High-Level Accumulation of Exogenous Small RNAs Not Affecting Endogenous Small RNA Biogenesis and Function in Plants

SummaryPlant small RNAs regulate key physiological mechanisms through post-transcriptional and transcriptional silencing of gene expression. sRNAs fall into two major categories: those that are reliant on RNA Dependent RNA Polymerases (RDRs) for biogenesis and those that aren’t. Known RDR-dependent sRNAs include phased and repeat-associated short interfering RNAs, while known RDR-independent sRNAs are primarily microRNAs and other hairpin-derived sRNAs. In this study, we produced and analyzed small RNA-seq libraries from rdr1/rdr2/rdr6 triple mutant plants. Only a small fraction of all sRNA loci were RDR1/RDR2/RDR6-independent; most of these were microRNA loci or associated with predicted hairpin precursors. We found 58 previously annotated microRNA loci that were reliant on RDR1, −2, or −6 function, casting doubt on their classification. We also found 38 RDR1/2/6-independent small RNA loci that are not MIRNAs or otherwise hairpin-derived, and did not fit into other known paradigms for small RNA biogenesis. These 38 small RNA-producing loci have novel biogenesis mechanisms, and are frequently located in the vicinity of protein-coding genes. Altogether, our analysis suggest that these 38 loci represent one or more new types of small RNAs in Arabidopsis thaliana.Significance StatementSmall RNAs regulate gene expression in plants and are produced through a variety of previously-described mechanisms. Here, we examine a set of previously undiscovered small RNA-producing loci that are produced by novel mechanisms.

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Translation and codon usage regulate Argonaute slicer activity to trigger small RNA biogenesis

10.1101/2020.09.04.282863 ◽

2020 ◽

Author(s):

Meetali Singh ◽

Eric Cornes ◽

Blaise Li ◽

Piergiuseppe Quarato ◽

Loan Bourdon ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Embryonic Development ◽

Rna Polymerase ◽

Codon Usage ◽

Small Rna ◽

Small Rnas ◽

Rna Dependent Rna Polymerase ◽

Coding Sequences ◽

Germ Granules ◽

Rna Biogenesis

In the Caenorhabditis elegans germline, thousands of mRNAs are concomitantly expressed with antisense 22G-RNAs, which are loaded into the Argonaute CSR-1. Despite their essential functions for animal fertility and embryonic development, how CSR-1 22G-RNAs are produced remains unknown. Here, we show that CSR-1 slicer activity is primarily involved in triggering the synthesis of small RNAs on the coding sequences of germline mRNAs and post-transcriptionally regulates a fraction of targets. CSR-1-cleaved mRNAs prime the RNA-dependent RNA polymerase, EGO-1, to synthesize 22G-RNAs in phase with ribosome translation in the cytoplasm, in contrast to other 22G-RNAs mostly synthesized in germ granules. Moreover, codon optimality and efficient translation antagonize CSR-1 slicing and 22G-RNAs biogenesis. We propose that codon usage differences encoded into mRNA sequences might be a conserved strategy in eukaryotes to regulate small RNA biogenesis and Argonaute targeting.

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Diversification of small RNA amplification mechanisms for targeting transposon-related sequences in ciliates

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1903491116 ◽

2019 ◽

Vol 116 (29) ◽

pp. 14639-14644 ◽

Cited By ~ 3

Author(s):

Masatoshi Mutazono ◽

Tomoko Noto ◽

Kazufumi Mochizuki

Keyword(s):

Transposable Elements ◽

Small Rna ◽

Signal Amplification ◽

Small Rnas ◽

Ciliated Protozoa ◽

Rna Amplification ◽

Somatic Nucleus ◽

Dna Elimination ◽

Internal Eliminated Sequences ◽

Rna Biogenesis

The silencing of repetitive transposable elements (TEs) is ensured by signal amplification of the initial small RNA trigger, which occurs at distinct steps of TE silencing in different eukaryotes. How such a variety of secondary small RNA biogenesis mechanisms has evolved has not been thoroughly elucidated. Ciliated protozoa perform small RNA-directed programmed DNA elimination of thousands of TE-related internal eliminated sequences (IESs) in the newly developed somatic nucleus. In the ciliate Paramecium, secondary small RNAs are produced after the excision of IESs. In this study, we show that in another ciliate, Tetrahymena, secondary small RNAs accumulate at least a few hours before their derived IESs are excised. We also demonstrate that DNA excision is dispensable for their biogenesis in this ciliate. Therefore, unlike in Paramecium, small RNA amplification occurs before IES excision in Tetrahymena. This study reveals the remarkable diversity of secondary small RNA biogenesis mechanisms, even among ciliates with similar DNA elimination processes, and thus raises the possibility that the evolution of TE-targeting small RNA amplification can be traced by investigating the DNA elimination mechanisms of ciliates.

