Olivier Voinnet

ETH Zurich | ETH Zürich · Department of Biology

Unlike in metazoans, plant microRNAs (miRNAs) undergo stepwise nuclear maturation before engaging cytosolic, sequence-complementary transcripts in association with the silencing effector protein ARGONAUTE1 (AGO1). Since their discovery, how and under which form plant miRNAs translocate to the cytosol has remained unclear, as has their sub-cellular...

Loaded into ARGONAUTE(AGO) proteins, eukaryotic micro(mi)RNAs regulate gene expression via cleavage, translational repression, and/or accelerated decay of sequence-complementary target transcripts. Despite their importance in development, cell identity maintenance and stress responses, how individual miRNAs contribute to spatial gene regulation wit...

In RNA interference (RNAi), the RNase III Dicer processes long double-stranded RNA (dsRNA) into short interfering RNA (siRNA), which, when loaded into ARGONAUTE (AGO) family proteins, execute gene silencing. Remarkably, RNAi can act non-cell autonomously: it is graft transmissible, and plasmodesmata-associated proteins modulate its cell-to-cell spr...

Co-evolution between hosts' and parasites' genomes shapes diverse pathways of acquired immunity based on silencing small (s)RNAs. In plants, sRNAs cause heterochromatinization, sequence-degeneration and, ultimately, loss-of-autonomy of most transposable elements (TEs). Recognition of newly-invasive plant TEs, by contrast, involves an innate antivir...

Regulation of intercellular small RNA (sRNA)-mediated gene silencing in plants is commonly ascribed to hypothetical mechanisms involving movement channels. In this commentary, I present a complementary, perhaps counter-intuitive view, in which sRNA movement is also crucially regulated by the cell-autonomous silencing machinery found in silencing-em...

In RNA interference (RNAi), small-interfering (si)RNAs processed from double-stranded RNA guide ARGONAUTE(AGO) proteins to silence sequence-complementary RNA/DNA. RNAi can propagate locally and systemically in plants, but despite recent mechanistic advances, basic questions remain unaddressed. For instance, RNAi is inferred to diffuse through plasm...

Transient transgenic expression accelerates pharming and facilitates protein studies in plants. One embodiment of the approach involves leaf infiltration of Agrobacterium strains whose T‐DNA are engineered with the gene(s)‐of‐interest. However, gene expression during “agro‐infiltration” is intrinsically and universally impeded by the onset of post‐...

Co-evolution between hosts' and parasites' genomes shapes diverse pathways of acquired immunity based on silencing small (s)RNAs. In plants, sRNAs cause heterochromatinization, sequence degeneration, and, ultimately, loss of autonomy of most transposable elements (TEs). Recognition of newly invasive plant TEs, by contrast, involves an innate antivi...

The Arabidopsis genome encodes ten Argonautes proteins showing distinct expression pattern as well as intracellular localisation during sexual reproduction.

In RNA interference (RNAi), small-interfering (si)RNAs processed from double-stranded RNA guide ARGONAUTE(AGO) proteins to silence sequence-complementary RNA/DNA. Plant RNAi can propagate locally and systemically, but despite recent mechanistic advances, basic questions/hurdles remain unaddressed. For instance, RNAi is inferred to diffuse through p...

Plant microRNAs (miRNAs) guide cytosolic post-transcriptional gene silencing of sequence-complementary transcripts within the producing cells, as well as in distant cells and tissues. Here, we used an artificial miRNA-based system (amiRSUL) in Arabidopsis thaliana to explore the still elusive mechanisms of inter-cellular miRNA movement via forward...

Supplementary Information related to Devers et al., 2020, Nature Plants including: Supplementary Figures 1-15; Supplementary Discussion; Supplementary Tables 1 and 2

Arabidopsis encodes ten ARGONAUTE (AGO) effectors of RNA silencing, canonically loaded with either 21‐22 nucleotide (nt) long small RNAs (sRNA) to mediate post‐transcriptional‐gene‐silencing (PTGS) or 24nt sRNAs to promote RNA‐directed‐DNA‐methylation. Using full‐locus constructs, we characterized the expression, biochemical properties, and possibl...

Diverse classes of silencing small (s)RNAs operate via ARGONAUTE-family proteins within RNA-induced-silencing-complexes (RISCs). Here, we have streamlined various embodiments of a Q-sepharose-based RISC-purification method that relies on conserved biochemical properties of all ARGONAUTEs. We show, in multiple benchmarking assays, that the resulting...

