Gene Silencing by RNA Interference and the Role of Small Interfering RNAs

2004 ◽  
Author(s):  
Natasha Caplen
Keyword(s):  
Rna Interference ◽  
Gene Silencing ◽  
Small Interfering Rnas

scholarly journals Viroids as a Tool to Study RNA-Directed DNA Methylation in Plants

Cells ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 1187
Author(s):  
Michael Wassenegger ◽  
Athanasios Dalakouras
Keyword(s):  
Dna Methylation ◽  
Rna Interference ◽  
Gene Silencing ◽  
Plant Cells ◽  
Transcriptional Gene Silencing ◽  
Rnai Machinery ◽  
Double Stranded Rna ◽  
Post Transcriptional Gene Silencing ◽  
Cumulative Amount

Viroids are plant pathogenic, circular, non-coding, single-stranded RNAs (ssRNAs). Members of the Pospiviroidae family replicate in the nucleus of plant cells through double-stranded RNA (dsRNA) intermediates, thus triggering the host’s RNA interference (RNAi) machinery. In plants, the two RNAi pillars are Post-Transcriptional Gene Silencing (PTGS) and RNA-directed DNA Methylation (RdDM), and the latter has the potential to trigger Transcriptional Gene Silencing (TGS). Over the last three decades, the employment of viroid-based systems has immensely contributed to our understanding of both of these RNAi facets. In this review, we highlight the role of Pospiviroidae in the discovery of RdDM, expound the gradual elucidation through the years of the diverse array of RdDM’s mechanistic details and propose a revised RdDM model based on the cumulative amount of evidence from viroid and non-viroid systems.

scholarly journals Human Argonaute2 and Argonaute3 are catalytically activated by different lengths of guide RNA

2020 ◽  
Author(s):  
Mi Seul Park ◽  
GeunYoung Sim ◽  
Audrey C. Kehling ◽  
Kotaro Nakanishi
Keyword(s):  
Gene Silencing ◽  
Fold Increase ◽  
Specific Gene ◽  
Small Interfering Rnas ◽  
Guide Rna ◽  
Guide Rnas ◽  
Argonaute Proteins ◽  
Complementary Rnas ◽  
Standing Question

AbstractRNA interfering is a eukaryote-specific gene silencing by 20∼23 nucleotide (nt) microRNAs and small interfering RNAs that recruit Argonaute proteins to complementary RNAs for degradation. In humans, Argonaute2 (AGO2) has been known as the only slicer while Argonaute3 (AGO3) barely cleaves RNAs. Therefore, the intrinsic slicing activity of AGO3 remains controversial and a long-standing question. Here, we report 14-nt 3′ end-shortened variants of let-7a, miR-27a, and specific miR-17-92 families that make AGO3 an extremely competent slicer by an ∼ 82-fold increase in target cleavage. These RNAs, named cleavage-inducing tiny guide RNAs (cityRNAs), conversely lower the activity of AGO2, demonstrating that AGO2 and AGO3 have different optimum guide lengths for target cleavage. Our study sheds light on the role of tiny guide RNAs.

scholarly journals Development of an RNA interference (RNAi) gene knockdown protocol in the anaerobic gut fungus Pecoramyces ruminantium strain C1A

2017 ◽  
Author(s):  
Shelby S Calkins ◽  
Nicole C Elledge ◽  
Stephen M. Marek ◽  
M B. Couger ◽  
Mostafa S Elshahed ◽  
...  
Keyword(s):  
Rna Interference ◽  
Gene Silencing ◽  
Neurospora Crassa ◽  
Dna Helicase ◽  
Small Interfering Rnas ◽  
Gene Encoding ◽  
Interacting Protein ◽  
Genes Encoding ◽  
Lactate Levels ◽  
Electron Sink

Members of the anaerobic gut fungi (AGF) reside in rumen, hindgut, and feces of ruminant and non-ruminant herbivorous mammals and reptilian herbivores. No protocols for gene insertion, deletion, silencing, or mutation are currently available for the AGF, rendering gene-targeted molecular biological manipulations unfeasible. Here, we developed and optimized an RNA interference (RNAi)-based protocol for targeted gene silencing in the anaerobic gut fungus Pecoramyces ruminantium strain C1A. Analysis of the C1A genome identified genes encoding enzymes required for RNA silencing in fungi (Dicer, Argonaute, Neurospora crassa QDE-3 homolog DNA helicase, Argonaute-interacting protein, and Neurospora crassa QIP homolog exonuclease); and the competency of C1A germinating spores for RNA uptake was confirmed using fluorescently labeled small interfering RNAs (siRNA). Addition of chemically-synthesized siRNAs targeting D-lactate dehydrogenase (ldhD) gene to C1A germinating spores resulted in marked target gene silencing; as evident by significantly lower ldhD transcriptional levels, a marked reduction in the D-LDH specific enzymatic activity in intracellular protein extracts, and a reduction in D-lactate levels accumulating in the culture supernatant. Comparative transcriptomic analysis of untreated versus siRNA-treated cultures identified a few off-target siRNA-mediated gene silencing effects. As well, significant differential up-regulation of the gene encoding NAD-dependent 2-hydroxyacid dehydrogenase (Pfam00389) in siRNA-treated C1A cultures was observed, which could possibly compensate for loss of D-LDH as an electron sink mechanism in C1A. The results demonstrate the feasibility of RNAi in anaerobic fungi, and opens the door for gene silencing-based studies in this fungal clade.

scholarly journals Exploring the functions of RNA interference pathway proteins: some functions are more RISCy than others?

