Citrus exocortis viroid causes ribosomal stress in tomato plants

Abstract 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.

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Ethylene is Involved in Symptom Development and Ribosomal Stress of Tomato Plants upon Citrus Exocortis Viroid Infection

Plants ◽

10.3390/plants9050582 ◽

2020 ◽

Vol 9 (5) ◽

pp. 582

Author(s):

Francisco Vázquez Prol ◽

M. Pilar López-Gresa ◽

Ismael Rodrigo ◽

José María Bellés ◽

Purificación Lisón

Keyword(s):

Ribosome Biogenesis ◽

Ethylene Biosynthesis ◽

Northern Blotting ◽

Phenotypic Characterization ◽

Disease Development ◽

Tomato Plants ◽

Susceptibility To Infection ◽

Citrus Exocortis Viroid ◽

Viroid Infection ◽

Ribosomal Stress

Citrus exocortis viroid (CEVd) is known to cause different symptoms in citrus trees, and its mechanism of infection has been studied in tomato as an experimental host, producing ribosomal stress on these plants. Some of the symptoms caused by CEVd in tomato plants resemble those produced by the phytohormone ethylene. The present study is focused on elucidating the relationship between CEVd infection and ethylene on disease development. To this purpose, the ethylene insensitive Never ripe (Nr) tomato mutants were infected with CEVd, and several aspects such as susceptibility to infection, defensive response, ethylene biosynthesis and ribosomal stress were studied. Phenotypic characterization revealed higher susceptibility to CEVd in these mutants, which correlated with higher expression levels of both defense and ethylene biosynthesis genes, as well as the ribosomal stress marker SlNAC082. In addition, Northern blotting revealed compromised ribosome biogenesis in all CEVd infected plants, particularly in Nr mutants. Our results indicate a higher ethylene biosynthesis in Nr mutants and suggest an important role of this phytohormone in disease development and ribosomal stress caused by viroid infection.

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A Novel Conserved RNA-binding Domain Protein, RBD-1, Is Essential For Ribosome Biogenesis

Molecular Biology of the Cell ◽

10.1091/mbc.e02-03-0138 ◽

2002 ◽

Vol 13 (10) ◽

pp. 3683-3695 ◽

Cited By ~ 6

Author(s):

Petra Björk ◽

Göran Baurén ◽

ShaoBo Jin ◽

Yong-Guang Tong ◽

Thomas R. Bürglin ◽

...

Keyword(s):

Ribosome Biogenesis ◽

Rna Binding ◽

Ribosomal Subunit ◽

Binding Domain ◽

Dependent Manner ◽

Binding Domains ◽

40S Ribosomal Subunit ◽

Rna Binding Domain

Synthesis of the ribosomal subunits from pre-rRNA requires a large number of trans-acting proteins and small nucleolar ribonucleoprotein particles to execute base modifications, RNA cleavages, and structural rearrangements. We have characterized a novel protein, RNA-binding domain-1 (RBD-1), that is involved in ribosome biogenesis. This protein contains six consensus RNA-binding domains and is conserved as to sequence, domain organization, and cellular location from yeast to human. RBD-1 is essential in Caenorhabditis elegans. In the dipteran Chironomus tentans, RBD-1 (Ct-RBD-1) binds pre-rRNA in vitro and anti-Ct-RBD-1 antibodies repress pre-rRNA processing in vivo. Ct-RBD-1 is mainly located in the nucleolus in an RNA polymerase I transcription-dependent manner, but it is also present in discrete foci in the interchromatin and in the cytoplasm. In cytoplasmic extracts, 20–30% of Ct-RBD-1 is associated with ribosomes and, preferentially, with the 40S ribosomal subunit. Our data suggest that RBD-1 plays a role in structurally coordinating pre-rRNA during ribosome biogenesis and that this function is conserved in all eukaryotes.

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Characterization of small RNAs derived from Citrus exocortis viroid (CEVd) in infected tomato plants

Virology ◽

10.1016/j.virol.2007.05.011 ◽

2007 ◽

Vol 367 (1) ◽

pp. 135-146 ◽

Cited By ~ 53

Author(s):

Raquel Martín ◽

Catalina Arenas ◽

José-Antonio Daròs ◽

Alejandra Covarrubias ◽

José Luis Reyes ◽

...

