato spindle tuber viroid; PSTVd) in host plants (e.g., tomato). This algorithm, mimicking the RNA silencing mechanism thought to be linked to viroid pathogenicity, requires only the genome sequences of the viroids and host plants. It involves three steps: alignment of synthetic short sequences of the viroids to the host plant genome, calculation of the alignment coverage, and clustering of the viroids based on coverage using UMAP and DBSCAN. Validation through inoculation experiments confirmed the effectiveness of the algorithm in predicting the severity of disease symptoms induced by viroids. As the algorithm only requires the genome sequence data, it may be applied to any viroid and plant combination. These findings underscore a correlation between viroid pathogenicity and the genome sequences of viroid isolates and host plants, potentially aiding in the prevention of viroid outbreaks and the breeding of viroid-resistant crops.


Asunto(s)
Genoma Viral , Enfermedades de las Plantas , Solanum lycopersicum , Viroides , Solanum lycopersicum/virología , Enfermedades de las Plantas/virología , Viroides/genética , Viroides/patogenicidad , Genoma Viral/genética , Algoritmos , Genoma de Planta
3.
Plant Dis ; 108(7): 2181-2189, 2024 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-38522091

RESUMEN

Peach latent mosaic viroid (PLMVd) infects peach trees in China and induces a conspicuous albino phenotype (peach calico, PC) that is closely associated with variants containing a 12-to-14 nucleotide hairpin insertion capped by a U-rich loop. Initially, PC disease distribution was limited to parts of Italy, and it was first detected in the field in China in 2019. To explore the molecular and biological characteristics of PLMVd PC isolates in peach in China, we conducted a comprehensive analysis of disease phenotype development and investigated the data-associated pathogenicity and in vivo dynamics of the Chinese isolate PC-A2 using slash-inoculation into GF-305 peach seedlings. Inoculated seedlings displayed PC symptoms much earlier following topping treatment, and PLMVd infectivity was further assessed using bioassay and semiquantitative RT-PCR experiments. Evolutionary analysis showed that the PC isolate and its progeny variants clustered into a single phylogroup distinct from reference PC-C40 isolates from Italy and PC-K1 and PC-K2 from South Korea. Some PC-A2 progeny variants from green leaves of PC-expressing seedlings showed unbalanced point mutations in hairpin stems compared with the PC-C40 reference sequence and constituted a new stem insertion type. The results reveal associations between the recessive phenotypes of peach albino symptoms and base variation in hairpin stem insertions relative to the PC-C40/chloroplastic heat shock protein 90 reference sequence.


Asunto(s)
Enfermedades de las Plantas , Prunus persica , Viroides , Enfermedades de las Plantas/virología , Prunus persica/virología , China , Viroides/genética , Viroides/fisiología , Viroides/patogenicidad , Viroides/aislamiento & purificación , Filogenia , Mutación , Fenotipo , ARN Viral/genética , Plantones/virología , Hojas de la Planta/virología
4.
Viruses ; 15(3)2023 03 04.
Artículo en Inglés | MEDLINE | ID: mdl-36992390

RESUMEN

Hop latent viroid (HLVd) is the biggest concern for cannabis and hop growers worldwide. Although most HLVd-infected plants remain asymptomatic, research on hops has demonstrated a decrease in both the α-bitter acid and terpene content of hop cones, which affects their economic value. The HLVd-associated "dudding" or "duds" disease of cannabis was first reported in 2019 in California. Since then, the disease has become widespread in cannabis-growing facilities across North America. Although severe yield loss associated with duds disease has been recorded, little scientific information is available to growers in order to contain HLVd. Consequently, this review aims to summarise all of the scientific information available on HLVd so as to be able to understand the effect of HLVd on yield loss, cannabinoid content, terpene profile, disease management and inform crop protection strategies.


