FUS stimulates microRNA biogenesis by facilitating co-transcriptional Drosha recruitment

Morlando, Mariangela; Modigliani, Stefano Dini; Torrelli, Giulia; Rosa, Alessandro; Carlo, V. Di; Caffarelli, Elisa; Bozzoni, Irene

doi:10.1038/emboj.2012.319

Cited by 206 publications

(204 citation statements)

References 40 publications

(84 reference statements)

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“…2b). In contrast, no upregulation was observed for the miR-15a and miR-432 miRNAs, previously shown to be unaffected by alteration in FUS levels 16 . Moreover, as a consequence of the miR-141/200a increase obtained after Dox induction of flag-FUS, we observed that the Zeb1-previously reported to be post-transcriptionally repressed by the members of miR-200 family 17 -was downregulated (Fig.…”

Section: Resultsmentioning

confidence: 50%

“…Figure 3a shows that FUS interacts in vitro with both pri-miR-141 and pri-miR-200a. Specific binding also occurs with the positive control pri-miR-9-2, while no interaction is observed with the negative control, pri-miR-15a 16 . According to previous data indicating the chromatin localization of FUS 16 , chromatin immunorecipitation with anti-FUS antibodies revealed a specific localization of FUS on the chromosomal loci encoding for miR-141 and miR-200a, while no localization was detected on the negative control, miR-15a (Fig.…”

Section: Resultsmentioning

confidence: 94%

“…FUS was previously described as a Drosha interactor 20 and shown to enhance miRNA expression by direct binding to nascent pri-miRNAs on chromatin and facilitating co-transcriptional processing 16,21 . To dissect the mechanism by which FUS affects miR-141/200a biogenesis, we therefore tested these features on miR-141 and miR-200a.…”

Section: Resultsmentioning

confidence: 99%

“…epB-Puro-TT-derived plasmids for overexpressing RFP, RFP-FUS and flag-FUS fusion cDNAs are described in Morlando et al 16 The FUS-G48A mutant construct was obtained by inverse PCR amplification on FUS-WT plasmid (SC320263, OriGene Technologies) using the oligonucleotides FUS-G48A forward (FW) and reverse (REV), while the FUS-D3 0 -UTR mutant construct was generated by PCR amplification on FUS-WT plasmid using the oligonuclotides FUS-D3 0 -UTR FW and REV. HA-FUS-WT and HA-FUS-G48A were generated by insertion of the HA tag in both FUS-WT and FUS-G48A plasmids through a reverse PCR using the HA-5 0 FUS FW and REV oligonucleotides.…”

Section: Ha-fus-g48amentioning

confidence: 99%

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An ALS-associated mutation in the FUS 3′-UTR disrupts a microRNA–FUS regulatory circuitry

et al. 2014

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While the physiologic functions of the RNA-binding protein FUS still await thorough characterization, the pathonegetic role of FUS mutations in amyotrophic lateral sclerosis (ALS) is clearly established. Here we find that a human FUS mutation that leads to increased protein expression, and was identified in two ALS patients with severe outcome, maps to the seed sequence recognized by miR-141 and miR-200a in the 3 0 -UTR of FUS. We demonstrate that FUS and these microRNAs are linked by a feed-forward regulatory loop where FUS upregulates miR-141/200a, which in turn impact FUS protein synthesis. We also show that Zeb1, a target of miR-141/200a and transcriptional repressor of these two microRNAs, is part of the circuitry and reinforces it. Our results reveal a possible correlation between deregulation of this regulatory circuit and ALS pathogenesis, and open interesting perspectives in the treatment of these mutations through ad hoc-modified microRNAs.