2005 ◽  
Vol 387 (3) ◽  
pp. 561-571 ◽  
Author(s):  
Katarzyna JARONCZYK ◽  
Jon B. CARMICHAEL ◽  
Tom C. HOBMAN
Keyword(s):  
Rna Interference ◽  
Gene Silencing ◽  
Cell Cycle Regulators ◽  
Small Interfering Rnas ◽  
Piwi Domain ◽  
Therapeutic Technique ◽  
Silencing Pathways ◽  
Laboratory Tool ◽  
Molecular Therapeutic ◽  
Regulate Gene

PPD (PAZ Piwi domain) proteins and the Dicer family have been the subjects of intense study over the last 6 years. These proteins have well-established roles in RNAi (RNA interference), a process that relies on siRNAs (small interfering RNAs) or miRNAs (microRNAs) to mediate specificity. The development of techniques for applying RNAi as a laboratory tool and a molecular therapeutic technique has rapidly outpaced our understanding of the biology of this process. However, over the last 2 years, great strides have been made towards elucidating how PPD proteins and Dicer regulate gene-silencing at the pre- and post-transcriptional levels. In addition, evidence is beginning to emerge that suggests that these proteins have additional siRNA-independent roles as cell-cycle regulators. In the present review, we summarize the well-known roles of these two classes of proteins in gene-silencing pathways, as well as explore the evidence for novel roles of PPD and Dicer proteins.

scholarly journals Human Argonaute2 and Argonaute3 are catalytically activated by different lengths of guide RNA

2020 ◽  
Vol 117 (46) ◽  
pp. 28576-28578
Author(s):  
Mi Seul Park ◽  
GeunYoung Sim ◽  
Audrey C. Kehling ◽  
Kotaro Nakanishi
Keyword(s):  
Gene Silencing ◽  
Specific Gene ◽  
Small Interfering Rnas ◽  
Guide Rna ◽  
Guide Rnas ◽  
Argonaute Proteins ◽  
Rna Interfering ◽  
Complementary Rnas ◽  
Standing Question

RNA interfering is a eukaryote-specific gene silencing by 20∼23-nucleotide (nt) microRNAs and small interfering RNAs that recruit Argonaute proteins to complementary RNAs for degradation. In humans, Argonaute2 (AGO2) has been known as the only slicer while Argonaute3 (AGO3) barely cleaves RNAs. Therefore, the intrinsic slicing activity of AGO3 remains controversial and a long-standing question. Here, we report 14-nt 3′ end-shortened variants of let-7a, miR-27a, and specific miR-17–92 families that make AGO3 an extremely competent slicer, increasing target cleavage up to ∼82-fold in some instances. These RNAs, named cleavage-inducing tiny guide RNAs (cityRNAs), conversely lower the activity of AGO2, demonstrating that AGO2 and AGO3 have different optimum guide lengths for target cleavage. Our study sheds light on the role of tiny guide RNAs.

scholarly journals Development of an RNA interference (RNAi) gene knockdown protocol in the anaerobic gut fungusPecoramyces ruminantiumstrain C1A

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e4276 ◽  
Author(s):  
Shelby S. Calkins ◽  
Nicole C. Elledge ◽  
Katherine E. Mueller ◽  
Stephen M. Marek ◽  
MB Couger ◽  
...  
Keyword(s):  
Rna Interference ◽  
Gene Silencing ◽  
Neurospora Crassa ◽  
Dna Helicase ◽  
Small Interfering Rnas ◽  
Gene Encoding ◽  
Interacting Protein ◽  
Genes Encoding ◽  
Lactate Levels ◽  
Electron Sink

Members of the anaerobic gut fungi (AGF) reside in rumen, hindgut, and feces of ruminant and non-ruminant herbivorous mammals and reptilian herbivores. No protocols for gene insertion, deletion, silencing, or mutation are currently available for the AGF, rendering gene-targeted molecular biological manipulations unfeasible. Here, we developed and optimized an RNA interference (RNAi)-based protocol for targeted gene silencing in the anaerobic gut fungusPecoramyces ruminantiumstrain C1A. Analysis of the C1A genome identified genes encoding enzymes required for RNA silencing in fungi (Dicer, Argonaute,Neurospora crassaQDE-3 homolog DNA helicase, Argonaute-interacting protein, andNeurospora crassaQIP homolog exonuclease); and the competency of C1A germinating spores for RNA uptake was confirmed using fluorescently labeled small interfering RNAs (siRNA). Addition of chemically-synthesized siRNAs targeting D-lactate dehydrogenase (ldhD) gene to C1A germinating spores resulted in marked target gene silencing; as evident by significantly lowerldhDtranscriptional levels, a marked reduction in the D-LDH specific enzymatic activity in intracellular protein extracts, and a reduction in D-lactate levels accumulating in the culture supernatant. Comparative transcriptomic analysis of untreated versus siRNA-treated cultures identified a few off-target siRNA-mediated gene silencing effects. As well, significant differential up-regulation of the gene encoding NAD-dependent 2-hydroxyacid dehydrogenase (Pfam00389) in siRNA-treated C1A cultures was observed, which could possibly compensate for loss of D-LDH as an electron sink mechanism in C1A. The results demonstrate the feasibility of RNAi in anaerobic fungi, and opens the door for gene silencing-based studies in this fungal clade.