Asunto(s)
Cannabis , Carlavirus , Enfermedades de las Plantas , Viroides , Cannabis/química , Cannabis/virología , Humulus/química , Humulus/virología , América del Norte , Enfermedades de las Plantas/prevención & control , Enfermedades de las Plantas/virología , Viroides/patogenicidad , Viroides/fisiología , Carlavirus/patogenicidad , Carlavirus/fisiología , Especificidad del Huésped
5.
Mol Biol Rep ; 49(2): 1581-1586, 2022 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-34773552

RESUMEN

BACKGROUND: A 328-nucleotide variant of citrus bent leaf viroid (CBLVd) was characterized by citrus varieties in Malaysia. After the first report in Malaysia, the emerging CBLVd was detected in five citrus species, namely Citrofortunella microcarpa, Citrus aurantifolia, Citrus hystrix, Citrus maxima, and Citrus sinensis. METHODS AND RESULTS: CBLVd was detected in 23 out of 133 symptomatic samples through RT-PCR. Sequence analysis of the RT-PCR amplicons from this study showed 99-100% sequence identity to the reference CBLVd Jp isolate and CBLVd isolates reported in Malaysia. Inoculation of sap, obtained from a CBLVd positive sample, into 6-month old healthy C. microcarpa seedlings showed symptoms of slight leaf bending, reduced leaf size of matured leaves, and mild mosaic between 4 to 6 months after inoculation. Moreover, the observed symptoms of chlorosis, midvein necrosis, leaf rolling, and smalling of leaves in calamondin, C. microcarpa (Bunge) Wijnands, were not reported in earlier studies and opened a new avenue for the study of symptomology. The mechanical transmissibility of CBLVd in the inoculated seedlings was reconfirmed by RT-PCR assay and sequencing. CONCLUSIONS: Based on the results, the sequence similarity of CBLVd isolates from different areas of Malaysia showed no significant difference among each other and the reference isolate. The CBLVd is mechanically transmissible and could produce variable symptoms in different hosts.


Asunto(s)
Viroides/genética , Viroides/aislamiento & purificación , Viroides/patogenicidad , Secuencia de Bases/genética , Citrus/genética , Citrus/virología , Malasia/epidemiología , Conformación de Ácido Nucleico , Enfermedades de las Plantas/virología , Hojas de la Planta/genética , ARN Viral/genética , Programas Informáticos
6.
Cells ; 10(11)2021 11 01.
Artículo en Inglés | MEDLINE | ID: mdl-34831194

RESUMEN

Infectious dimeric RNA transcripts are a powerful tool for reverse genetic analyses in viroid studies. However, the construction of dimeric cDNA clones is laborious and time consuming, especially in mutational analyses by in vitro mutagenesis. In this study, we developed a system to synthesize a precisely monomeric linear RNA that could be transcribed in vitro directly from the cDNA clones of four viroid species. The cDNA clones were constructed such that RNA transcription was initiated at the guanine nucleotide of a predicted processing and ligation site in the viroid replication process. Although the transcribed RNAs were considered to possess 5'-triphosphate and 3'-hydroxyl termini, the RNA transcripts were infectious even without in vitro modifications. Additionally, infectivity was detected in the monomeric RNA transcripts, in which transcription was initiated at guanine nucleotides distinct from the predicted processing/ligation site. Moreover, monomeric viroid RNAs bearing 5'-monophosphate, 5'-hydroxyl, or 5'-capped termini were found to be infectious. Northern blot analysis of the pooled total RNA of the plants inoculated with the 5'-terminal modified RNA of potato spindle tuber viroid (PSTVd) indicated that maximum PSTVd accumulation occurred in plants with 5'-monophosphate RNA inoculation, followed by the plants with 5'-triphosphate RNA inoculation. Our system for synthesizing an infectious monomeric linear viroid RNA from a cDNA clone will facilitate mutational analyses by in vitro mutagenesis in viroid research.


