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dgcr8 knockout dgcr8 mouse escs  (Novus Biologicals)


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    Novus Biologicals dgcr8 knockout dgcr8 mouse escs
    ( a ) (left) Susceptibility (TCID 50 /ml) of miRNA deficient cells ( <t>Dgcr8</t> -/- , Dicer -/- ) and wild-type parental cells ( Dgcr8 +/+ (ESC1), Dicer +/+ (ESC2)) to TMEV infection, higher values represent higher susceptibility (n = 4, p-value<0.05, t-test). (right) Quantification of Influenza A replication after infection of the same cell lines, data show the average (n = 3)±s.d. (*) p-value<0.05 by t-test. ( b ) Quantification of Ifnb1 expression of ESCs lacking Dgcr8 or Dicer to stimulation with poly(I:C) and G 3 -YSD. Data show average (n = 3)±s.d., normalized to mock, (*) p-value<0.05 by t-test. ( c ) Quantification of TMEV vRNA upon JAK1/2 inhibition by Ruxolitinib treatment. Data show the average (n = 3)±s.d. ( d ) qRT-PCR analyses of ISGs expression after stimulation of wild-type and miRNA-deficient ESCs with IFN-β. Data show average (n = 3)±s.d., normalized to mock treated cells ( e, f ) Quantification of TMEV replication after infection in Dgcr8 ( e ) and Dicer ( f ) parental (+/+), deficient (-/-) and rescued (resc) cell lines. Data are normalized to miRNA-deficient cell lines susceptibility. Data show the average (n = 3)±s.d (*) p-value<0.05 by t-test. Northern blots for three stem-cell specific miRNAs, as control for knock-out and rescue of Dgcr8 and Dicer , are shown at the right of each panel.
    Dgcr8 Knockout Dgcr8 Mouse Escs, supplied by Novus Biologicals, used in various techniques. Bioz Stars score: 90/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/nba1+19349/pmc06529217-146-0-11?v=Novus+Biologicals
    Average 90 stars, based on 14 article reviews
    dgcr8 knockout dgcr8 mouse escs - by Bioz Stars, 2026-07
    90/100 stars

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    1) Product Images from "MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response"

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    Journal: eLife

    doi: 10.7554/eLife.44171

    ( a ) (left) Susceptibility (TCID 50 /ml) of miRNA deficient cells ( Dgcr8 -/- , Dicer -/- ) and wild-type parental cells ( Dgcr8 +/+ (ESC1), Dicer +/+ (ESC2)) to TMEV infection, higher values represent higher susceptibility (n = 4, p-value<0.05, t-test). (right) Quantification of Influenza A replication after infection of the same cell lines, data show the average (n = 3)±s.d. (*) p-value<0.05 by t-test. ( b ) Quantification of Ifnb1 expression of ESCs lacking Dgcr8 or Dicer to stimulation with poly(I:C) and G 3 -YSD. Data show average (n = 3)±s.d., normalized to mock, (*) p-value<0.05 by t-test. ( c ) Quantification of TMEV vRNA upon JAK1/2 inhibition by Ruxolitinib treatment. Data show the average (n = 3)±s.d. ( d ) qRT-PCR analyses of ISGs expression after stimulation of wild-type and miRNA-deficient ESCs with IFN-β. Data show average (n = 3)±s.d., normalized to mock treated cells ( e, f ) Quantification of TMEV replication after infection in Dgcr8 ( e ) and Dicer ( f ) parental (+/+), deficient (-/-) and rescued (resc) cell lines. Data are normalized to miRNA-deficient cell lines susceptibility. Data show the average (n = 3)±s.d (*) p-value<0.05 by t-test. Northern blots for three stem-cell specific miRNAs, as control for knock-out and rescue of Dgcr8 and Dicer , are shown at the right of each panel.
    Figure Legend Snippet: ( a ) (left) Susceptibility (TCID 50 /ml) of miRNA deficient cells ( Dgcr8 -/- , Dicer -/- ) and wild-type parental cells ( Dgcr8 +/+ (ESC1), Dicer +/+ (ESC2)) to TMEV infection, higher values represent higher susceptibility (n = 4, p-value<0.05, t-test). (right) Quantification of Influenza A replication after infection of the same cell lines, data show the average (n = 3)±s.d. (*) p-value<0.05 by t-test. ( b ) Quantification of Ifnb1 expression of ESCs lacking Dgcr8 or Dicer to stimulation with poly(I:C) and G 3 -YSD. Data show average (n = 3)±s.d., normalized to mock, (*) p-value<0.05 by t-test. ( c ) Quantification of TMEV vRNA upon JAK1/2 inhibition by Ruxolitinib treatment. Data show the average (n = 3)±s.d. ( d ) qRT-PCR analyses of ISGs expression after stimulation of wild-type and miRNA-deficient ESCs with IFN-β. Data show average (n = 3)±s.d., normalized to mock treated cells ( e, f ) Quantification of TMEV replication after infection in Dgcr8 ( e ) and Dicer ( f ) parental (+/+), deficient (-/-) and rescued (resc) cell lines. Data are normalized to miRNA-deficient cell lines susceptibility. Data show the average (n = 3)±s.d (*) p-value<0.05 by t-test. Northern blots for three stem-cell specific miRNAs, as control for knock-out and rescue of Dgcr8 and Dicer , are shown at the right of each panel.