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MiR&moRe2: A Bioinformatics Tool to Characterize microRNAs and microRNA-Offset RNAs from Small RNA-Seq Data

International Journal of Molecular Sciences ◽

10.3390/ijms21051754 ◽

2020 ◽

Vol 21 (5) ◽

pp. 1754 ◽

Cited By ~ 3

Author(s):

Enrico Gaffo ◽

Michele Bortolomeazzi ◽

Andrea Bisognin ◽

Piero Di Battista ◽

Federica Lovisa ◽

...

Keyword(s):

Small Rna ◽

Small Rnas ◽

Cell Types ◽

Small Rna Sequencing ◽

Rna Seq ◽

Sequencing Data ◽

Research Findings ◽

Human Blood Cell ◽

And Function ◽

Comprehensive Study

MicroRNA-offset RNAs (moRNAs) are microRNA-like small RNAs generated by microRNA precursors. To date, little is known about moRNAs and bioinformatics tools to inspect their expression are still missing. We developed miR&moRe2, the first bioinformatics method to consistently characterize microRNAs, moRNAs, and their isoforms from small RNA sequencing data. To illustrate miR&moRe2 discovery power, we applied it to several published datasets. MoRNAs identified by miR&moRe2 were in agreement with previous research findings. Moreover, we observed that moRNAs and new microRNAs predicted by miR&moRe2 were downregulated upon the silencing of the microRNA-biogenesis pathway. Further, in a sizeable dataset of human blood cell populations, tens of novel miRNAs and moRNAs were discovered, some of them with significantly varied expression levels among the cell types. Results demonstrate that miR&moRe2 is a valid tool for a comprehensive study of small RNAs generated from microRNA precursors and could help to investigate their biogenesis and function.

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Assembly and function of small RNA – Argonaute protein complexes

Biological Chemistry ◽

10.1515/hsz-2014-0116 ◽

2014 ◽

Vol 395 (6) ◽

pp. 611-629 ◽

Cited By ~ 42

Author(s):

Anne Dueck ◽

Gunter Meister

Keyword(s):

Gene Expression ◽

Gene Silencing ◽

Small Rna ◽

Small Rnas ◽

Atp Hydrolysis ◽

Protein Complexes ◽

Protein Family ◽

Argonaute Protein ◽

Argonaute Proteins ◽

And Function

Abstract Small RNAs such as microRNAs (miRNAs), short interfering RNAs (siRNAs) or Piwi-interacting RNAs (piRNAs) are important regulators of gene expression in various organisms. Small RNAs bind to a member of the Argonaute protein family and are incorporated into larger structures that mediate diverse gene silencing events. The loading of Argonaute proteins with small RNAs is aided by a number of auxiliary factors as well as ATP hydrolysis. This review will focus on the mechanisms of Argonaute loading in different organisms. Furthermore, we highlight the versatile functions of small RNA-Argonaute protein complexes in organisms from all three kingdoms of life.

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Translation and codon usage regulate Argonaute slicer activity to trigger small RNA biogenesis

10.21203/rs.3.rs-87655/v1 ◽

2020 ◽

Author(s):

Germano Cecere ◽

Meetali Singh ◽

Eric Cornes ◽

Blaise Li ◽

Piergiuseppe Quarato ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Embryonic Development ◽

Rna Polymerase ◽

Codon Usage ◽

Small Rna ◽

Small Rnas ◽

Rna Dependent Rna Polymerase ◽

Coding Sequences ◽

Germ Granules ◽

Rna Biogenesis

Abstract In the Caenorhabditis elegans germline, thousands of mRNAs are concomitantly expressed with antisense 22G-RNAs, which are loaded into the Argonaute CSR-1. Despite their essential functions for animal fertility and embryonic development, how CSR-1 22G-RNAs are produced remains unknown. Here, we show that CSR-1 slicer activity is primarily involved in triggering the synthesis of small RNAs on the coding sequences of germline mRNAs and post-transcriptionally regulates a fraction of targets. CSR-1-cleaved mRNAs prime the RNA-dependent RNA polymerase, EGO-1, to synthesize 22G-RNAs in phase with ribosome translation in the cytoplasm, in contrast to other 22G-RNAs mostly synthesized in germ granules. Moreover, codon optimality and efficient translation antagonize CSR-1 slicing and 22G-RNAs biogenesis. We propose that codon usage differences encoded into mRNA sequences might be a conserved strategy in eukaryotes to regulate small RNA biogenesis and Argonaute targeting.

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From transcription to translation: new insights in the structure and function of Argonaute protein

BioMolecular Concepts ◽

10.1515/bmc-2012-0024 ◽

2012 ◽

Vol 3 (6) ◽

pp. 545-559

Author(s):

Corinna Giorgi ◽

Carlo Cogoni ◽

Caterina Catalanotto

Keyword(s):

Small Rna ◽

Small Rnas ◽

Biological Function ◽

Rna Molecules ◽

Argonaute Proteins ◽

Ancestral Function ◽

Higher Eukaryotes ◽

Silencing Pathways ◽

Genome Protection ◽

And Function

AbstractArgonaute proteins play a central role in gene silencing pathways mediated by small RNA molecules. The ancestral function of small RNA-dependent silencing is related to genome protection against parasitic nucleic acids, such as transposons and viruses. However, new classes of small RNAs are continuously being uncovered in all higher eukaryotes in which they play important functions in processes ranging from embryonic development to differentiation to cell proliferation and metabolism. Small RNAs have variegated biogenesis pathways and accomplish distinct functions. Nevertheless, it appears that all small RNAs work merely as guides in recognizing the target RNAs invariably relying on the interaction with Argonaute proteins and associated factors for their biological function. Here, we discuss recent findings on the structure and regulation of mammalian Argonaute proteins and overview the various roles that these versatile proteins play in regulating gene expression.

