KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| Rabbit monoclonal anti-Ago2 | Cell Signaling Technology | Cat#2897; Clone: C34C6; RRID: AB_2096291 |
| Mouse monoclonal anti-β-actin | Sigma-Aldrich | Cat#A2228; Clone: AC-74; RRID: AB_476697 |
| Mouse monoclonal anti-Tubulin | Sigma-Aldrich | Cat#T9026; Clone: DM1A; RRID: AB_477593 |
| Mouse monoclonal anti-HaloTag | Promega | Cat#G9211; RRID: AB_2688011 |
| Rabbit polyclonal anti-FLAG | Sigma-Aldrich | Cat#F7425; RRID: AB_439687 |
| Rabbit polyclonal anti-GW182 (TNRC6A) | Bethyl | Cat#A302–329A; RRID: AB_1850240 |
| Donkey anti-Rabbit IgG polyclonal antibody (IRDye 800CW) | LI-COR Biosciences | Cat#926–32213; RRID: AB_621848 |
| Donkey anti-Mouse IgG polyclonal antibody (IRDye 680RD) | LI-COR Biosciences | Cat#926–68072; RRID: AB_10953628 |
| Donkey anti-Rabbit IgG, HRP-conjugated | GE Healthcare | Cat#NA934; RRID: AB_772206 |
| Sheep anti-Mouse IgG, HRP-conjugated | GE Healthcare | Cat#NA931; RRID: AB_772210 |
| Bacterial and Virus Strains | ||
| pAd-Cas9-EA (Ad-EA) | ViraQuest Inc.; Maddalo et al., 2014 | N/A |
| pAd-Cas9-BN (Ad-BN) | ViraQuest Inc.; Cook et al., 2017 | N/A |
| Ad-Cre | ViraQuest Inc. | N/A |
| Critical Commercial Assays | ||
| Dual-Luciferase Reporter Assay System | Promega | Cat#E1960 |
| RNeasy Mini Kit | QIAGEN | Cat#74104 |
| TruSeq Stranded mRNA LT Kit | Illumina | Cat#RS-122–2102 |
| TruSeq Stranded Total RNA LT Kit | Illumina | Cat#RS-122–1202 |
| Chemicals, Peptides, and Recombinant Proteins | ||
| HaloTag Ligands TMRDirect | Promega | Cat#G2991 |
| Janelia Fluor 646 HaloTag ligand | Promega | Cat#GA1120 |
| Janelia Fluor 549 HaloTag ligand | Promega | Cat#GA1110 |
| Amersham ECL Western Blotting Detection Reagent | GE Healthcare | Cat#RPN2106 |
| Odyssey Blocking Buffer (TBS) | LI-COR Biosciences | Cat#927–50003 |
| KnockOut DMEM | Gibco | Cat#10829018 |
| Fetal Bovine Serum, embryonic stem cell-qualified, US origin | Gibco | Cat#16141079 |
| ESGRO Leukemia Inhibitory Factor (LIF) | Millipore | Cat#ESG1107 |
| Penicillin-Streptomycin (5,000 U/mL) | Gibco | Cat#15070063 |
| GlutaMax Supplement | Gibco | Cat#35050061 |
| MEM non-essential amino acid solution (100X) | Sigma-Aldrich | Cat#M7145 |
| EmbryoMax Nucleosides (100X) | Millipore | Cat#ES-008-D |
| 2-Mercaptoethanol | BIO-RAD | Cat#1610710, CAS: 60–24-2 |
| DMEM | Gibco | Cat#11965118 |
| DMEM, no phenol red | Gibco | Cat#21063029 |
| Advanced DMEM/F12 | Gibco | Cat#12634028 |
| HEPES (1M) | Gibco | Cat#15630080 |
| Geneticin Selective Antibiotic (G418 Sulfate) | Gibco | Cat#10131035 |
| Halolink Resin | Promega | Cat#G1914 |
| Magne HaloTag Beads | Promega | Cat#G7282 |
| PhosStop, Phosphatase inhibitor | Roche | Cat#4906837001 |
| cOmplete, Mini, EDTA-free Protease Inhibitor Cocktail | Roche | Cat#11836170001 |
| AcTEV protease | Invitrogen | Cat#12575015 |
| Protease Inhibitor Cocktail, 50x | Promega | Cat#G6521 |
| RQ1 RNase-Free DNase | Promega | Cat#M6101 |
| RNase A | Affymetrix | Cat#70194Z |
| Alkaline Phosphatase, Calf Intestinal (CIAP) | Promega | Cat#M1821 |