    Techniques Used: Infection, Expressing, Inhibition, Quantitative RT-PCR, Northern Blot, Control, Knock-Out

    ( a ) RT-PCR of poly(I:C) activated ESCs. Only Dgcr8 -/- (lane 6, left panel) and Dicer -/- ( lane 5 and 6, right panel) express detectable Ifnb1 transcript (top panels). Actb amplification serves as a loading control. ( b ) Ifnb mRNA quantification by qRT-PCR after increasing amounts of poly(I:C) in Dgcr8 -/- and Dicer -/- mESCs. Data show the average (n = 3)±s.d., normalized to mock, (*) p-value<0.05 by t-test. ( c ) IFN-β ELISA analyses from mock and poly(I:C) stimulated wild-type and miRNA-deficient ESCs. Data show the average (n = 3)±s.d., (*) p-value<0.05 by two-tailed T test ( d ) TMEV vRNA quantification in wild-type and miRNA-deficient cell lines after mock or Ruxolitinib treatment (JAK1/2 inhibitor). Data show the average (n = 3)±s.d., (*) p value < 0.05 by Student’s t-test, n.s. non-significant ( e ) TCID50 quantification after TMEV infection of Dgcr8 (left) and Dicer (right) rescued cell lines.
    Figure Legend Snippet: ( a ) RT-PCR of poly(I:C) activated ESCs. Only Dgcr8 -/- (lane 6, left panel) and Dicer -/- ( lane 5 and 6, right panel) express detectable Ifnb1 transcript (top panels). Actb amplification serves as a loading control. ( b ) Ifnb mRNA quantification by qRT-PCR after increasing amounts of poly(I:C) in Dgcr8 -/- and Dicer -/- mESCs. Data show the average (n = 3)±s.d., normalized to mock, (*) p-value<0.05 by t-test. ( c ) IFN-β ELISA analyses from mock and poly(I:C) stimulated wild-type and miRNA-deficient ESCs. Data show the average (n = 3)±s.d., (*) p-value<0.05 by two-tailed T test ( d ) TMEV vRNA quantification in wild-type and miRNA-deficient cell lines after mock or Ruxolitinib treatment (JAK1/2 inhibitor). Data show the average (n = 3)±s.d., (*) p value < 0.05 by Student’s t-test, n.s. non-significant ( e ) TCID50 quantification after TMEV infection of Dgcr8 (left) and Dicer (right) rescued cell lines.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Amplification, Control, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Infection