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Small RNAs Asserting Big Roles in Mycobacteria

10.20944/preprints202107.0044.v3 ◽

2021 ◽

Author(s):

Fatma S. Coskun ◽

Przemyslaw Plocinski ◽

Nicolai S.C. van Oers

Keyword(s):

Small Rna ◽

Small Rnas ◽

Mammalian Cells ◽

Noncoding Rnas ◽

Locked Nucleic Acid ◽

Processing Enzymes ◽

Open Questions ◽

Rna Probes ◽

And Function

Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (Mtb), with 10.4 million new cases per year reported in the human population. Recent studies on the Mtb transcriptome have revealed the abundance of noncoding RNAs expressed at various phases of mycobacteria growth, in culture, in infected mammalian cells and in patients. Among these noncoding RNAs are both small RNAs (sRNAs) between 50-350 nts in length and smaller RNAs (sncRNA) <50 nts. In this review, we provide an up-to-date synopsis of the identification, designation, and function of these Mtb-encoded sRNAs and sncRNAs. The methodological advances including RNA sequencing strategies, small RNA antagonists and locked nucleic acid sequence specific RNA probes advancing the studies on these small RNA are described. Initial insights into the regulation of the small RNA expression and putative processing enzymes required for their synthesis and function are discussed. There are many open questions remaining about the biological and pathogenic roles of these small non-coding RNAs, and potential research directions needed to define the role of these mycobacterial noncoding RNAs summarized.

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Translation and codon usage regulate Argonaute slicer activity to trigger small RNA biogenesis

Nature Communications ◽

10.1038/s41467-021-23615-w ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Meetali Singh ◽

Eric Cornes ◽

Blaise Li ◽

Piergiuseppe Quarato ◽

Loan Bourdon ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Embryonic Development ◽

Rna Polymerase ◽

Codon Usage ◽

Small Rna ◽

Small Rnas ◽

Rna Dependent Rna Polymerase ◽

Coding Sequences ◽

Germ Granules ◽

Rna Biogenesis

AbstractIn the Caenorhabditis elegans germline, thousands of mRNAs are concomitantly expressed with antisense 22G-RNAs, which are loaded into the Argonaute CSR-1. Despite their essential functions for animal fertility and embryonic development, how CSR-1 22G-RNAs are produced remains unknown. Here, we show that CSR-1 slicer activity is primarily involved in triggering the synthesis of small RNAs on the coding sequences of germline mRNAs and post-transcriptionally regulates a fraction of targets. CSR-1-cleaved mRNAs prime the RNA-dependent RNA polymerase, EGO-1, to synthesize 22G-RNAs in phase with translating ribosomes, in contrast to other 22G-RNAs mostly synthesized in germ granules. Moreover, codon optimality and efficient translation antagonize CSR-1 slicing and 22G-RNAs biogenesis. We propose that codon usage differences encoded into mRNA sequences might be a conserved strategy in eukaryotes to regulate small RNA biogenesis and Argonaute targeting.

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L2 acquisition of the bei passive in Mandarin Chinese: A constructionist approach

Chinese as a Second Language Research ◽

10.1515/caslar-2020-0007 ◽

2020 ◽

Vol 9 (2) ◽

pp. 169-198

Author(s):

Chen Chen ◽

Feng-hsi Liu

Keyword(s):

Mandarin Chinese ◽

Choice Task ◽

L2 Acquisition ◽

Low Level ◽

Passive Construction ◽

Form And Function ◽

L2 Learners ◽

Constructionist Approach ◽

High Level ◽

And Function

Abstract A major claim in the constructionist approach to language acquisition is that grammar is learned by pairings of form and function. In this study we test this claim by examining how L2 learners of Mandarin Chinese acquire the bei passive construction, a construction that is associated with the meaning of adversity. Our goal is to find out whether L2 learners make the association between the passive and adversity. Participants performed a sentence choice task under four conditions: an adversative context with an adversative verb, an adversative context with a neutral verb, a neutral context with a neutral verb and a positive context with a neutral verb. In each context participants were asked to select either the bei passive construction or its active counterpart. We found that high-level learners consistently chose the bei passive significantly more in adversative contexts than in non-adversative contexts regardless of the connotations of the verbs, while low-level learners made the distinction half of the time. In addition, while low-level learners did not yet associate adversity with the form of the construction, high-level learners did. We conclude that L2 learners do learn the bei passive construction as a form-meaning pair. The constructionist approach is supported.

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