Table 2.
Functional microRNA arrayed screenings
| Library/method | Cell model | Screening method | Readouts | No of hits (examples) # | Refs. |
|---|---|---|---|---|---|
| 266 mouse miRNAs (Dharmacon)/LT SM | Mouse ESC DGCR8 knockout | Manual SP | Relative cell number (MTT assay) | 14 (miR-19a, miR-20a/b, miR-33, miR-93, miR-106a, miR-223, miR-291a/b-3p, miR-294, miR-295, miR-302b/c/d) | [96] |
| 379 mouse miRNAs (Ambion)/LT SM | $ MEFs from OG2 mice (Oct4-GFP) Reprograming (OKS cassette lentiviral) | Semi-Automated FM | Oct4-GFP+ iPSC Colony counts | 27, 14 validated (miR-130a/b, −148a, − 152, − 190, − 301, −301b, −669b, − 721 and miR-290 and miR-302 cluster members); siRNA KD of 130/301/721 family target Meox2 increased reprograming. | [90] |
| 52 candidates ESC-specific miRNAs/EP Amaxa, PB-CAG-miRNA piggybac vector | $ MEFs (Oct4-GFP-Puro) Reprograming (PB-CAG-OCKS transposon) | Manual FC (Cytomics FC500 series, Beckman Coulter) | Resistance to Puromycin and % of GFP+ cells. | 4 (miR-302 cluster, miR-25, miR-290, miR-298); miR-25 targets Wwp2 and Fbxw7. | [101] |
| 875 human miRs (Ambion)/LT SM | * Mouse ESCs Myh6-eGFP (cardiac-specific) | HCS (InCell Analyzer 1000, GE) | Integrated fluorescence intensity of Myh6-eGFP | 19 (let-7a, let-7b, let-7c, let-7d, let-7e, let-7f, let-7 g, let-7i, miR-98, mus-miR-202, miR-18a, miR-18b); KD of let-7 and miR-18 targets (Acvr1b and Smad2, respectively) promoted mesoderm differentiation. | [103, 104] |
| 570 Mouse miRNAs (Dharmacon)/LT SM | $ MEFs from OG2 mice (Oct4-GFP) Reprograming (OSK cassette retrovirus) | HCS (InCell Analyzer 2000, GE) | Oct4-GFP+ iPSC Colony counts | 16 (miRs − 294, −302a/b/d, −467d,-181b/d,-19a*, −34c*, −467d, − 294, − 677, − 451, −30d, − 590-3p, − 144, − 324-3p, − 455-5p). siRNA KD of miR-294 targets (Cdkn1a, Zfp148, Hivep2, Ddhd1, Dpysl2, Pten, Cfl2, 9530068E07Rik) or miR-181 targets (Bptf, Lin7c, Cpsf6, Nr2c2, Bclaf1, Nol8, Igf2bp2, Marcks) increased reprograming. | [102] |
| 40 mouse miRNAs (Dharmacon)/LT SM | Mouse ESC DGCR8 knockout | Manual (varied) | AP colony staining, morphology, cell cycle, gene expression pattern and Oct4 staining | 14 (miR-218-5p, 200c-3p, 129-5p, 135b-5p, 24-3p, 9-5p, 32-5p and 27a-3p); miR-27a/24 targets Oct4, Foxo1, gp130, Smads; increased reprogramming (anti-miRs − 27a and − 24). | [97] |
| 21 miR highly expressed or significantly upregulated during ESCs to EpiLC transition/LT SM | @ Dgcr8−/− Naïve mESCs to EpiSCs transition | Manual qRT-PCR | Naïve markers Rex1 and Klf2 and epiblast marker Fgf5 | 7 (miR-291a-3p, miR-294-3p, miR-295-3p, miR-302a/b/c/d-3p); siRNA KD of miR-290/302 family target Akt1 promoted Naïve to Primed transition.. | [98] |
| 379 mouse miRNAs (Ambion)/LT SM | $ MEFs from OG2 mice (Oct4-GFP) Reprograming (OKS cassette lentiviral) | Semi-Automated FM | Oct4-GFP+ iPSC Colony counts | 14 (miRNA-212/132); siRNA KD of miRNA-212/132 targets p300 and Jarid1a hampers reprogramming. | [92] |
| 31 human miRNAs differentially expressed upon differentiation of pluripotent cells (Ambion)/LT SM | Human NTERA2 EC and Human H1 ESCs | Manual HCS (ImageXpress Micro, Molecular Devices) | Multiparametric Phenotypic Profile (Hoechst/CellMask, anti-Oct4 and – CycB1) |
Lineage dependent (miR-22-3p, miR-27a-3p, miR-29a/b, miR-30a-5p)
Lineage dependent (miR-106a-5p, miR-302a/b/c/d-3p, miR-372-3p and miR-373-3p; miR-92a-3p, miR-363-3p); miR-363 targets NOTCH1 and PSEN1 |
(submitted, 2018) |
Notes: lipid transfection (LT), electroporation (EP), synthetic mimics (SM), flow cytometry (FC), spectrophotometry (SP), fluorometry (F), luminometry (L), transmitted light microscopy (TLM), fluorescence microscopy (FM), confocal fluorescence microscopy (CFM), high-content screening (HCS), alkaline phosphatase (AP). # Gain of function effect of introduced miRs could promote pluripotency/reprograming or inhibit Naïve-Primed transition or differentiation (“Pluripotency/Reprograming Effectors” or “Differentiation Inhibitors”); alternatively, miRs could be associated with reduced pluripotency/reprograming, Naïve-Primed transition or differentiation induction (“Pluripotency/Reprograming Repressors” or “Differentiation Inducers,” italicized in the table). Effect on reprograming efficiency (when evaluated) is mentioned with the specific miRNA (or anti-miR/inhibitor) tested in parenthesis. $ Reprograming. * Differentiation. @ Naïve-Primed Transition