Keyword(s):

Small Rnas ◽

Tomato Plants ◽

Citrus Exocortis Viroid ◽

Infected Tomato

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Regulation of Ribosomal Protein OperonsrplM-rpsI,rpmB-rpmG, andrplU-rpmAat the Transcriptional and Translational Levels

Journal of Bacteriology ◽

10.1128/jb.00187-16 ◽

2016 ◽

Vol 198 (18) ◽

pp. 2494-2502 ◽

Cited By ~ 14

Author(s):

Leonid V. Aseev ◽

Ludmila S. Koledinskaya ◽

Irina V. Boni

Keyword(s):

Ribosomal Proteins ◽

Ribosome Biogenesis ◽

Ribosomal Subunit ◽

General Rule ◽

Single Copy ◽

Translation Initiation Region ◽

Content Type ◽

Regulatory Pathways ◽

E Coli

ABSTRACTIt is widely assumed that in the best-characterized model bacteriumEscherichia coli, transcription units encoding ribosomal proteins (r-proteins) and regulation of their expression have been already well defined. However, transcription start sites for severalE. colir-protein operons have been established only very recently, so that information concerning the regulation of these operons at the transcriptional or posttranscriptional level is still missing. This paper describes for the first time thein vivoregulation of three r-protein operons,rplM-rpsI,rpmB-rpmG, andrplU-rpmA. The results demonstrate that transcription of all three operons is subject to ppGpp/DksA-dependent negative stringent control under amino acid starvation, in parallel with the rRNA operons. By using single-copy translational fusions with the chromosomallacZgene, we show here that at the translation level only one of these operons,rplM-rpsI, is regulated by the mechanism of autogenous repression involving the 5′ untranslated region (UTR) of the operon mRNA, whilerpmB-rpmGandrplU-rpmAare not subject to this type of regulation. This may imply that translational feedback control is not a general rule for modulating the expression ofE. colir-protein operons. Finally, we report that L13, a primary protein in 50S ribosomal subunit assembly, serves as a repressor ofrplM-rpsIexpressionin vivo, acting at a target within therplMtranslation initiation region. Thus, L13 represents a novel example of regulatory r-proteins in bacteria.IMPORTANCEIt is important to obtain a deeper understanding of the regulatory mechanisms responsible for coordinated and balanced synthesis of ribosomal components. In this paper, we highlight the major role of a stringent response in regulating transcription of three previously unexplored r-protein operons, and we show that only one of them is subject to feedback regulation at the translational level. Improved knowledge of the regulatory pathways controlling ribosome biogenesis may promote the development of novel antibacterial agents.

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The final step of 40S ribosomal subunit maturation is controlled by a dual key lock

10.1101/2020.07.29.226936 ◽

2020 ◽

Author(s):

Laura Plassart ◽

Ramtin Shayan ◽

Christian Montellese ◽

Dana Rinaldi ◽

Natacha Larburu ◽

...

Keyword(s):

Ribosomal Subunit ◽

Ribosomal Subunits ◽

40S Ribosomal Subunit ◽

Inactive Form ◽

Final Maturation ◽

Translation Apparatus ◽

Translation Accuracy ◽

Functional Analyses

Preventing premature interaction of preribosomes with the translation apparatus is essential to translation accuracy. Hence, the final maturation step releasing functional 40S ribosomal subunits, namely processing of the 18S ribosomal RNA 3′ end, is safeguarded by protein DIM2, which both interacts with the endoribonuclease NOB1 and masks the rRNA cleavage site. To elucidate the control mechanism that unlocks NOB1 activity, we performed cryo-EM analysis of late human pre-40S particles purified using a catalytically-inactive form of ATPase RIO1. These structures, together with in vivo and in vitro functional analyses, support a model in which ATPloaded RIO1 cooperates with ribosomal protein RPS26/eS26 to displace DIM2 from the 18S rRNA 3′ end, thereby triggering final cleavage by NOB1; release of ADP then leads to RIO1 dissociation from the 40S subunit. This dual key lock mechanism requiring RIO1 and RPS26 guarantees the precise timing of pre-40S particle conversion into translation-competent ribosomal subunits.

