In Vitro Assembly of Plant RNA-Induced Silencing Complexes Facilitated by Molecular Chaperone HSP90

Iki, Taichiro; Yoshikawa, Manabu; Nishikiori, Masaki; Jaudal, Mauren; Matsumoto-Yokoyama, Eiko; Mitsuhara, Ichiro; Meshi, Tetsuo; Ishikawa, Masayuki

doi:10.1016/j.molcel.2010.05.014

Cited by 296 publications

(319 citation statements)

References 50 publications

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“…Efficient RISC assembly requires the incorporation of small RNAs in both animal and plant cells (34)(35)(36). Thus, we selected mutations that are known to affect miRNA biogenesis and accumulation, including mutations in the doublestranded RNA-binding protein DRB1 (also known as HYL1) and Dicer homolog DCL1 mediating processing of most miRNA precursors (37,38), the RNA methyltransferase HEN1 (39) critical for miRNA stability and HASTY (HST), the Arabidopsis ortholog of Exp5 required for the nuclear export and/or stability of miRNAs (40).…”

Section: Resultsmentioning

confidence: 99%

Degradation of the antiviral component ARGONAUTE1 by the autophagy pathway

Derrien

Baumberger

Schepetilnikov

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

256

288

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Posttranscriptional gene silencing (PTGS) mediated by siRNAs is an evolutionarily conserved antiviral defense mechanism in higher plants and invertebrates. In this mechanism, viral-derived siRNAs are incorporated into the RNA-induced silencing complex (RISC) to guide degradation of the corresponding viral RNAs. In Arabidopsis, a key component of RISC is ARGONAUTE1 (AGO1), which not only binds to siRNAs but also carries the RNA slicer activity. At present little is known about posttranslational mechanisms regulating AGO1 turnover. Here we report that the viral suppressor of RNA silencing protein P0 triggers AGO1 degradation by the autophagy pathway. Using a P0-inducible transgenic line, we observed that AGO1 degradation is blocked by inhibition of autophagy. The engineering of a functional AGO1 fluorescent reporter protein further indicated that AGO1 colocalizes with autophagy-related (ATG) protein 8a (ATG8a) positive bodies when degradation is impaired. Moreover, this pathway also degrades AGO1 in a nonviral context, especially when the production of miRNAs is impaired. Our results demonstrate that a selective process such as ubiquitylation can lead to the degradation of a key regulatory protein such as AGO1 by a degradation process generally believed to be unspecific. We anticipate that this mechanism will not only lead to degradation of AGO1 but also of its associated proteins and eventually small RNAs.

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Section: Resultsmentioning

confidence: 99%

Degradation of the antiviral component ARGONAUTE1 by the autophagy pathway

Derrien

Baumberger

Schepetilnikov

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

256

288

View full text Add to dashboard Cite

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“…HSP90 plays important roles in plant defense, especially in defense against microbial pathogens (Hubert et al, 2003;Takahashi et al, 2003;Sangster and Queitsch, 2005;Zhang et al, 2015). HSP90 also is implicated in chaperoning AGO1 (Iki et al, 2010). Our double mutant and expression analyses, however, suggest a synergistic, rather than a simple epistatic or additive, relationship.…”

Section: Discussionmentioning

confidence: 62%

The Mechanistic Underpinnings of anago1-Mediated, Environmentally Dependent, and Stochastic Phenotype