Asunto(s)
ARN Viral/genética , Sitio de Iniciación de la Transcripción , Viroides/genética , Viroides/patogenicidad , Secuencia de Bases , Cucumis sativus/virología , ADN Complementario/genética , Solanum lycopersicum/virología , Enfermedades de las Plantas/virología , Procesamiento Postranscripcional del ARN/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Viral/aislamiento & purificación , Factores de Tiempo
7.
Arch Virol ; 166(11): 3157-3163, 2021 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-34396472

RESUMEN

The genus Coleviroid, family Pospiviroidae, comprises six known viroids, all infecting Plectranthus scutellarioides (Coleus blumei; coleus). In 2017, a novel viroid-like RNA sequence that shares ca. 65% identity with Coleus blumei viroid 1 (CbVd-1) was identified in a coleus cultivar infected by multiple coleviroids. Further sequence and secondary structure analyses are consistent with the discovery of a seventh viroid in the genus Coleviroid: tentatively named "Coleus blumei viroid 7" (CbVd-7). The viroid appears to be the product of a natural recombination event between CbVd-1 and Coleus blumei viroid 5. We prove CbVd-7 to be infectious and in turn demonstrate the ability of all known coleviroid left- and right-arm segments to recombine. With a length of 234 nucleotides, this is the smallest viroid described to date.


Asunto(s)
Plectranthus/virología , Virus Reordenados/genética , Recombinación Genética , Viroides/genética , Genoma Viral , Filogenia , Viroides/aislamiento & purificación , Viroides/patogenicidad
8.
Int J Mol Sci ; 22(12)2021 Jun 08.
Artículo en Inglés | MEDLINE | ID: mdl-34201240

RESUMEN

Infectious viroid clones consist of dimeric cDNAs used to generate transcripts which mimic the longer-than-unit replication intermediates. These transcripts can be either generated in vitro or produced in vivo by agro-inoculation. We have designed a new plasmid, which allows both inoculation methods, and we have compared them by infecting Solanum lycopersicum and Solanum melongena with clones of Citrus exocortis virod (CEVd), Tomato chlorotic dwarf viroid (TCDVd), and Potato spindle tuber viroid (PSTVd). Our results showed more uniform and severe symptoms in agro-inoculated plants. Viroid accumulation and the proportion of circular and linear forms were different depending on the host and the inoculation method and did not correlate with the symptoms, which correlated with an increase in PR1 induction, accumulation of the defensive signal molecules salicylic (SA) and gentisic (GA) acids, and ribosomal stress in tomato plants. The alteration in ribosome biogenesis was evidenced by both the upregulation of the tomato ribosomal stress marker SlNAC082 and the impairment in 18S rRNA processing, pointing out ribosomal stress as a novel signature of the pathogenesis of nuclear-replicating viroids. In conclusion, this updated binary vector has turned out to be an efficient and reproducible method that will facilitate the studies of viroid-host interactions.


Asunto(s)
Enfermedades de las Plantas/virología , Plásmidos/administración & dosificación , ARN Viral/genética , Ribosomas/metabolismo , Solanum lycopersicum/virología , Viroides/clasificación , Viroides/aislamiento & purificación , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismo , Enfermedades de las Plantas/genética , Plásmidos/genética , Ribosomas/genética , Viroides/patogenicidad
9.
Int J Mol Sci ; 22(6)2021 Mar 10.
Artículo en Inglés | MEDLINE | ID: mdl-33801996

RESUMEN

Viroids are tiny single-stranded circular RNA pathogens that infect plants. Viroids do not encode any proteins, yet cause an assortment of symptoms. The following review describes viroid classification, molecular biology and spread. The review also discusses viroid pathogenesis, host interactions and detection. The review concludes with a description of future prospects in viroid research.


Asunto(s)
ARN Circular/genética , ARN Viral/genética , Viroides/genética , Replicación Viral/genética , Resistencia a la Enfermedad/genética , Interacciones Huésped-Patógeno , Modelos Genéticos , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/virología , Plantas/genética , Plantas/virología , Viroides/clasificación , Viroides/patogenicidad , Virulencia/genética
10.
Plant Dis ; 105(10): 2785-2791, 2021 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-33560883