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Chaperone-E3 Ligase Complex HSP70-CHIP Mediates Ubiquitination of Ribosomal Protein S3

International Journal of Molecular Sciences ◽

10.3390/ijms19092723 ◽

2018 ◽

Vol 19 (9) ◽

pp. 2723 ◽

Cited By ~ 2

Author(s):

Inwoo Hwang ◽

Sung-Woo Cho ◽

Jee-Yin Ahn

Keyword(s):

Ribosomal Protein ◽

Ribosome Biogenesis ◽

Ribosomal Subunit ◽

E3 Ligase ◽

Interacting Protein ◽

Akt Activation ◽

Protein Levels ◽

40S Ribosomal Subunit ◽

Subsequent Degradation ◽

Ribosomal Protein S3

In addition to its role in ribosome biogenesis, ribosomal protein S3 (RPS3), a component of the 40S ribosomal subunit, has been suggested to possess several extraribosomal functions, including an apoptotic function. In this study, we demonstrated that in the mouse brain, the protein levels of RPS3 were altered by the degree of nutritional starvation and correlated with neuronal apoptosis. After endurable short-term starvation, the apoptotic function of RPS3 was suppressed by Akt activation and Akt-mediated T70 phosphorylation, whereas after prolonged starvation, the protein levels of RPS3 notably increased, and abundant neuronal death occurred. These events coincided with ubiquitination and subsequent degradation of RPS3, controlled by HSP70 and the cochaperone E3 ligase: carboxy terminus of heat shock protein 70-interacting protein (CHIP). Thus, our study points to an extraribosomal role of RPS3 in balancing neuronal survival or death depending on the degree of starvation through CHIP-mediated polyubiquitination and degradation.

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Effects of starvation, diabetes and acute insulin treatment on the regulation of polypeptide-chain initiation in rat skeletal muscle

Biochemical Journal ◽

10.1042/bj2230687 ◽

1984 ◽

Vol 223 (3) ◽

pp. 687-696 ◽

Cited By ~ 63

Author(s):

C S Harmon ◽

C G Proud ◽

V M Pain

Keyword(s):

Skeletal Muscle ◽

Protein Synthesis ◽

Polypeptide Chain ◽

Ribosomal Subunit ◽

Diabetic Rats ◽

Initiation Factor ◽

Chain Initiation ◽

Low Concentrations ◽

40S Ribosomal Subunit

The rate of protein synthesis in skeletal muscle is greatly decreased in response to diabetes and starvation. Analysis of polyribosome profiles indicates that polypeptide-chain initiation is impaired under these conditions. To identify the step in initiation that is affected, we assayed the incorporation of [35S]methionyl-tRNAfMet into [35S]methionyl-tRNAfMet . 40S-ribosomal-subunit initiation complexes in cell-free extracts based on postmitochondrial supernatants prepared from gastrocnemius muscle. Extracts from either starved or diabetic rats were 30-40% less active in forming these complexes compared with those derived from fed or insulin-maintained controls respectively. This change could be reversed by treatment of either starved or diabetic rats with insulin in vivo 30 min before death. Formation of 40S initiation complexes by extracts from either fed or starved rats could be stimulated by the addition of exogenous purified initiation factor eIF-2, but extracts from starved or diabetic rats were more sensitive than controls to stimulation by low concentrations of the factor. These results provide evidence for the acute regulation by insulin of protein synthesis in skeletal muscle at the level of polypeptide-chain initiation, and suggest that in this tissue, as in certain other eukaryotic systems, control of initiation appears to be mediated by changes in the activity of initiation factor eIF-2.