    ( a, b ) Susceptibility of Dgcr8 -/- , Dicer -/- and parental cells to TMEV infection after inhibition of IRF3 (BX795) ( a ) and Nf-κB (BMS345541) ( b ), normalized to mock-treated cells. ( c ) Heat map of significantly differentially expressed proteins (p<0.05) in the absence ( Dgcr8 -/- ) or presence ( Dgcr8 resc ) of miRNAs identified by STRING analysis. ( d ) Western blot analysis of MAVS expression in miRNA-deficient cells ( Dgcr8 -/- and Dicer -/- , lanes 2 and 5), wild-type counterparts ( Dgcr8 +/+ and Dicer +/+ , lanes 1 and 4) and respective rescued ESCs lines ( Dgcr8 resc and Dicer resc , lanes 3 and 6). MAVS quantification normalized to Tubulin and relative to wild-type levels is shown at the top of the panel. 10.7554/eLife.44171.010 Figure 3—source data 1. Source data of mass spectrometry results.
    Figure Legend Snippet: ( a, b ) Susceptibility of Dgcr8 -/- , Dicer -/- and parental cells to TMEV infection after inhibition of IRF3 (BX795) ( a ) and Nf-κB (BMS345541) ( b ), normalized to mock-treated cells. ( c ) Heat map of significantly differentially expressed proteins (p<0.05) in the absence ( Dgcr8 -/- ) or presence ( Dgcr8 resc ) of miRNAs identified by STRING analysis. ( d ) Western blot analysis of MAVS expression in miRNA-deficient cells ( Dgcr8 -/- and Dicer -/- , lanes 2 and 5), wild-type counterparts ( Dgcr8 +/+ and Dicer +/+ , lanes 1 and 4) and respective rescued ESCs lines ( Dgcr8 resc and Dicer resc , lanes 3 and 6). MAVS quantification normalized to Tubulin and relative to wild-type levels is shown at the top of the panel. 10.7554/eLife.44171.010 Figure 3—source data 1. Source data of mass spectrometry results.

    Techniques Used: Infection, Inhibition, Western Blot, Expressing, Mass Spectrometry

    ( a ) Treatment of mESCs with inhibitors of IRF3 and NF-κB activation improves viral replication to a much greater extend in the Dgcr8 -/- and Dicer -/- cells compared to the parental cells, suggesting that the inhibitory effect that miRNAs have on the IFN response is upstream of these two transcription factors. ( b ) Western blot analyses of phospho-IRF3 upon stimulation of Dgcr8 -/- ESCs with poly(I:C) in the presence or absence of BX759 (inhibitor IRF3 phosphorylation). Tubulin serves as a loading control ( c ) Quantification of Tnf mRNA upon poly(I:C) stimulation in Dgcr8 +/+ and Dgcr8 -/- in the presence or absence of BMS345541 (inhibitor of NFkB activation). Data show the average (n = 2) normalized to Gapdh ( d ) Quantification of oxidative phosphorylation activity by Rhodamine 123 uptake assay in Dgcr8 (left) and Dicer (right) cell lines. ( e ) qRT-PCR quantification of Mavs mRNA. Mavs expression is higher in the absence of miRNAs ( Dgcr8 -/- and Dicer -/- ), when compared to parental cell lines ( Dgcr8 +/+ and Dicer +/+ ). Data show the average (n = 3)±s.d, (*) denotes p<0.05, t-test. ( f ) Western blot analyses for MDA5, RIG-I and PKR for wild-type cell lines (+/+), Dgcr8 and Dicer KO (-/-), and the respective rescued cell lines (resc). Quantification of the signal normalized to Tubulin and relative to wild-type is shown at the top of each panel.
    Figure Legend Snippet: ( a ) Treatment of mESCs with inhibitors of IRF3 and NF-κB activation improves viral replication to a much greater extend in the Dgcr8 -/- and Dicer -/- cells compared to the parental cells, suggesting that the inhibitory effect that miRNAs have on the IFN response is upstream of these two transcription factors. ( b ) Western blot analyses of phospho-IRF3 upon stimulation of Dgcr8 -/- ESCs with poly(I:C) in the presence or absence of BX759 (inhibitor IRF3 phosphorylation). Tubulin serves as a loading control ( c ) Quantification of Tnf mRNA upon poly(I:C) stimulation in Dgcr8 +/+ and Dgcr8 -/- in the presence or absence of BMS345541 (inhibitor of NFkB activation). Data show the average (n = 2) normalized to Gapdh ( d ) Quantification of oxidative phosphorylation activity by Rhodamine 123 uptake assay in Dgcr8 (left) and Dicer (right) cell lines. ( e ) qRT-PCR quantification of Mavs mRNA. Mavs expression is higher in the absence of miRNAs ( Dgcr8 -/- and Dicer -/- ), when compared to parental cell lines ( Dgcr8 +/+ and Dicer +/+ ). Data show the average (n = 3)±s.d, (*) denotes p<0.05, t-test. ( f ) Western blot analyses for MDA5, RIG-I and PKR for wild-type cell lines (+/+), Dgcr8 and Dicer KO (-/-), and the respective rescued cell lines (resc). Quantification of the signal normalized to Tubulin and relative to wild-type is shown at the top of each panel.