RESUMEN

Viruses and viroids prevalent in a population of 42 wild grapevines (i.e., free-living, uncultivated grapevines; Vitis spp.) were compared with those in a population of 85 cultivated grapevines collected in Tennessee, United States by RNA sequencing analysis of pools of ribosomal RNA-depleted total RNA. The sequences of 10 viruses (grapevine fleck virus, grapevine leafroll-associated virus 2, grapevine rupestris stem pitting-associated virus, grapevine Syrah virus 1, grapevine vein-clearing virus, grapevine virus B, grapevine virus E, tobacco ringspot virus, tomato ringspot virus, and a novel nano-like virus) and two viroids (hop stunt viroid and grapevine yellow speckle viroid 1) were detected in both grapevine populations. Sequences of four viruses (grapevine associated tymo-like virus, grapevine leafroll-associated virus 3, grapevine red blotch virus, and grapevine virus H) were identified only from cultivated grapevines. High, moderate, and low numbers of sequence reads were identified only from wild grapevines for a novel caulimovirus, an enamovirus, and alfalfa mosaic virus, respectively. The presence of most virus sequences and both viroids was verified independently in the original samples by reverse-transcription PCR followed by Sanger sequencing. Comparison of viral sequences shared by both populations showed that cultivated and wild grapevines harbored distinct sequence variants, which suggests that there was limited virus movement between the two populations. Collectively, this study represents the first unbiased survey of viruses and viroids in both cultivated and wild grapevines within a defined geographic region.


Asunto(s)
Enfermedades de las Plantas/virología , Viroides , Vitis , ARN Viral/genética , Tennessee , Viroides/genética , Viroides/patogenicidad , Vitis/virología
11.
Curr Opin Virol ; 47: 32-37, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33460914

RESUMEN

Viroids are single-stranded circular noncoding RNAs that infect plants. The noncoding nature indicates that viroids must harness their RNA genomes to redirect host machinery for infection. Therefore, the viroid model provides invaluable opportunities for delineating fundamental principles of RNA structure-function relationships and for dissecting the composition and mechanism of RNA-related cellular machinery. There are two viroid families, Pospiviroidae and Avsunviroidae. Members of both families replicate via the RNA-based rolling-circle mechanism with some variations. Viroid replication is generally divided into three steps: transcription, cleavage, and ligation. Decades of studies have uncovered numerous viroid RNA structures with a regulatory role in replication and multiple enzymes critical for the three replication steps. This review discusses these findings and highlights the latest discoveries. Future studies will continue to elucidate regulatory factors and mechanism of host machinery exploited by viroids and provide new insights into host-viroid interactions in the context of pathogenesis.


Asunto(s)
Viroides/fisiología , Replicación Viral/fisiología , ARN Polimerasas Dirigidas por ADN/metabolismo , Interacciones Huésped-Patógeno , Motivos de Nucleótidos , Enfermedades de las Plantas/virología , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , ARN Viral/química , ARN Viral/genética , ARN Viral/metabolismo , Transcripción Genética , Proteínas Virales/genética , Proteínas Virales/metabolismo , Viroides/clasificación , Viroides/patogenicidad
12.
Mol Plant Pathol ; 22(2): 153-162, 2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33305492

RESUMEN

TAXONOMY: Hop stunt viroid (HSVd) is the type species of the genus Hostuviroid (family Pospiviroidae). The other species of this genus is Dahlia latent viroid, which presents an identical central conserved region (CCR) but lacks other structural hallmarks present in Hop stunt viroid. HSVd replication occurs in the nucleus through an asymmetric rolling-circle model as in the other members of the family Pospiviroidae, which also includes the genera Pospiviroid, Cocadviroid, Apscaviroid, and Coleoviroid. PHYSICAL PROPERTIES: Hop stunt viroid consists of a single-stranded, circular RNA of 295-303 nucleotides depending on isolates and sequence variants. The most stable secondary structure is a rod-like or quasi-rod-like conformation with two characteristic domains: a CCR and a terminal conserved hairpin similar to that of cocadviroids. HSVd lacks a terminal conserved region. HOSTS AND SYMPTOMS: HSVd infects a very broad range of natural hosts and has been reported to be the causal agent of five different diseases (citrus cachexia, cucumber pale fruit, peach and plum apple apricot distortion, and hop stunt). It is distributed worldwide. TRANSMISSION: HSVd is transmitted mechanically and by seed.