A bstract During sexual reproduction, development of a totipotent zygote from the fusion of highly differentiated gametes is accompanied by dynamic regulation of gene expression. This notably involves RNA silencing operated by Argonautes (AGO) effector proteins. While AGOs’ roles during Arabidopsis somatic life have been extensively investigated, l...

RNAi mediated by small-interfering RNAs (siRNAs) operates via transcriptional (TGS) and posttranscriptional gene silencing (PTGS). In Arabidopsis thaliana, TGS relies on DICER-LIKE-3 (DCL3)-dependent 24-nt siRNAs loaded into AGO4-clade ARGONAUTE effector proteins. PTGS operates via DCL4-dependent 21-nt siRNAs loaded into AGO1-clade proteins. We set...

Background Immunoglobulin A vasculitis (IgAV) is a small vessel vasculitis with IgA deposits in the vessel wall, manifesting with skin purpura, arthralgia/arthritis, gastrointestinal (GI) and renal complications. Altered serum miRNA signatures can reflect disease-specific pathology, which makes serum miRNAs promising disease biomarkers. We have rec...

In a reductionist perspective, plant silencing small (s)RNAs are often classified as mediating nuclear transcriptional-gene-silencing (TGS) or cytosolic post-transcriptional-gene-silencing (PTGS). Among the PTGS diagnostics is the association of AGOs and their sRNA cargoes with the translation apparatus. In Arabidopsis, this is observed for AGO1 lo...

The Extracellular RNA Communication Consortium (ERCC) was launched to accelerate progress in the new field of extracellular RNA (exRNA) biology and to establish whether exRNAs and their carriers, including extracellular vesicles (EVs), can mediate intercellular communication and be utilized for clinical applications. Phase 1 of the ERCC focused on...

Arabidopsis encodes ten ARGONAUTE (AGO) effectors of RNA silencing, canonically loaded with either 21-22nt small RNAs (sRNA) to mediate post-transcriptional-gene-silencing (PTGS) or 24nt sRNAs to promote RNA-directed-DNA-methylation. Using full-locus constructs, we characterized the expression, biochemical properties, and possible modes of action o...

In Arabidopsis thaliana, ARGONAUTE1 (AGO1) plays a central role in microRNA (miRNA) and small interfering RNA (siRNA)-mediated silencing and is a key component in antiviral responses. The polerovirus F-box P0 protein triggers AGO1 degradation as a viral counter-defence. Here, we identified a motif in AGO1 that is required for its interaction with t...

In eukaryotes, the RNase-III Dicer often produces length/sequence microRNA (miRNA) variants, called "isomiRs", owing to intrinsic structural/sequence determinants of miRNA precursors (pre-miRNAs). Here, we combined biophysics, genetics and biochemistry to study Arabidopsis miR168, the key feedback-regulator of the central plant silencing effector p...

The authors were made aware of possible issues with some figures published in the above-mentioned article. All figures were checked one by one, and clarifications are presented here pertaining to the complexity of some of the original mountings. The present amendments are based on the original raw data provided to the editors. Apart from an error m...

Retroelements, the prevalent class of plant transposons, have major impacts on host genome integrity and evolution. They produce multiple proteins from highly compact genomes and, similar to viruses, must have evolved original strategies to optimize gene expression, although this aspect has been seldom investigated thus far. Here, we have establish...

Significance As obligate photosynthetic and sessile organisms, plants are particularly exposed to the damaging effects of excess light and UV wavelengths, which can impact genome integrity by inducing DNA sequence alterations. As a response, plants have evolved efficient genome surveillance processes, some of which appear to also overlap with mecha...

In plants, tasiRNAs form a class of endogenous secondary siRNAs produced through the action of RNA-DEPENDENT-RNA-POLYMERASE-6 (RDR6) upon microRNA-mediated cleavage of non-coding TAS RNAs. In Arabidopsis thaliana, TAS1, TAS2 and TAS4 tasiRNA production proceeds via a single cleavage event mediated by 22nt-long or/and asymmetric miRNAs in an ARGONAU...

Phytoviruses encode viral suppressors of RNA-silencing (VSRs) to counteract the plant antiviral silencing response, which relies on virus-derived small interfering (si)RNAs processed by Dicer RNaseIII enzymes and subsequently loaded into ARGONAUTE (AGO) effector proteins. Here, a tobacco cell-free system was engineered to recapitulate the key steps...