    Techniques Used: Activation Assay, Western Blot, Phospho-proteomics, Control, Activity Assay, Quantitative RT-PCR, Expressing

    ( a ) Dual luciferase assay with Mavs , Rig-I and Mda5 3’UTRs in miRNA-deficient cells lines ( Dgcr8 -/- and Dicer -/- ). Data show the average (n = 3)±s.d normalized to Renilla and relative to the parental lines, (*) p-value<0.05 by t-test ( b ) Western blot of cell line overexpressing MAVS lacking the 3’UTR in Dgcr8 +/+ cells (lane 3). MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( c ) Susceptibility (TCID 50 /ml) of same cells lines as in ( b ) to TMEV infection (left panel) and quantification of viral RNA after TMEV infections in the same cell lines (right panel). Data show the average (n = 5)±s.d. (*) p-value<0.05 by t-test ( d ) Ifnb mRNA expression after poly(I:C) transfection of the same cell lines as in ( b ), average is represented (n = 3)±s.d, normalized to Dgcr8 +/+ cell line, (*) p-value<0.05 by t-test.
    Figure Legend Snippet: ( a ) Dual luciferase assay with Mavs , Rig-I and Mda5 3’UTRs in miRNA-deficient cells lines ( Dgcr8 -/- and Dicer -/- ). Data show the average (n = 3)±s.d normalized to Renilla and relative to the parental lines, (*) p-value<0.05 by t-test ( b ) Western blot of cell line overexpressing MAVS lacking the 3’UTR in Dgcr8 +/+ cells (lane 3). MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( c ) Susceptibility (TCID 50 /ml) of same cells lines as in ( b ) to TMEV infection (left panel) and quantification of viral RNA after TMEV infections in the same cell lines (right panel). Data show the average (n = 5)±s.d. (*) p-value<0.05 by t-test ( d ) Ifnb mRNA expression after poly(I:C) transfection of the same cell lines as in ( b ), average is represented (n = 3)±s.d, normalized to Dgcr8 +/+ cell line, (*) p-value<0.05 by t-test.

    Techniques Used: Luciferase, Western Blot, Infection, Expressing, Transfection

    ( a ) Transfection of miRNA mimics miR-125a-5p, miR-125b-5p, miR-185–5p and miR-673–5p in Dgcr8 -/- cells followed by MAVS western blot. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( b ) Quantification of TMEV replication by qRT-PCR in the same cell lines as in ( a ) (n = 3) ( c ) Western blot analysis of MAVS expression in Dgcr8 +/+ cells transfected with antagomirs against miR-125a-5p, miR-125b-5p and miR-673–5p. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( d ) Western blot analysis of MAVS expression in miR-673 -/- cell lines. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( e ) Quantification of TMEV vRNA in miR-673 CRISPR knock-out cell lines in a Dgcr8 +/+ background. Data show the average (n = 3)±s.d (*) p-value<0.05 by t-test. ( f ) Quantification of TMEV vRNA shown as fold-change compared to mock upon JAK1/2 inhibition by Ruxolitinib treatment. Data show the average (n = 3)±s.d ( g ) qRT-PCR quantification of mmu-miR-673–5p expression after retinoic acid (RA) differentiation of ESC. Data show the average (n = 2)±s.e.m, normalized to U6 snRNA, (*) p-val <0.05 by t-test.
    Figure Legend Snippet: ( a ) Transfection of miRNA mimics miR-125a-5p, miR-125b-5p, miR-185–5p and miR-673–5p in Dgcr8 -/- cells followed by MAVS western blot. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( b ) Quantification of TMEV replication by qRT-PCR in the same cell lines as in ( a ) (n = 3) ( c ) Western blot analysis of MAVS expression in Dgcr8 +/+ cells transfected with antagomirs against miR-125a-5p, miR-125b-5p and miR-673–5p. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( d ) Western blot analysis of MAVS expression in miR-673 -/- cell lines. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( e ) Quantification of TMEV vRNA in miR-673 CRISPR knock-out cell lines in a Dgcr8 +/+ background. Data show the average (n = 3)±s.d (*) p-value<0.05 by t-test. ( f ) Quantification of TMEV vRNA shown as fold-change compared to mock upon JAK1/2 inhibition by Ruxolitinib treatment. Data show the average (n = 3)±s.d ( g ) qRT-PCR quantification of mmu-miR-673–5p expression after retinoic acid (RA) differentiation of ESC. Data show the average (n = 2)±s.e.m, normalized to U6 snRNA, (*) p-val <0.05 by t-test.