Asunto(s)
Virus de Plantas/patogenicidad , ARN Viral/fisiología , Viroides/patogenicidad , Epigénesis Genética , Variación Genética , Genoma Viral , Interacciones Huésped-Patógeno , Enfermedades de las Plantas/virología , Virus de Plantas/genética , Viroides/genética , Replicación Viral
13.
Int J Mol Sci ; 21(19)2020 Oct 05.
Artículo en Inglés | MEDLINE | ID: mdl-33027943

RESUMEN

While the potato spindle tuber viroid (PSTVd) variant, PSTVd-Dahlia (PSTVd-D or PSTVd-Dwt) induces very mild symptoms in tomato cultivar 'Rutgers', PSTVd-Intermediate (PSTVd-I or PSTVd-Iwt) induces severe symptoms. These two variants differ by nine nucleotides, of which six mutations are located in the terminal left (TL) to the pathogenicity (P) domains. To evaluate the importance of mutations located in the TL to the P domains, ten types of point mutants were created by swapping the nucleotides between the two viroid variants. Bioassay in tomato plants demonstrated that two mutants created on PSTVd-Iwt at positions 42 and 64 resulted in symptom attenuation. Phenotypic and RT-qPCR analysis revealed that mutation at position 42 of PSTVd-Iwt significantly reduced disease severity and accumulation of the viroid, whereas mutation at position 64 showed a significant reduction in stunting when compared to the PSTVd-Iwt infected plant. RT-qPCR analysis on pathogenesis-related protein 1b1 and chalcone synthase genes showed a direct correlation with symptom severity whereas the expansin genes were down-regulated irrespective of the symptom severity. These results indicate that the nucleotides at positions 42 and 64 are in concert with the ones at positions 43, 310, and 311/312, which determines the slower and stable accumulation of PSTVd-D without eliciting excessive host defense responses thus contributing in the attenuation of disease symptom.


Asunto(s)
Dahlia/química , Enfermedades de las Plantas/genética , Solanum lycopersicum/genética , Viroides/efectos de los fármacos , Solanum lycopersicum/crecimiento & desarrollo , Solanum lycopersicum/virología , Nucleótidos/genética , Enfermedades de las Plantas/virología , Extractos Vegetales/química , Extractos Vegetales/farmacología , Virus de Plantas/efectos de los fármacos , Virus de Plantas/patogenicidad , Virus ARN/genética , Virus ARN/patogenicidad , ARN Viral/genética , Viroides/patogenicidad
14.
Int J Mol Sci ; 21(20)2020 Oct 13.
Artículo en Inglés | MEDLINE | ID: mdl-33066322

RESUMEN

Tomato (Lycopersicum esculentum L.) and pepper (Capsicum annuum L.) plants belonging to the family Solanaceae are cultivated worldwide. The rapid development of next-generation sequencing (NGS) technology facilitates the identification of viruses and viroids infecting plants. In this study, we carried out metatranscriptomics using RNA sequencing followed by bioinformatics analyses to identify viruses and viroids infecting tomato and pepper plants in Vietnam. We prepared a total of 16 libraries, including eight tomato and eight pepper libraries derived from different geographical regions in Vietnam. We identified a total of 602 virus-associated contigs, which were assigned to 18 different virus species belonging to nine different viral genera. We identified 13 different viruses and two viroids infecting tomato plants and 12 viruses and two viroids infecting pepper plants with viruses as dominantly observed pathogens. Our results showed that multiple infection of different viral pathogens was common in both plants. Moreover, geographical region and host plant were two major factors to determine viral populations. Taken together, our results provide the comprehensive overview of viral pathogens infecting two important plants in the family Solanaceae grown in Vietnam.