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Section: Resultsmentioning

confidence: 50%

Section: Resultsmentioning

confidence: 94%

Section: Resultsmentioning

confidence: 99%

Section: Ha-fus-g48amentioning

confidence: 99%

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An ALS-associated mutation in the FUS 3′-UTR disrupts a microRNA–FUS regulatory circuitry

et al. 2014

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“…In the light of the involvement of DROSHA in DDR signaling, its predominantly nuclear localization and of its reported co-transcriptional role in the context of microRNA processing (Morlando et al, 2012), (Gromak et al, 2013), we wondered if this endoribonuclase is recruited to sites of DNA damage to locally process newly synthetized dilncRNA into DDRNAs. To test this hypothesis, we took advantage of the DIvA cellular system (for DSB inducible via AsiSI), (Iacovoni et al, 2010), (Aymard et al, 2014), , (Aymard et al, 2017) a clonal U2OS cell line that stably expresses the AsiSI restriction enzyme fused with a modified oestrogen receptor ligand binding domain.…”

Section: Drosha Is Recruited To Dsbsmentioning

confidence: 99%

DROSHA associates to DNA damage sites and is required for DNA repair

Cabrini

Roncador

Galbiati

et al. 2018

Preprint

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The DNA damage response (DDR) is the signaling cascade through which a cell recognizes DNA lesions, and promotes their resolution via the repair pathways of Non-Homologous End Joining (NHEJ), or Homologous Recombination (HR). We recently demonstrated that DROSHA boosts DDR signaling by processing damage-induced long non-coding RNAs into smaller DNA damage response RNAs (DDRNAs). However, the location at which DROSHA exerts its DDR functions, relative to sites of DNA damage, remains unknown.To investigate DROSHA's localization during DDR activation, we used the DiVA cellular system, which allows the controlled induction of several DNA double strand breaks (DSBs) in the human genome. Indeed, by genome wide chromatin immunoprecipitation followed by next generation sequencing, we demonstrate that DROSHA associates with DSBs. In support of this, DSB-recruitment of DROSHA is detectable at the single-cell level by Proximity Ligation Assay between DROSHA and known DDR markers, and by DNA damage in situ ligation followed by Proximity Ligation Assay (DI-PLA), which demonstrates proximity of DROSHA to DNA ends. DROSHA recruitment occurs at both genic and inter-genic DSBs, suggesting that its recruitment is independent from ongoing transcription preceding damage generation. DROSHA's recruitment to DNA lesions occurs throughout the cell cycle, and with a preference for NHEJ-prone DSBs. Consistently, inhibition of the HR pathway increases DROSHA recruitment, and DROSHA knock down strongly impairs NHEJ efficiency in a GFP-reporter cellular system for monitoring NHEJ DNA repair. Overall, these results demonstrate that DROSHA acts locally at sites of DNA damage to promote NHEJ DNA repair.

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TDP‐43 and FUS en route from the nucleus to the cytoplasm

Ederle¹,

2017

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Misfolded or mislocalized RNA‐binding proteins (RBPs) and, consequently, altered mRNA processing, can cause neuronal dysfunction, eventually leading to neurodegeneration. Two prominent examples are the RBPs TAR DNA‐binding protein of 43 kDa (TDP‐43) and fused in sarcoma (FUS), which form pathological messenger ribonucleoprotein aggregates in patients suffering from amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two devastating neurodegenerative disorders. Here, we review the multiple functions of TDP‐43 and FUS in mRNA processing, both in the nucleus and in the cytoplasm. We discuss how TDP‐43 and FUS may exit the nucleus and how defects in both nuclear and cytosolic mRNA processing events, and possibly nuclear export defects, may contribute to neurodegeneration and ALS/FTD pathogenesis.

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FUS stimulates microRNA biogenesis by facilitating co-transcriptional Drosha recruitment

Cited by 206 publications

References 40 publications

An ALS-associated mutation in the FUS 3′-UTR disrupts a microRNA–FUS regulatory circuitry

An ALS-associated mutation in the FUS 3′-UTR disrupts a microRNA–FUS regulatory circuitry

DROSHA associates to DNA damage sites and is required for DNA repair

TDP‐43 and FUS en route from the nucleus to the cytoplasm

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