    Techniques Used: Transfection, Western Blot, Quantitative RT-PCR, Expressing, CRISPR, Knock-Out, Inhibition

    ( a ) Dgcr8 -/- cells were transfected with miRNA mimics for 24 hr, followed by RNA extraction and Mavs mRNA quantification by qRT-PCR. Transfection of all miRNA mimics result in (non-significant) decrease of Mavs levels. ( b ) Dgcr8 -/- mESCs were first transfected with mimics, as in ( a ) followed by TMEV infections 36 hr later, and measurement of the TCID50. All mimics except for mir-185-5p resulted in higher TCID50 values compared to mock transfected cells, suggesting that in mESC cells targeting Mavs with these miRNAs leads to a higher susceptibility to viral infection.
    Figure Legend Snippet: ( a ) Dgcr8 -/- cells were transfected with miRNA mimics for 24 hr, followed by RNA extraction and Mavs mRNA quantification by qRT-PCR. Transfection of all miRNA mimics result in (non-significant) decrease of Mavs levels. ( b ) Dgcr8 -/- mESCs were first transfected with mimics, as in ( a ) followed by TMEV infections 36 hr later, and measurement of the TCID50. All mimics except for mir-185-5p resulted in higher TCID50 values compared to mock transfected cells, suggesting that in mESC cells targeting Mavs with these miRNAs leads to a higher susceptibility to viral infection.

    Techniques Used: Transfection, RNA Extraction, Quantitative RT-PCR, Infection

    ( a ) Genomic sequencing of miR-673 locus contained in chromosome 12. In WT, the position of the two miRNAs encoded by the locus, miR-673–5p and miR-673–3p are indicated by yellow boxes. For miR-673 -/- clones (1,2 and 3) both sequenced alleles are shown with the indicated deletions induced by CRISPR. ( b ) Quantification of mmu-miR-673–5p abundance by qRT-PCR. Data show the average (n = 3)±s.d, normalized to U6 snRNA. (n.d, not detected) ( c ) Quantification of Mavs mRNA levels by RT-qPCR. Data show the average of at least (n = 3)±s.d, normalized to Actb , and relative to Dgcr8 +/+ .
    Figure Legend Snippet: ( a ) Genomic sequencing of miR-673 locus contained in chromosome 12. In WT, the position of the two miRNAs encoded by the locus, miR-673–5p and miR-673–3p are indicated by yellow boxes. For miR-673 -/- clones (1,2 and 3) both sequenced alleles are shown with the indicated deletions induced by CRISPR. ( b ) Quantification of mmu-miR-673–5p abundance by qRT-PCR. Data show the average (n = 3)±s.d, normalized to U6 snRNA. (n.d, not detected) ( c ) Quantification of Mavs mRNA levels by RT-qPCR. Data show the average of at least (n = 3)±s.d, normalized to Actb , and relative to Dgcr8 +/+ .