Plant RNA silencing operates via RNA-directed DNA-methylation (RdDM) to repress transcription or by targeting mRNAs via posttranscriptional gene silencing (PTGS). These pathways rely on distinct Dicer-like (DCL) proteins that process doublestranded RNA (dsRNA) into small-interfering RNAs (siRNAs). Here, we explored the expression and subcellular lo...

Systemic, posttranscriptional silencing of transgenes in Nicotiana benthamiana was initiated in localized regions of the plant by introduction of transgene-homologous DNA fragments, including those without a promoter. Following this initiation step, a sequence-specific signal of gene silencing spread from cells that had received the ectopic DNA via...

[This corrects the article DOI: 10.1371/journal.ppat.1002035.].

Underlying data for Fig 5. (A) Original films and coomassie stainings used for mounting Fig 5A. Left panels: Western analysis of DCL1. Right panels: Western analysis of AGO1. (B) Original films used for mounting Fig 5B. Fig 5B was mounted from films obtained by sequentially stripping and re-hybridizing a single Northern blot membrane with several d...

Western blot analysis of the suppressor lines used in Jay et al., 2011. P15 was detected from 20 μg of total proteins from seedlings. The P15 antibody was used at a 1/10 000 dilution. P19 was detected from 100 μg of total proteins from seedlings. The P19 antibody was used at a 1/5000 dilution. Hc-Pro was detected from 20 μg of total proteins from s...

Materials and methods used for the analysis of the suppressor lines used in Jay et al., 2011, corresponding to S4 Fig, S5 Fig, S7 File and S8 File. (PDF)

Underlying data for Fig 4. (A) Original film (left) and ethidium bromide staining (right) used for mounting Fig 4C and Fig 4G. Both figures were mounted from the same blot, hybridized with a mix of random-labeled PCR products corresponding to the 35S terminator and HcPro, allowing simultaneous detection of the P19 and HcPro transcripts (bottom lane...

Genotyping of the suppressor lines used in Jay et al., 2011. (A) Total gDNA loaded on 1.2% agarose gel. (B) Primers used for PCR and sequencing. (C) ACTIN2 amplification (PCR of 28 cycles using primers Actin2-For and Actin2-Rev) loaded on 1.2% agarose gel. (D) PCR amplification of the CHS RNAi transgene (PCR of 30 cycles using primers 35S-For and C...

Raw sequencing data of the genomic DNA of the suppressor lines used in Jay et al., 2011, downloaded from GATC, in the ab1 format. (ZIP)

Detailed steps of the mounting of the corrected panels from Jay et al., 2011. (PDF)

Underlying data for Fig 6. (A) Original film (left) used for mounting Fig 6B and original ethidium bromide staining of the corresponding high molecular Northern gel (right). The blot was hybridized with a mix of random-labeled PCR products corresponding to the 35S terminator and HcPro, allowing detection of the P6 transcripts (upper lane). Blue rec...

Contigs of the raw sequences from the S7 File, for P15, P19 and HcPro from the suppressor lines used in Jay et al., 2011, using the CLC Genomics Workbench software. The hygromycin selection gene present in the HcPro transgenic line was also assembled. (ZIP)

In the Report “A plant miRNA contributes to antibacterial resistance by repressing auxin signaling,” a loading control panel in Fig. 2A was incorrectly assembled. The authors have retrieved the original small RNA blots with their cognate rRNA loading controls. The original data show that the Report’s biological conclusion (miR393 is induced in resp...

Small regulatory RNAs are fundamental in eukaryotic and prokaryotic gene regulation. In plants, an important element of post-transcriptional control is effected by 20–24 nt microRNAs (miRNAs) and short interfering RNAs (siRNAs) bound to the ARGONAUTE1 (AGO1) protein in an RNA induced silencing complex (RISC). AGO1 may cleave target mRNAs with small...

Replication of Cauliflower mosaic virus (CaMV), a plant double-stranded DNA virus, requires the viral translational transactivator protein P6. Although P6 is known to form cytoplasmic inclusion bodies (viroplasms) so far considered essential for virus biology, a fraction of the protein is also present in the nucleus. Here, we report that monomeric...