scholarly journals Cross-Kingdom Small RNAs Among Animals, Plants and Microbes

Cells ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 371 ◽  
Author(s):  
Zeng ◽  
Gupta ◽  
Jiang ◽  
Yang ◽  
Gong ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Interference ◽  
Gene Silencing ◽  
Horizontal Transfer ◽  
Small Rnas ◽  
Signal Transmission ◽  
Fungal Diseases ◽  
Small Interfering Rnas ◽  
Eukaryotic Species ◽  
Non Coding Rnas

Small RNAs (sRNAs), a class of regulatory non-coding RNAs around 20~30-nt long, including small interfering RNAs (siRNAs) and microRNAs (miRNAs), are critical regulators of gene expression. Recently, accumulating evidence indicates that sRNAs can be transferred not only within cells and tissues of individual organisms, but also across different eukaryotic species, serving as a bond connecting the animal, plant, and microbial worlds. In this review, we summarize the results from recent studies on cross-kingdom sRNA communication. We not only review the horizontal transfer of sRNAs among animals, plants and microbes, but also discuss the mechanism of RNA interference (RNAi) signal transmission via cross-kingdom sRNAs. We also compare the advantages of host-induced gene silencing (HIGS) and spray-induced gene silencing (SIGS) technology and look forward to their applicable prospects in controlling fungal diseases.

scholarly journals Development of an RNA interference (RNAi) gene knockdown protocol in the anaerobic gut fungus Pecoramyces ruminantium strain C1A

2017 ◽  
Author(s):  
Shelby S Calkins ◽  
Nicole C Elledge ◽  
Stephen M. Marek ◽  
M B. Couger ◽  
Mostafa S Elshahed ◽  
...  
Keyword(s):  
Rna Interference ◽  
Gene Silencing ◽  
Neurospora Crassa ◽  
Dna Helicase ◽  
Small Interfering Rnas ◽  
Gene Encoding ◽  
Interacting Protein ◽  
Genes Encoding ◽  
Lactate Levels ◽  
Electron Sink

Members of the anaerobic gut fungi (AGF) reside in rumen, hindgut, and feces of ruminant and non-ruminant herbivorous mammals and reptilian herbivores. No protocols for gene insertion, deletion, silencing, or mutation are currently available for the AGF, rendering gene-targeted molecular biological manipulations unfeasible. Here, we developed and optimized an RNA interference (RNAi)-based protocol for targeted gene silencing in the anaerobic gut fungus Pecoramyces ruminantium strain C1A. Analysis of the C1A genome identified genes encoding enzymes required for RNA silencing in fungi (Dicer, Argonaute, Neurospora crassa QDE-3 homolog DNA helicase, Argonaute-interacting protein, and Neurospora crassa QIP homolog exonuclease); and the competency of C1A germinating spores for RNA uptake was confirmed using fluorescently labeled small interfering RNAs (siRNA). Addition of chemically-synthesized siRNAs targeting D-lactate dehydrogenase (ldhD) gene to C1A germinating spores resulted in marked target gene silencing; as evident by significantly lower ldhD transcriptional levels, a marked reduction in the D-LDH specific enzymatic activity in intracellular protein extracts, and a reduction in D-lactate levels accumulating in the culture supernatant. Comparative transcriptomic analysis of untreated versus siRNA-treated cultures identified a few off-target siRNA-mediated gene silencing effects. As well, significant differential up-regulation of the gene encoding NAD-dependent 2-hydroxyacid dehydrogenase (Pfam00389) in siRNA-treated C1A cultures was observed, which could possibly compensate for loss of D-LDH as an electron sink mechanism in C1A. The results demonstrate the feasibility of RNAi in anaerobic fungi, and opens the door for gene silencing-based studies in this fungal clade.

scholarly journals Chemical strategies for strand selection in short-interfering RNAs

RSC Advances ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 2415-2426
Author(s):  
Andrew J. Varley ◽  
Jean-Paul Desaulniers
Keyword(s):  
Rna Interference ◽  
Gene Silencing ◽  
Rnai Pathway ◽  
Specific Gene ◽  
Small Interfering Rnas ◽  
Chemical Strategies

Therapeutic small interfering RNAs (siRNAs) are double stranded RNAs capable of potent and specific gene silencing through activation of the RNA interference (RNAi) pathway.

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