    Techniques Used: Genomic Sequencing, Clone Assay, CRISPR, Quantitative RT-PCR

    ( a ) TMEV vRNA quantification in wild-type and miRNA-deficient cell lines after mock or Ruxolitinib treatment (JAK1/2 inhibitor). Data show the average (n = 3)±s.d., (*) p value < 0.05 by Student’s t-test, n.s. non-significant ( b ) Growth curves of ESCs. Data show the average (n = 3)±s.d. ( c ) No obvious morphological changes were observed between wild-type ( Dgcr8 +/+ ) and miR-673-deficient ESCs ( d ) Quantification of Nanog and Pouf5f1 pluripotent markers upon knocking out miR-673 by CRISPR in comparison to wild-type ESCs, data show the average (n = 3)±s.e.m, (n.s.) non-significant by t-test. NIH3T3 serves as a negative control for pluripotent maker expression ( e ) Quantification of Nanog and Pouf51 expression after reintroduction of miR-673 mimics and controls for 24 or 48 hr. Data show the average (n = 3)±s.e.m. (n.s.) non-significant by t-test.
    Figure Legend Snippet: ( a ) TMEV vRNA quantification in wild-type and miRNA-deficient cell lines after mock or Ruxolitinib treatment (JAK1/2 inhibitor). Data show the average (n = 3)±s.d., (*) p value < 0.05 by Student’s t-test, n.s. non-significant ( b ) Growth curves of ESCs. Data show the average (n = 3)±s.d. ( c ) No obvious morphological changes were observed between wild-type ( Dgcr8 +/+ ) and miR-673-deficient ESCs ( d ) Quantification of Nanog and Pouf5f1 pluripotent markers upon knocking out miR-673 by CRISPR in comparison to wild-type ESCs, data show the average (n = 3)±s.e.m, (n.s.) non-significant by t-test. NIH3T3 serves as a negative control for pluripotent maker expression ( e ) Quantification of Nanog and Pouf51 expression after reintroduction of miR-673 mimics and controls for 24 or 48 hr. Data show the average (n = 3)±s.e.m. (n.s.) non-significant by t-test.

    Techniques Used: CRISPR, Comparison, Negative Control, Expressing



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    cell line mouse dgcr8 ko embryonic stem cell pmid - by Bioz Stars, 2026-07
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    nba1 19349 dgcr8 ko mouse embryonic stem cell line  (Novus Biologicals)
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    Novus Biologicals nba1 19349 dgcr8 ko mouse embryonic stem cell line
    ( A–D ) mRNA stability or translation level changes of ESCC miRNA targets versus all mRNAs. The p value was calculated with Mann-Whitney test. A/B) mRNA stability changes in <t>Dgcr8</t> KO ( A ) or Ddx6 KO ( B ) cells. n = 3 for wild-type, n = 4 for Ddx6 KO (2 replicates of each Ddx6 KO line), n = 3 for Dgcr8 KO. C/D) Translation level changes in Dgcr8 KO ( C ) or Ddx6 KO ( D ) cells. n = 3 for each genotype. ( E ) Translation level changes of individual ESCC miRNA targets in Dgcr8 KO and Ddx6 KO ESCs. See also .
    Nba1 19349 Dgcr8 Ko Mouse Embryonic Stem Cell Line, supplied by Novus Biologicals, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ( a ) (left) Susceptibility (TCID 50 /ml) of miRNA deficient cells ( Dgcr8 -/- , Dicer -/- ) and wild-type parental cells ( Dgcr8 +/+ (ESC1), Dicer +/+ (ESC2)) to TMEV infection, higher values represent higher susceptibility (n = 4, p-value<0.05, t-test). (right) Quantification of Influenza A replication after infection of the same cell lines, data show the average (n = 3)±s.d. (*) p-value<0.05 by t-test. ( b ) Quantification of Ifnb1 expression of ESCs lacking Dgcr8 or Dicer to stimulation with poly(I:C) and G 3 -YSD. Data show average (n = 3)±s.d., normalized to mock, (*) p-value<0.05 by t-test. ( c ) Quantification of TMEV vRNA upon JAK1/2 inhibition by Ruxolitinib treatment. Data show the average (n = 3)±s.d. ( d ) qRT-PCR analyses of ISGs expression after stimulation of wild-type and miRNA-deficient ESCs with IFN-β. Data show average (n = 3)±s.d., normalized to mock treated cells ( e, f ) Quantification of TMEV replication after infection in Dgcr8 ( e ) and Dicer ( f ) parental (+/+), deficient (-/-) and rescued (resc) cell lines. Data are normalized to miRNA-deficient cell lines susceptibility. Data show the average (n = 3)±s.d (*) p-value<0.05 by t-test. Northern blots for three stem-cell specific miRNAs, as control for knock-out and rescue of Dgcr8 and Dicer , are shown at the right of each panel.