| T4 RNA Ligase 1 | New England Biolabs | Cat#M0204 |
| T4 Polynucleotide Kinase | New England Biolabs | Cat#M0201 |
| SuperScript III Reverse Transcriptase | Invitrogen | Cat#18080044 |
| Accuprime Pfx Supermix | Invitrogen | Cat#12344040 |
| Proteinase K, recombinant, PCR Grade | Roche | Cat#3115836001 |
| Phenol solution | Sigma-Aldrich | Cat#P4682 |
| Chloroform – isoamyl alcohol mixture | Sigma-Aldrich | Cat#25668 |
| Novex TBE-Urea Gels, 15%, 10 well | Invitrogen | Cat#EC6885BOX |
| Novex TBE Gels, 6%, 10 well | Invitrogen | Cat#EC6265BOX |
| SYBR Green I Nucleic Acid Gel Stain | Invitrogen | Cat#S7563 |
| Dynabeads MyOne Silane | ThermoFisher Scientific | Cat#37002D |
| TRIzol Reagent | Invitrogen | Cat#15596026 |
| RNase-Free DNase Set | QIAGEN | Cat#79254 |
| NEBuilder HiFi DNA Assembly Master Mix | New England Biolabs | Cat#E2621L |
| cOmplete Protease Inhibitor Cocktail (Mass Spectrometry) | Roche | Cat#11836153001 |
| Trypsin Gold, Mass Spectrometry Grade | Promega | Cat#V5280 |
| Lysyl Endopeptidase (Lys-C) | Wako Chemicals | Cat#129–02541 |
| TMT10plex Isobaric Label Reagent Set plus TMT11–131C Label Reagent | Thermo Fisher Scientific | Cat#A34808 |
| Oligonucleotides | ||
| See Table S1. | N/A | N/A |
| Deposited Data | ||
| HEAP libraries and RNA-seq datasets | This paper | GSE139349 |
| TT-FHAgo2 mESC iCLIP | Bosson et al., 2014 | GSE61348 |
| TT-FHAgo2 mESC RNAseq | Bosson et al., 2014 | GSE61348 |
| CLEAR-CLIP | Moore et al., 2015 | GSE73059 |
| miR-17~92 E9.5 embryo RNA-seq | Han et al., 2015 | GSE63813 |
| miR-124 overexpression CAD | Makeyev et al., 2007 | GSE8498 |
| miRbase version 21 | Kozomara et al., 2018 | ftp://mirbase.org/pub/mirbase/21/ |
| TargetScan | Agarwal et al., 2015 | http://www.targetscan.org/mmu_71/mmu_71_data_download/miR_Family_Info.txt.zip |
| Experimental Models: Cell Lines | ||
| Ago2-/- MEFs | Alexander Tarakhovsky Laboratory; O’Carroll et al., 2007 | N/A |
| Ago2Halo-LSL/+ mESCs | This paper | N/A |
| Ago2Halo/+ mESCs | This paper | N/A |
| Ago2Halo-LSL MEFs | This paper | N/A |
| Ago2Halo MEFs | This paper | N/A |
| KRasG12D;Trp53-/- (KP) NSCLC cell lines | This paper | N/A |
| V6.5 mESCs | Rudolf Jaenisch Laboratory | N/A |
| Experimental Models: Organisms/Strains | ||
| Mouse: KRasLSL-G12D/+: B6.129S4-Krastm4Tyj/J | Jackson et al., 2001 | JAX Stock No. 008179 |
| Mouse: Trp53fl/fl: B6.129P2-Trp53tm1Brn/J | Marino et al., 2000 | JAX Stock No. 008462 |
| Mouse: β-actin-Flpe: B6;SJL-Tg(ACTFLPe)9205Dym/J | Memorial Sloan Kettering Cancer Center Mouse Genetics Core Facility; Rodriguez et al., 2000 | JAX Stock No. 003800 |
| Mouse: CAG-Cre: Tg(CAG-cre)13Miya | Memorial Sloan Kettering Cancer Center Mouse Genetics Core Facility; Sakai and Miyazaki, 1997 | N/A |
| Mouse: Ago2Halo-LSL | This paper | N/A |
| Mouse: Ago2Halo | This paper | N/A |
| Mouse: miR-17~92-/- | Ventura et al., 2008 | N/A |
| Recombinant DNA | ||
| MSCV-PIG-Empty | Mayr and Bartel, 2009 | Addgene: 21654 |
| MSCV-PIG-Halo | This paper | N/A |
| MSCV-PIG-Ago2 | This paper | N/A |
| MSCV-PIG-Halo-Ago2 | This paper | N/A |
