MicroRNA Biogenesis via Splicing and Exosome-Mediated Trimming in Drosophila

Flynt, Alex S.; Greimann, Jaclyn C.; Chung, Wei-Jen; Lima, Christopher D.; Lai, Eric C.

doi:10.1016/j.molcel.2010.06.014

Cited by 162 publications

(157 citation statements)

References 42 publications

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“…It was previously reported that the exosome trims 3 ′ -tailed mirtrons (Flynt et al 2010). We used siRNA to knock down Dis3 (also known as Rrp44), a core component of both the nuclear and cytoplasmic exosome complexes (Tomecki and Dziembowski 2010), followed by transfection of pHSUR4.…”

Section: Resultsmentioning

confidence: 99%

The host Integrator complex acts in transcription-independent maturation of herpesvirus microRNA 3′ ends

et al. 2015

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Herpesvirus saimiri (HVS) is an oncogenic γ-herpesvirus that produces microRNAs (miRNAs) by cotranscription of precursor miRNA (pre-miRNA) hairpins immediately downstream from viral small nuclear RNAs (snRNA). The host cell Integrator complex, which recognizes the snRNA 3 ′ end processing signal (3 ′ box), generates the 5 ′ ends of HVS pre-miRNA hairpins. Here, we identify a novel 3 ′ box-like sequence (miRNA 3 ′ box) downstream from HVS premiRNAs that is essential for miRNA biogenesis. In vivo knockdown and rescue experiments confirmed that the 3 ′ end processing of HVS pre-miRNAs also depends on Integrator activity. Interaction between Integrator and HVS primary miRNA (pri-miRNA) substrates that contain only the miRNA 3 ′ box was confirmed by coimmunoprecipitation and an in situ proximity ligation assay (PLA) that we developed to localize specific transient RNA-protein interactions inside cells. Surprisingly, in contrast to snRNA 3 ′ end processing, HVS pre-miRNA 3 ′ end processing by Integrator can be uncoupled from transcription, enabling new approaches to study Integrator enzymology.

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Section: Resultsmentioning

confidence: 99%

The host Integrator complex acts in transcription-independent maturation of herpesvirus microRNA 3′ ends

et al. 2015

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“…If DBR1 is a slow enzyme compared with nuclear exonucleases or if artificial mirtrons are unusually stable as lariats, partial digestion of the 39 end of the lariat could prevent functional hairpin formation or, alternatively, result in variable hairpin formation, which could underlie the difference in mirtron and shRNA efficiency. In support, a recent study has shown that miR-1017, a 39-tailed mirtron in Drosophila melanogaster, is processed by 39 exonuclease to form the pre-miRNA (Flynt et al 2010). Sequencing results of artificial mirtrons may have missed these truncated 39 sequences because RNA was sized pre-selection, thus further work needs to be undertaken to elucidate the debranching step in mirtron biogenesis and the rules governing the intrinsic stability of introns.…”

Section: Dmpk-mirt 5 Knockdown Is Dependent On Splicing and Exportin-mentioning

confidence: 99%

Artificial mirtron-mediated gene knockdown: Functional DMPK silencing in mammalian cells

Seow¹,

Sibley²,

Wood³

2012

RNA

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Mirtrons are introns that form pre-miRNA hairpins after splicing to produce RNA interference (RNAi) effectors distinct from Drosha-dependent intronic miRNAs. Here we present a design algorithm for artificial mirtrons and demonstrate, for the first time, efficient gene knockdown of myotonic dystrophy protein kinase (DMPK) target sequences in Renilla luciferase 39 UTR and subsequently pathogenic DMPK mRNA, causative of Type I myotonic dystrophy, using artificial mirtrons cloned as eGFP introns. Deep sequencing of artificial mirtrons suggests that functional mature transcripts corresponding to the designed sequence were produced in high abundance. They were further shown to be splicing-dependent, Drosha-independent, and partially dependent on exportin-5, resulting in the precise generation of pre-miRNAs. In a murine myoblast line containing a pathogenic copy of human DMPK with more than 500 CUG repeats, the DMPK artificial mirtron corrected DM1-associated splicing abnormalities of the Serca-1 mRNA, demonstrating the therapeutic potential of mirtron-mediated RNAi. Thus, further development and exploitation of the unique properties of mirtrons will benefit future research and therapeutic RNAi applications as an alternative to conventional RNAi strategies.

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“…Both ends of mirtrons are defined by the splice sites. A related, mirtron-like source of small RNAs requires both splicing and exosome-mediated trimming to extract the pre-microRNA hairpin [24,25]. In this case only one end of the precursor hairpin is defined by the splicing reaction.…”

Section: Introductionmentioning

confidence: 99%

MicroRNA or Not MicroRNA?

Langenberger

Bartschat²,

Hertel

et al. 2011

Lecture Notes in Computer Science

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Abstract. The avalanche of next generation sequencing data has led to a rapid increase of annotated microRNAs in the last few years. Many of them are specific to individual species or rather narrow clades. A closer inspection of the current version of miRBase shows that dozens of entries conflict with other ncRNAs, in particular snoRNAs. With few exceptions, these cases show little similarities to canonical microRNAs, however, and thus they should be considered as mis-annotations.

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MicroRNA Biogenesis via Splicing and Exosome-Mediated Trimming in Drosophila

Cited by 162 publications

References 42 publications

The host Integrator complex acts in transcription-independent maturation of herpesvirus microRNA 3′ ends

The host Integrator complex acts in transcription-independent maturation of herpesvirus microRNA 3′ ends

Artificial mirtron-mediated gene knockdown: Functional DMPK silencing in mammalian cells

MicroRNA or Not MicroRNA?

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