Asunto(s)
Capsicum/virología , Metagenómica/métodos , Tipificación Molecular/métodos , Virus de Plantas/genética , Solanum lycopersicum/virología , Transcriptoma , Viroides/genética , Virus de Plantas/patogenicidad , Vietnam , Viroides/patogenicidad
15.
PLoS One ; 15(7): e0236481, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32716919

RESUMEN

RNA-dependent RNA polymerase 6 (RDR6) is one of the key factors in plant defense responses and suppresses virus or viroid invasion into shoot apical meristem (SAM) in Nicotiana benthamiana. To evaluate the role of Solanum lycopersicum (Sl) RDR6 upon viroid infection, SlRDR6-suppressed (SlRDR6i) 'Moneymaker' tomatoes were generated by RNA interference and inoculated with intermediate or lethal strain of potato spindle tuber viroid (PSTVd). Suppression of SlRDR6 did not change disease symptoms of both PSTVd strains in 'Moneymaker' tomatoes. Analysis of PSTVd distribution in shoot apices by in situ hybridization revealed that both PSTVd strains similarly invade the basal part but not apical part including pluripotent stem cells of SAM in SlRDR6i plants at a low rate unlike a previous report in N. benthamiana. In addition, unexpectedly, amount of PSTVd accumulation was apparently lower in SlRDR6i plants than in control tomatoes transformed with empty cassette in early infection especially in the lethal strain. Meanwhile, SlRDR6 suppression did not affect the seed transmission rates of PSTVd. These results indicate that RDR6 generally suppresses PSTVd invasion into SAM in plants, while suppression of RDR6 does not necessarily elevate amount of PSTVd accumulation. Additionally, our results suggest that host factors such as RDR1 other than RDR6 may also be involved in the protection of SAM including pluripotent stem cells from PSTVd invasion and effective RNA silencing causing the decrease of PSTVd accumulation during early infection in tomato plants.


Asunto(s)
Meristema/citología , Meristema/virología , Proteínas de Plantas/metabolismo , Células Madre Pluripotentes/virología , ARN Polimerasa Dependiente del ARN/metabolismo , Solanum lycopersicum/enzimología , Solanum lycopersicum/virología , Viroides/patogenicidad , Regulación de la Expresión Génica de las Plantas , Genoma Viral , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores de Tiempo , Viroides/aislamiento & purificación
16.
Virus Res ; 286: 198090, 2020 09.
Artículo en Inglés | MEDLINE | ID: mdl-32634444

RESUMEN

Viroids with small, non-coding circular RNA genome can induce diseases in many plant species. The extend of infection symptoms depends on environmental conditions, viroid strain, and host plant species and cultivar. Pathogen recognition leads to massive transcriptional reprogramming to favor defense responses over normal cellular functions. To better understand the interaction between plant host and potato spindle tuber viroid (PSTVd) variants that differ in their virulence, comparative transcriptomic analysis was performed by an RNA-seq approach. The changes of gene expression were analyzed at the time point when subtle symptoms became visible in plants infected with the severe PSTVd-S23 variant, while those infected with the mild PSTVd-M variant looked like non-infected healthy plants. Over 3000 differentially expressed genes (DEGs) were recognized in both infections, but the majority of them were specific for infection with the severe variant. In both infections recognized DEGs were mainly related to biotic stress, hormone metabolism and signaling, transcription regulation, protein degradation, and transport. The DEGs related to cell cycle and microtubule were uniquely down-regulated only in the PSTVd-S23-infected plants. Similarly, expression of transcription factors from C2C2-GATA and growth-regulating factor (GRF) families was only altered upon infection with the severe variant. Both PSTVd variants triggered plant immune response; however expression of genes encoding crucial factors of this process was markedly more changed in the plants infected with the severe variant than in those with the mild one.


Asunto(s)
Ciclo Celular/genética , Enfermedades de las Plantas/genética , Solanum lycopersicum/virología , Viroides/genética , Regulación de la Expresión Génica de las Plantas , Enfermedades de las Plantas/virología , Proteínas de Plantas/genética , Virus ARN/genética , ARN Viral/genética , RNA-Seq , Transcriptoma , Viroides/patogenicidad
17.
Int J Mol Sci ; 21(7)2020 Apr 03.
Artículo en Inglés | MEDLINE | ID: mdl-32260277

