ARGONAUTE2 cooperates with SWI/SNF complex to determine nucleosome occupancy at human Transcription Start Sites

Carissimi, Claudia; Laudadio, Ilaria; Cipolletta, Emanuela; Gioiosa, Silvia; Mihailovich, Marija; Bonaldi, Tiziana; Macino, Giuseppe; Fulci, Valerio

doi:10.1093/nar/gku1387

Cited by 38 publications

(41 citation statements)

References 57 publications

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“…The previous studies identifying AGO2 interacting partners focused on preparations from whole cells, ribosomal gradient fractions, or chromatin (Meister et al 2005;Höck et al 2007;Robb and Rana 2007;Landthaler et al 2008;Weinmann et al 2009;Ameyar-Zazoua et al 2012;Frohn et al 2012;Carissimi et al 2015). One of our goals was to analyze cytoplasmic and nuclear interactions using extracts isolated from each compartment.…”

Section: Purification Of Nuclear or Cytoplasmic Ago2mentioning

confidence: 99%

“…We also tested the potential for AGO2 interactions with the protein SMARCC1 that had been identified in a whole cell study (Carissimi et al 2015). SMARCC1 is an intriguing candidate because the protein is primarily nuclear and has functions in the SWI/SNF complex.…”

Section: Evaluating Interactions With Predicted Protein Partnersmentioning

confidence: 99%

“…Harel-Bellan and colleagues examined interactions between proteins with Flag-AGO1 and Flag-AGO2 and identified SRSF splicing factors as candidate interacting partners (Ameyar-Zazoua et al 2012). Another recent study by Carissimi et al (2015) in whole cell extracts reported interactions between endogenous AGO2 and SWI/SNF proteins normally found in the nucleus.…”

Section: Introductionmentioning

confidence: 99%

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Stable association of RNAi machinery is conserved between the cytoplasm and nucleus of human cells

Kalantari

Hicks

et al. 2016

RNA

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Argonaute 2 (AGO2), the catalytic engine of RNAi, is typically associated with inhibition of translation in the cytoplasm. AGO2 has also been implicated in nuclear processes including transcription and splicing. There has been little insight into AGO2's nuclear interactions or how they might differ relative to cytoplasm. Here we investigate the interactions of cytoplasmic and nuclear AGO2 using semi-quantitative mass spectrometry. Mass spectrometry often reveals long lists of candidate proteins, complicating efforts to rigorously discriminate true interacting partners from artifacts. We prioritized candidates using orthogonal analytical strategies that compare replicate mass spectra of proteins associated with Flag-tagged and endogenous AGO2. Interactions with TRNC6A, TRNC6B, TNRC6C, and AGO3 are conserved between nuclei and cytoplasm. TAR binding protein interacted stably with cytoplasmic AGO2 but not nuclear AGO2, consistent with strand loading in the cytoplasm. Our data suggest that interactions between functionally important components of RNAi machinery are conserved between the nucleus and cytoplasm but that accessory proteins differ. Orthogonal analysis of mass spectra is a powerful approach to streamlining identification of protein partners.

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Section: Purification Of Nuclear or Cytoplasmic Ago2mentioning

confidence: 99%

Section: Evaluating Interactions With Predicted Protein Partnersmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Stable association of RNAi machinery is conserved between the cytoplasm and nucleus of human cells

Kalantari

Hicks

et al. 2016

RNA

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“…Alternatively, phenotypic differences could be related to functions of AGO1 other than miRNA target repression. Indeed, the implication of AGO proteins in several pathways such as nonsense-mediated mRNA decay (Choe et al, 2010), alternative splicing (AmeyarZazoua et al, 2012;Yang et al, 2012b;Taliaferro et al, 2013;Alló et al, 2014), DNA double-strand break repair (Michalik et al, 2012;Wei et al, 2012), nucleosome occupancy at transcription start sites (Carissimi et al, 2015), and quality control of mRNAs encoding proteins entering the secretory pathway (Karamyshev et al, 2014) has been proposed during the last years. If that is the case, the conditional slicer-deficient alleles might be a valuable tool to investigate novel functions of plant AGO1 by decoupling these from target regulation.…”