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Xenopus LSm Proteins Bind U8 snoRNA via an Internal Evolutionarily Conserved Octamer Sequence

Molecular and Cellular Biology ◽

10.1128/mcb.22.12.4101-4112.2002 ◽

2002 ◽

Vol 22 (12) ◽

pp. 4101-4112 ◽

Cited By ~ 33

Author(s):

Nenad Tomasevic ◽

Brenda A. Peculis

Keyword(s):

Ribosome Biogenesis ◽

Rna Binding ◽

Ribosomal Subunit ◽

High Specificity ◽

Sm Proteins ◽

U6 Snrna ◽

Evolutionarily Conserved

ABSTRACT U8 snoRNA plays a unique role in ribosome biogenesis: it is the only snoRNA essential for maturation of the large ribosomal subunit RNAs, 5.8S and 28S. To learn the mechanisms behind the in vivo role of U8 snoRNA, we have purified to near homogeneity and characterized a set of proteins responsible for the formation of a specific U8 RNA-binding complex. This 75-kDa complex is stable in the absence of added RNA and binds U8 with high specificity, requiring the conserved octamer sequence present in all U8 homologues. At least two proteins in this complex can be cross-linked directly to U8 RNA. We have identified the proteins as Xenopus homologues of the LSm (like Sm) proteins, which were previously reported to be involved in cytoplasmic degradation of mRNA and nuclear stabilization of U6 snRNA. We have identified LSm2, -3, -4, -6, -7, and -8 in our purified complex and found that this complex associates with U8 RNA in vivo. This purified complex can bind U6 snRNA in vitro but does not bind U3 or U14 snoRNA in vitro, demonstrating that the LSm complex specifically recognizes U8 RNA.

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The Heat Shock Protein YbeY Is Required for Optimal Activity of the 30S Ribosomal Subunit

Journal of Bacteriology ◽

10.1128/jb.00448-10 ◽

2010 ◽

Vol 192 (18) ◽

pp. 4592-4596 ◽

Cited By ~ 22

Author(s):

Aviram Rasouly ◽

Chen Davidovich ◽

Eliora Z. Ron

Keyword(s):

Heat Shock ◽

Ribosomal Subunit ◽

Heat Shock Gene ◽

Wild Type ◽

Lower Efficiency ◽

Ribosomal Subunits ◽

Translation Machinery ◽

Shock Gene

ABSTRACT The highly conserved bacterial ybeY gene is a heat shock gene whose function is not fully understood. Previously, we showed that the YbeY protein is involved in protein synthesis, as Escherichia coli mutants with ybeY deleted exhibit severe translational defects in vivo. Here we show that the in vitro activity of the translation machinery of ybeY deletion mutants is significantly lower than that of the wild type. We also show that the lower efficiency of the translation machinery is due to impaired 30S small ribosomal subunits.

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yRACK1/Asc1 proxiOMICs—Towards Illuminating Ships Passing in the Night

Cells ◽

10.3390/cells8111384 ◽

2019 ◽

Vol 8 (11) ◽

pp. 1384 ◽

Cited By ~ 1

Author(s):

Kerstin Schmitt ◽

Oliver Valerius

Keyword(s):

Quality Control ◽

Binding Proteins ◽

Ribosomal Subunit ◽

Stress Factors ◽

Head Region ◽

Ubiquitin E3 Ligase ◽

Mrna Binding Proteins ◽

40S Ribosomal Subunit ◽

Mrna Binding

Diverse signals and stress factors regulate the activity and homeostasis of ribosomes in all cells. The Saccharomyces cerevisiae protein Asc1/yRACK1 occupies an exposed site at the head region of the 40S ribosomal subunit (hr40S) and represents a central hub for signaling pathways. Asc1 strongly affects protein phosphorylation and is involved in quality control pathways induced by translation elongation arrest. Therefore, it is important to understand the dynamics of protein formations in the Asc1 microenvironment at the hr40S. We made use of the in vivo protein-proximity labeling technique Biotin IDentification (BioID). Unbiased proxiOMICs from two adjacent perspectives identified nucleocytoplasmic shuttling mRNA-binding proteins, the deubiquitinase complex Ubp3-Bre5, as well as the ubiquitin E3 ligase Hel2 as neighbors of Asc1. We observed Asc1-dependency of hr40S localization of mRNA-binding proteins and the Ubp3 co-factor Bre5. Hel2 and Ubp3-Bre5 are described to balance the mono-ubiquitination of Rps3 (uS3) during ribosome quality control. Here, we show that the absence of Asc1 resulted in massive exposure and accessibility of the C-terminal tail of its ribosomal neighbor Rps3 (uS3). Asc1 and some of its direct neighbors together might form a ribosomal decision tree that is tightly connected to close-by signaling modules.

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