RESUMEN

The mediator (MED) represents a large, conserved, multi-subunit protein complex that regulates gene expression through interactions with RNA polymerase II and enhancer-bound transcription factors. Expanding research accomplishments suggest the predominant role of plant MED subunits in the regulation of various physiological and developmental processes, including the biotic stress response against bacterial and fungal pathogens. However, the involvement of MED subunits in virus/viroid pathogenesis remains elusive. In this study, we investigated for the first time the gene expression modulation of selected MED subunits in response to five viroid species (Apple fruit crinkle viroid (AFCVd), Citrus bark cracking viroid (CBCVd), Hop latent viroid (HLVd), Hop stunt viroid (HSVd), and Potato spindle tuber viroid (PSTVd)) in two model plant species (Nicotiana tabacum and N. benthamiana) and a commercially important hop (Humulus lupulus) cultivar. Our results showed a differential expression pattern of MED subunits in response to a viroid infection. The individual plant MED subunits displayed a differential and tailored expression pattern in response to different viroid species, suggesting that the MED expression is viroid- and plant species-dependent. The explicit evidence obtained from our results warrants further investigation into the association of the MED subunit with symptom development. Together, we provide a comprehensive portrait of MED subunit expression in response to viroid infection and a plausible involvement of MED subunits in fine-tuning transcriptional reprogramming in response to viroid infection, suggesting them as a potential candidate for rewiring the defense response network in plants against pathogens.


Asunto(s)
Humulus/virología , Complejo Mediador/genética , Nicotiana/virología , Viroides/patogenicidad , Perfilación de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Humulus/genética , Hojas de la Planta/genética , Hojas de la Planta/microbiología , Proteínas de Plantas/genética , Virus de Plantas , Especificidad de la Especie , Nicotiana/genética , Viroides/genética
18.
Nucleic Acids Res ; 48(6): 3134-3155, 2020 04 06.
Artículo en Inglés | MEDLINE | ID: mdl-32083649

RESUMEN

While G/U pairs are present in many RNAs, the lack of molecular studies to characterize the roles of multiple G/U pairs within a single RNA limits our understanding of their biological significance. From known RNA 3D structures, we observed that the probability a G/U will form a Watson-Crick (WC) base pair depends on sequence context. We analyzed 17 G/U pairs in the 359-nucleotide genome of Potato spindle tuber viroid (PSTVd), a circular non-coding RNA that replicates and spreads systemically in host plants. Most putative G/U base pairs were experimentally supported by selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE). Deep sequencing PSTVd genomes from plants inoculated with a cloned master sequence revealed naturally occurring variants, and showed that G/U pairs are maintained to the same extent as canonical WC base pairs. Comprehensive mutational analysis demonstrated that nearly all G/U pairs are critical for replication and/or systemic spread. Two selected G/U pairs were found to be required for PSTVd entry into, but not for exit from, the host vascular system. This study identifies critical roles for G/U pairs in the survival of an infectious RNA, and increases understanding of structure-based regulation of replication and trafficking of pathogen and cellular RNAs.


Asunto(s)
Virus de Plantas/genética , ARN no Traducido/genética , ARN Viral/genética , Viroides/genética , Genoma Viral/genética , Mutación , Conformación de Ácido Nucleico , Virus de Plantas/patogenicidad , Solanum tuberosum/virología , Viroides/patogenicidad , Virosis/genética , Virosis/virología , Replicación Viral/genética
19.
PLoS Pathog ; 15(12): e1008110, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31790500

