Substrate-specific structural rearrangements of human Dicer

Taylor, David W.; Ma, Enbo; Shigematsu, Hideki; Cianfrocco, Michael A.; Noland, Cameron L.; Nagayama, Kuniaki; Nogales, Eva; Doudna, Jennifer A.; Wang, Hong Wei

doi:10.1038/nsmb.2564

Cited by 94 publications

(111 citation statements)

References 59 publications

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“…However, this is not to say that dsRNA binding does not contribute to the interaction. LGP2, Dicer, PACT and PKR all contain one or more dsRNA‐binding domains, and binding to ligand induces conformational rearrangements that may alter their ability to associate with other proteins (Cole, 2007; Taylor et al , 2013; Heyam et al , 2015; Uchikawa et al , 2016). Our experimental results with different LGP2 domains likely reflect a scenario in which RNA‐bound CTD undergoes structural rearrangements that allow it to associate with Dicer, an interaction that is then further strengthened in a cooperative manner by the presence of the helicase domain.…”

Section: Discussionmentioning

confidence: 99%

The RIG‐I‐like receptor LGP2 inhibits Dicer‐dependent processing of long double‐stranded RNA and blocks RNA interference in mammalian cells

et al. 2018

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In vertebrates, the presence of viral RNA in the cytosol is sensed by members of the RIG‐I‐like receptor (RLR) family, which signal to induce production of type I interferons (IFN). These key antiviral cytokines act in a paracrine and autocrine manner to induce hundreds of interferon‐stimulated genes (ISGs), whose protein products restrict viral entry, replication and budding. ISGs include the RLRs themselves: RIG‐I, MDA5 and, the least‐studied family member, LGP2. In contrast, the IFN system is absent in plants and invertebrates, which defend themselves from viral intruders using RNA interference (RNAi). In RNAi, the endoribonuclease Dicer cleaves virus‐derived double‐stranded RNA (dsRNA) into small interfering RNAs (siRNAs) that target complementary viral RNA for cleavage. Interestingly, the RNAi machinery is conserved in mammals, and we have recently demonstrated that it is able to participate in mammalian antiviral defence in conditions in which the IFN system is suppressed. In contrast, when the IFN system is active, one or more ISGs act to mask or suppress antiviral RNAi. Here, we demonstrate that LGP2 constitutes one of the ISGs that can inhibit antiviral RNAi in mammals. We show that LGP2 associates with Dicer and inhibits cleavage of dsRNA into siRNAs both in vitro and in cells. Further, we show that in differentiated cells lacking components of the IFN response, ectopic expression of LGP2 interferes with RNAi‐dependent suppression of gene expression. Conversely, genetic loss of LGP2 uncovers dsRNA‐mediated RNAi albeit less strongly than complete loss of the IFN system. Thus, the inefficiency of RNAi as a mechanism of antiviral defence in mammalian somatic cells can be in part attributed to Dicer inhibition by LGP2 induced by type I IFNs. LGP2‐mediated antagonism of dsRNA‐mediated RNAi may help ensure that viral dsRNA substrates are preserved in order to serve as targets of antiviral ISG proteins.

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Section: Discussionmentioning

confidence: 99%

The RIG‐I‐like receptor LGP2 inhibits Dicer‐dependent processing of long double‐stranded RNA and blocks RNA interference in mammalian cells

et al. 2018

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“…Conformation Population Analysis-Conformation population analysis similar to a previous study was conducted (23). Particle measurements were done using ImageJ (24).…”

Section: Methodsmentioning

confidence: 99%

Conformational Flexibility and Subunit Arrangement of the Modular Yeast Spt-Ada-Gcn5 Acetyltransferase Complex

Setiaputra

Ross

et al. 2015

Journal of Biological Chemistry

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Background:The Saccharomyces cerevisiae Spt-Ada-Gcn5 acetyltransferase (SAGA) complex regulates transcription through chromatin modification and other mechanisms. Results: The overall structure of SAGA and the arrangement of all subunits within this complex were determined. Conclusion: SAGA is flexible and is composed of core modules that support peripheral catalytic modules. Significance: Understanding the structural mechanisms of SAGA multifunctionality improves the understanding of other chromatin-modifying complexes.

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“…However, full-length mammalian Dicer proteins appear unable to process long dsRNAs into small interfering RNAs (siRNAs) Flemr et al 2013;Taylor et al 2013) and infection of mammalian somatic cells with RNA viruses does not result in the detection of significant levels of viral siRNAs (Parameswaran et al 2010;Cullen et al 2013). Drosophila, in contrast, encodes two Dicer proteins, dDcr1 and dDcr2.…”

Section: Synthetic Microrna Duplexes Efficiently Program Risc In Dicementioning

confidence: 99%

Derivation and characterization of Dicer- and microRNA-deficient human cells

Bogerd

Whisnant

Kennedy

et al. 2014

RNA

147

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We have used genome editing to generate inactivating deletion mutations in all three copies of the dicer (hdcr) gene present in the human cell line 293T. As previously shown in murine ES cells lacking Dicer function, hDcr-deficient 293T cells are severely impaired for the production of mature microRNAs (miRNAs). Nevertheless, RNA-induced silencing complexes (RISCs) present in these hDcr-deficient cells are readily programmed by transfected, synthetic miRNA duplexes to repress mRNAs bearing either fully or partially complementary targets, including targets bearing incomplete seed homology to the introduced miRNA. Using these hDcr-deficient 293T cells, we demonstrate that human pre-miRNA processing can be effectively rescued by ectopic expression of the Drosophila Dicer 1 protein, but only in the presence of the PB isoform of Loquacious (Loqs-PB), the fly homolog of the hDcr cofactor TRBP. In contrast, Drosophila Dicer 2, even in the presence of its cofactors Loqs-PD and R2D2, was unable to support human pre-miRNA processing. Interestingly, although ectopic Drosophila Dicer 1/Loqs-PB or hDcr both rescued pre-miRNA processing effectively in these hDcr-deficient cells, there were significant differences in the ratio of the miRNA isoforms that were produced, especially in the case of miR-30 family members, and we also noted differences in the relative expression level of miRNAs vs. passenger strands for a subset of human miRNAs. These data demonstrate that the mechanisms underlying the accurate processing of pre-miRNAs are largely, but not entirely, conserved between mammalian and insect cells.

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Substrate-specific structural rearrangements of human Dicer

Cited by 94 publications

References 59 publications

The RIG‐I‐like receptor LGP2 inhibits Dicer‐dependent processing of long double‐stranded RNA and blocks RNA interference in mammalian cells

The RIG‐I‐like receptor LGP2 inhibits Dicer‐dependent processing of long double‐stranded RNA and blocks RNA interference in mammalian cells

Conformational Flexibility and Subunit Arrangement of the Modular Yeast Spt-Ada-Gcn5 Acetyltransferase Complex

Derivation and characterization of Dicer- and microRNA-deficient human cells

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