2016

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98195-5065 (G.A.M., T.L., C.Q.)The crucial role of microRNAs in plant development is exceedingly well supported; their importance in environmental robustness is studied in less detail. Here, we describe a novel, environmentally dependent phenotype in hypomorphic argonaute1 (ago1) mutants and uncover its mechanistic underpinnings in Arabidopsis (Arabidopsis thaliana). AGO1 is a key player in microRNA-mediated gene regulation. We observed transparent lesions on embryonic leaves of ago1 mutant seedlings. These lesions increased in frequency in full-spectrum light. Notably, the lesion phenotype was most environmentally responsive in ago1-27 mutants. This allele is thought to primarily affect translational repression, which has been linked with the response to environmental perturbation. Using several lines of evidence, we found that these lesions represent dead and dying tissues due to an aberrant hypersensitive response. Although all three canonical defense hormone pathways (salicylic acid, jasmonate, and jasmonate/ethylene pathways) were up-regulated in ago1 mutants, we demonstrate that jasmonate perception drives the lesion phenotype. Double mutants of ago1 and coronatine insensitive1, the jasmonate receptor, showed greatly decreased frequency of affected seedlings. The chaperone HEAT SHOCK PROTEIN 90 (HSP90), which maintains phenotypic robustness in the face of environmental perturbations, is known to facilitate AGO1 function. HSP90 perturbation has been shown previously to upregulate jasmonate signaling and to increase plant resistance to herbivory. Although single HSP90 mutants showed subtly elevated levels of lesions, double mutant analysis disagreed with a simple epistatic model for HSP90 and AGO1 interaction; rather, both appeared to act nonadditively in producing lesions. In summary, our study identifies AGO1 as a major, largely HSP90-independent, factor in providing environmental robustness to plants. ARGONAUTE1 (AGO1) is an ancient, highly conserved essential protein. One of 10 functionally distinct AGO proteins in the plant Arabidopsis (Arabidopsis thaliana), AGO1 uses microRNAs (miRNAs) to decrease the expression of target genes. AGO1 and the miRNA pathway are crucial for many regulatory processes throughout development and in response to environmental stimuli. As frequent components of feedforward loops, miRNAs also play important roles in controlling gene expression noise and maintaining the phenotypic robustness of specific traits in several organisms, including plants ( Lempe et al., 2013).AGO1 and miRNAs also are critical for proper plant responses to abiotic stress. The expression levels of miRNAs respond to heat shock, cold, drought, oxidative stress, UV light exposure, and hypoxia (Sunkar et al., 2012). For example, in response to drought, the expression of miR169 decreases and the expression of its target gene, encoding the transcription factor NFYA5, increases in Arabidopsis (Li et al., 2008). Increased expression of NFYA5 supports plant survival in drought conditions. Transgenic plants ove...

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“…It has been proposed that the chaperone machinery supports a loading-competent state of fly Ago2, allowing it to accept siRNA duplexes from Dcr-2-R2D2 (Miyoshi et al, 2010;Iwasaki et al, 2015). In plants, studies using tobacco BY-2 lysate demonstrated that HSP90 binds to Arabidopsis AGO1 and AGO4 and promotes the loading of sRNA duplexes (Iki et al, 2010;Ye et al, 2012). The cochaperones of HSP90 are capable of modulating the ATP-hydrolyzing rate of HSP90, recruiting client proteins, or playing chaperone roles on their own (Röhl et al, 2013).…”

Section: Loading Of Small Rnas Into Ago Family Proteinsmentioning

confidence: 99%

RNAi in Plants: An Argonaute-Centered View

2016

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Argonaute (AGO) family proteins are effectors of RNAi in eukaryotes. AGOs bind small RNAs and use them as guides to silence target genes or transposable elements at the transcriptional or posttranscriptional level. Eukaryotic AGO proteins share common structural and biochemical properties and function through conserved core mechanisms in RNAi pathways, yet plant AGOs have evolved specialized and diversified functions. This Review covers the general features of AGO proteins and highlights recent progress toward our understanding of the mechanisms and functions of plant AGOs.

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In Vitro Assembly of Plant RNA-Induced Silencing Complexes Facilitated by Molecular Chaperone HSP90

Cited by 296 publications

References 50 publications

Degradation of the antiviral component ARGONAUTE1 by the autophagy pathway

Degradation of the antiviral component ARGONAUTE1 by the autophagy pathway

The Mechanistic Underpinnings of anago1-Mediated, Environmentally Dependent, and Stochastic Phenotype

RNAi in Plants: An Argonaute-Centered View

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