RESUMEN

Viroids are small, non-protein-coding RNAs which can induce disease symptoms in a variety of plant species. Potato (Solanum tuberosum L.) is the natural host of Potato spindle tuber viroid (PSTVd) where infection results in stunting, distortion of leaves and tubers and yield loss. Replication of PSTVd is accompanied by the accumulation of viroid-derived small RNAs (sRNAs) proposed to play a central role in disease symptom development. Here we report that PSTVd sRNAs direct RNA silencing in potato against StTCP23, a member of the TCP (teosinte branched1/Cycloidea/Proliferating cell factor) transcription factor family genes that play an important role in plant growth and development as well as hormonal regulation, especially in responses to gibberellic acid (GA). The StTCP23 transcript has 21-nucleotide sequence complementarity in its 3' untranslated region with the virulence-modulating region (VMR) of PSTVd strain RG1, and was downregulated in PSTVd-infected potato plants. Analysis using 3' RNA ligase-mediated rapid amplification of cDNA ends (3' RLM RACE) confirmed cleavage of StTCP23 transcript at the expected sites within the complementarity with VMR-derived sRNAs. Expression of these VMR sRNA sequences as artificial miRNAs (amiRNAs) in transgenic potato plants resulted in phenotypes reminiscent of PSTVd-RG1-infected plants. Furthermore, the severity of the phenotypes displayed was correlated with the level of amiRNA accumulation and the degree of amiRNA-directed down-regulation of StTCP23. In addition, virus-induced gene silencing (VIGS) of StTCP23 in potato also resulted in PSTVd-like phenotypes. Consistent with the function of TCP family genes, amiRNA lines in which StTCP23 expression was silenced showed a decrease in GA levels as well as alterations to the expression of GA biosynthesis and signaling genes previously implicated in tuber development. Application of GA to the amiRNA plants minimized the PSTVd-like phenotypes. Taken together, our results indicate that sRNAs derived from the VMR of PSTVd-RG1 direct silencing of StTCP23 expression, thereby disrupting the signaling pathways regulating GA metabolism and leading to plant stunting and formation of small and spindle-shaped tubers.


Asunto(s)
Genes de Plantas , Enfermedades de las Plantas/virología , Solanum tuberosum/virología , Viroides/patogenicidad , Virulencia/fisiología , Regulación de la Expresión Génica de las Plantas/fisiología , Interferencia de ARN/fisiología , Virus ARN , ARN Viral , Solanum tuberosum/genética , Factores de Transcripción
20.
Nucleic Acids Res ; 47(16): 8649-8661, 2019 09 19.
Artículo en Inglés | MEDLINE | ID: mdl-31392997

RESUMEN

Viroids are naked RNAs that do not code for any known protein and yet are able to infect plants causing severe diseases. Because of their RNA nature, many studies have focused on the involvement of viroids in RNA-mediated gene silencing as being their pathogenesis mechanism. Here, the alterations caused by the Citrus exocortis viroid (CEVd) on the tomato translation machinery were studied as a new aspect of viroid pathogenesis. The presence of viroids in the ribosomal fractions of infected tomato plants was detected. More precisely, CEVd and its derived viroid small RNAs were found to co-sediment with tomato ribosomes in vivo, and to provoke changes in the global polysome profiles, particularly in the 40S ribosomal subunit accumulation. Additionally, the viroid caused alterations in ribosome biogenesis in the infected tomato plants, affecting the 18S rRNA maturation process. A higher expression level of the ribosomal stress mediator NAC082 was also detected in the CEVd-infected tomato leaves. Both the alterations in the rRNA processing and the induction of NAC082 correlate with the degree of viroid symptomatology. Taken together, these results suggest that CEVd is responsible for defective ribosome biogenesis in tomato, thereby interfering with the translation machinery and, therefore, causing ribosomal stress.


Asunto(s)
Enfermedades de las Plantas/genética , Biosíntesis de Proteínas , ARN de Planta/genética , ARN Ribosómico 18S/genética , Ribosomas/metabolismo , Solanum lycopersicum/genética , Viroides/genética , Citrus/virología , Interacciones Huésped-Patógeno/genética , Solanum lycopersicum/metabolismo , Solanum lycopersicum/virología , Biogénesis de Organelos , Enfermedades de las Plantas/virología , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Hojas de la Planta/virología , Interferencia de ARN , ARN de Planta/antagonistas & inhibidores , ARN de Planta/metabolismo , ARN Ribosómico 18S/antagonistas & inhibidores , ARN Ribosómico 18S/metabolismo , ARN Viral/genética , ARN Viral/metabolismo , Proteínas Ribosómicas/genética , Proteínas Ribosómicas/metabolismo , Ribosomas/genética , Estrés Fisiológico/genética , Viroides/metabolismo , Viroides/patogenicidad
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