Section: Slicer Activity In Mirna Biogenesis and Accumulationmentioning

confidence: 99%

mRNA Decay of Most Arabidopsis miRNA Targets Requires Slicer Activity of AGO1

2016

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“…AGO2 was shown to promote transcriptional gene silencing in a microRNA (miRNA)-dependent manner (12) and to affect nucleosome occupancy at certain transcription start sites through interaction with the SWI/SNF complex (13). Furthermore, AGO1 has been shown to interact with actively transcribed genes and enhancer regions (14,15).…”

mentioning

confidence: 99%

Argonaute 2 Binds Directly to tRNA Genes and Promotes Gene Repression in cis

Woolnough

Atwood

Giles

2015

Molecular and Cellular Biology

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To further our understanding of the RNAi machinery within the human nucleus, we analyzed the chromatin and RNA binding of Argonaute 2 (AGO2) within human cancer cell lines. Our data indicated that AGO2 binds directly to nascent tRNA and 5S rRNA, and to the genomic loci from which these RNAs are transcribed, in a small RNA-and DICER-independent manner. AGO2 chromatin binding was not observed at non-TFIIIC-dependent RNA polymerase III (Pol III) genes or at extra-TFIIIC (ETC) sites, indicating that the interaction is specific for TFIIIC-dependent Pol III genes. A genome-wide analysis indicated that loss of AGO2 caused a global increase in mRNA expression level among genes that flank AGO2-bound tRNA genes. This effect was shown to be distinct from that of the disruption of DICER, DROSHA, or CTCF. We propose that AGO2 binding to tRNA genes has a novel and important regulatory role in human cells. The RNA interference (RNAi) machinery has several important functions that are conserved from fission yeast to humans. In the cytoplasm of eukaryotic cells, Argonaute proteins promote degradation and/or translational repression of specific mRNAs through tethering by a complementary small RNA (1). The nuclear functions of the RNAi machinery appear to be diverse across species and include roles in promoting heterochromatin formation, regulating alternative splicing, and promoting insulator function (2-5). The ability of both Argonaute 1 (AGO1) and AGO2 to alter splicing in human cells depends upon small RNAs derived from exonic sequence, which tether Argonaute proteins to nascent transcripts (3, 6). Tethering of each Argonaute protein was shown to be capable of recruiting a histone lysine methyltransferase, which altered the RNA polymerase II (Pol II) elongation rate and thus facilitated alternative splicing. This interaction between Argonaute proteins and the transcriptional apparatus is conserved in Drosophila melanogaster (7).Despite the similarities between Argonaute proteins, AGO2 is the only human Argonaute protein believed to have the endonuclease activity (Slicer) due to a unique amino-terminal region (8-10), and Ago2 but not Ago1 gene deletions are embryonic lethal in mice (11). AGO2 was shown to promote transcriptional gene silencing in a microRNA (miRNA)-dependent manner (12) and to affect nucleosome occupancy at certain transcription start sites through interaction with the SWI/SNF complex (13). Furthermore, AGO1 has been shown to interact with actively transcribed genes and enhancer regions (14, 15). Recent work has identified expanded binding capabilities of AGO2, which include binding to longer RNAs such as pre-miRNAs (65 to 75 nucleotides [nt]) and full-length tRNAs (ϳ75 nt) (16)(17)(18)(19)(20). To wit, AGO2 was recently shown to functionally interact with DICER in human nuclei but was unable to load duplex small interfering RNA (siRNA), indicating that nuclear RNAi processes may proceed through a mechanism distinct from that observed in the cytoplasm (21). These findings indicate that AGO2 may have roles beyo...

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ARGONAUTE2 cooperates with SWI/SNF complex to determine nucleosome occupancy at human Transcription Start Sites

Cited by 38 publications

References 57 publications

Stable association of RNAi machinery is conserved between the cytoplasm and nucleus of human cells

Stable association of RNAi machinery is conserved between the cytoplasm and nucleus of human cells

mRNA Decay of Most Arabidopsis miRNA Targets Requires Slicer Activity of AGO1

Argonaute 2 Binds Directly to tRNA Genes and Promotes Gene Repression in cis

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