At the request of the authors, PLOS Pathogens is retracting this publication following an investigation into concerns about the origin and assembly of Figure 6 and a mounting mistake in Figure 1B. The Northern blot depicted in Figure 6 contains several band duplications affecting the panels labelled 'IP@HA' and 'total RNA'. The figure was provided...

The Extracellular RNA (exRNA) Communication Consortium was launched by the National Institutes of Health to focus on the extent to which RNA might function in a non-cell-autonomous manner. With the availability of increasingly sensitive tools, small amounts of RNA can be detected in serum, plasma, and other bodily fluids. The exact mechanism(s) by...

In transgenic and nontransgenic plants, viruses are both initiators and targets of a defense mechanism that is similar to posttranscriptional gene silencing (PTGS). Recently, it was found that potyviruses and cucumoviruses encode pathogenicity determinants that suppress this defense mechanism. Here, we test diverse virus types for the ability to su...

The above article from The EMBO Journal , published online on 22 April 2010, has been retracted by agreement between the authors, the journal Chief Editor and Head of Scientific Publications, EMBO, Bernd Pulverer, and John Wiley & Sons Ltd. The authors' and editors' statements are as follows. Following an investigation conducted by the The EMBO Jo...

[This corrects the article DOI: 10.1371/journal.ppat.1003883.].

Disclosed are a variety of methods for achieving enhanced expression from a target nucleotide sequence in a plant e.g. comprising the step of transiently introducing into a tissue of a plant (e.g. a leaf) a first nucleic acid comprising the target nucleotide sequence and a second nucleic acid encoding a Post Transcriptional Gene Silencing (PTGS) su...

(+)strand RNA viruses have to overcome various points of restriction in the host to establish successful infection. In plants, this includes RNA silencing. To uncover additional bottlenecks to RNA virus infection, we genetically attenuated the impact of RNA silencing on transgenically expressed Potato virus X (PVX), a (+)strand RNA virus that repli...

RNA silencing is an evolutionarily conserved mechanism triggered by double-stranded RNA that is processed into 21- to 24-nt small interfering (si)RNA or micro (mi)RNA by RNaseIII-like enzymes called Dicers. Gene regulations by RNA silencing have fundamental implications in a large number of biological processes that include antiviral defense, maint...

Although silent transposons in plants can be reactivated by stress or during development, their potential deleterious effects are prevented by transposon-derived epigenetically activated small interfering RNAs (easiRNAs). A new study shows how serendipitous interactions between reactivated transposons and endogenous microRNAs might initiate easiRNA...

In eukaryoticRNAsilencing, RNase-III classes of enzymes in the Dicer family process double-stranded RNA of cellular or exogenous origin into small- RNA(sRNA) molecules. sRNAs are then loaded into effector proteins known as ARGONAUTEs (AGOs), which, as part of RNA-induced silencing complexes, target complementary RNA or DNA for silencing. Land plant...

RNA-DEPENDENT RNA POLYMERASE 6 (RDR6) is a key RNA silencing factor initially characterized in transgene silencing and virus resistance. This enzyme also contributes to the biosynthesis of endogenous short interfering RNAs (siRNAs) from non-coding RNAs, transposable elements and protein-coding transcripts. One class of protein-coding transcripts th...

Author Summary A basal network of gene regulation orchestrates the processes ensuring maintenance of genome integrity. Eukaryotic small RNAs generated by the RNAse-III Dicer have emerged as central players in this network, by mediating gene silencing at the transcriptional or post-transcriptional level via RNA interference (RNAi). To gain insight i...

RNA silencing is a central regulator of gene expression in most eukaryotes and acts both at the transcriptional level through DNA methylation and at the post-transcriptional level through direct mRNA interference mediated by small RNAs. In plants and invertebrates, the same pathways also function directly in host defence against viruses by targetin...

In antiviral RNA interference (RNAi), the DICER enzyme processes virus-derived double-stranded RNA (dsRNA) into small interfering RNAs (siRNAs) that guide ARGONAUTE proteins to silence complementary viral RNA. As a counterdefense, viruses deploy viral suppressors of RNAi (VSRs). Well-established in plants and invertebrates, the existence of antivir...

Transposable elements (TEs) contribute to genome size, organization and evolution. In plants, their activity is primarily controlled by transcriptional gene silencing (TGS), usually investigated at steady states, reflecting how long-established silent conditions are maintained, faithfully reiterated or temporarily modified. How active, invasive TEs...