| pKOII-Halo-lox-IRES-lox-Ago2 | This paper | N/A |
| pMB1610_pRR-puro | Flemr et al., 2015 | Addgene: 65853 |
| PX330 | Cong et al., 2013, Ran et al., 2013 | Addgene: 42230 |
| PX333 | Maddalo et al., 2014 | Addgene: 64073 |
| piS0 | Yekta et al., 2004 | Addgene: 12178 |
| piS1 | Addgene | Addgene: 12179 |
| psico luc + Cre | Ventura et al., 2004 | N/A |
| psico CD8 + Cre | Ventura et al., 2004 | N/A |
| pBABE-SV40-puro | Zhao et al., 2003 | Addgene:13970 |
| pTRETightBI-RY-0 | Mukherji et al., 2011 | Addgene: 31463 |
| pTRETightBI-RY-1pf | Mukherji et al., 2011 | Addgene: 31467 |
| pTRETightBI-RY-4 | Mukherji et al., 2011 | Addgene: 31465 |
| rtTA-N144 | Richner et al., 2015 | Addgene: 66810 |
| psiCheck2 | Promega | C8021 |
| psiCheck2-Pten-miR-29-WT | This paper | N/A |
| psiCheck2-Pten-miR-29-MUT | This paper | N/A |
| psiCheck2-Adrb2-let-7-WT | This paper | N/A |
| psiCheck2-Adrb2-let-7-MUT | This paper | N/A |
| psiCheck2-Taf7-miR-21-WT | This paper | N/A |
| psiCheck2-Taf7-miR-21-MUT | This paper | N/A |
| Turn-T6BWT-EYFP | This paper | N/A |
| Turn-T6BMUT-EYFP | This paper | N/A |
| Software and Algorithms | ||
| STAR (v2.5.3a) | Dobin et al., 2013 | https://github.com/alexdobin/STAR |
| Cutadapt (v1.15 and v1.17) | Martin, 2011 | https://cutadapt.readthedocs.io |
| Bowtie2 (v2.3.4) | Langmead and Salzberg, 2012 | http://bowtie-bio.sourceforge.net/bowtie2 |
| DESeq2 (v1.20.0, v1.6.3 and 1.22.1) | Love et al., 2014 | https://bioconductor.org/packages/release/bioc/html/DESeq2.html |
| IDR | Li et al., 2011 | https://github.com/nboley/idr |
| Deeptools (v3.1.3) | Ramirez et al., 2016 | https://deeptools.readthedocs.io |
| Bedtools (v2.23.0) | Quinlan and Hall, 2010 | https://bedtools.readthedocs.io |
| kentUtils | Kent et al., 2010 | https://github.com/ENCODE-DCC/kentUtils |
| Limma (v3.38.3) | Ritchie et al., 2015 | http://bioconductor.org/packages/release/bioc/html/limma.html |
| featureCounts (v1.6.3) | Liao et al., 2014 | http://bioinf.wehi.edu.au/featureCounts |
| Genomation (v1.14.0) | Akalin et al., 2015 | https://bioconductor.org/packages/release/bioc/html/genomation.html |
| HISAT2 (v0.1.6-beta) | Kim et al., 2019 | https://ccb.jhu.edu/software/hisat2 |
| GEOquery (v2.50.5) | Davis and Meltzer, 2007 | https://bioconductor.org/packages/release/bioc/html/GEOquery.html |
| Samtools (v1.3.1) | Li et al., 2009 | http://www.htslib.org |
| Parametric Analysis of Gene Set Enrichment | Kim and Volsky, 2005 | N/A |
| HOMER | Heinz et al., 2010 | http://homer.ucsd.edu/homer/ |
| CLIPanalyze (v0.0.8) | unpublished | https://bitbucket.org/leslielab/clipanalyze |
| Sequest | Eng et al., 1994 | http://proteomicsresource.washington.edu/protocols06/sequest.php |
| Odyssey | LI-COR Biosciences | https://www.licor.com/bio/ |
| IGV | Robinson et al., 2011 | http://software.broadinstitute.org/software/igv/ |
| ZEN | ZEISS | https://www.zeiss.com/microscopy/int/downloads.html |
| Fiji | NIH | https://imagej.net/Fiji |
| Other | ||
| Superose 6 column | GE Healthcare | Cat#10/300 GL |
| Sep-Pak C18 3 cc Vac Cartridge, 200 mg | Waters | Cat#WAT054945 |