    Journal: eLife

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    doi: 10.7554/eLife.44171

    Figure Lengend Snippet: ( a ) (left) Susceptibility (TCID 50 /ml) of miRNA deficient cells ( Dgcr8 -/- , Dicer -/- ) and wild-type parental cells ( Dgcr8 +/+ (ESC1), Dicer +/+ (ESC2)) to TMEV infection, higher values represent higher susceptibility (n = 4, p-value<0.05, t-test). (right) Quantification of Influenza A replication after infection of the same cell lines, data show the average (n = 3)±s.d. (*) p-value<0.05 by t-test. ( b ) Quantification of Ifnb1 expression of ESCs lacking Dgcr8 or Dicer to stimulation with poly(I:C) and G 3 -YSD. Data show average (n = 3)±s.d., normalized to mock, (*) p-value<0.05 by t-test. ( c ) Quantification of TMEV vRNA upon JAK1/2 inhibition by Ruxolitinib treatment. Data show the average (n = 3)±s.d. ( d ) qRT-PCR analyses of ISGs expression after stimulation of wild-type and miRNA-deficient ESCs with IFN-β. Data show average (n = 3)±s.d., normalized to mock treated cells ( e, f ) Quantification of TMEV replication after infection in Dgcr8 ( e ) and Dicer ( f ) parental (+/+), deficient (-/-) and rescued (resc) cell lines. Data are normalized to miRNA-deficient cell lines susceptibility. Data show the average (n = 3)±s.d (*) p-value<0.05 by t-test. Northern blots for three stem-cell specific miRNAs, as control for knock-out and rescue of Dgcr8 and Dicer , are shown at the right of each panel.

    Article Snippet: Dgcr8 knockout ( Dgcr8 -/- ) mouse ESCs were purchased from Novus Biologicals (NBA1-19349) and the parental strain, v6.5 ( Dgcr8 +/+ , also named in the text ESC1) from ThermoFisher (MES1402).

    Techniques: Infection, Expressing, Inhibition, Quantitative RT-PCR, Northern Blot, Control, Knock-Out

    ( a ) RT-PCR of poly(I:C) activated ESCs. Only Dgcr8 -/- (lane 6, left panel) and Dicer -/- ( lane 5 and 6, right panel) express detectable Ifnb1 transcript (top panels). Actb amplification serves as a loading control. ( b ) Ifnb mRNA quantification by qRT-PCR after increasing amounts of poly(I:C) in Dgcr8 -/- and Dicer -/- mESCs. Data show the average (n = 3)±s.d., normalized to mock, (*) p-value<0.05 by t-test. ( c ) IFN-β ELISA analyses from mock and poly(I:C) stimulated wild-type and miRNA-deficient ESCs. Data show the average (n = 3)±s.d., (*) p-value<0.05 by two-tailed T test ( d ) TMEV vRNA quantification in wild-type and miRNA-deficient cell lines after mock or Ruxolitinib treatment (JAK1/2 inhibitor). Data show the average (n = 3)±s.d., (*) p value < 0.05 by Student’s t-test, n.s. non-significant ( e ) TCID50 quantification after TMEV infection of Dgcr8 (left) and Dicer (right) rescued cell lines.

    Journal: eLife

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    doi: 10.7554/eLife.44171

    Figure Lengend Snippet: ( a ) RT-PCR of poly(I:C) activated ESCs. Only Dgcr8 -/- (lane 6, left panel) and Dicer -/- ( lane 5 and 6, right panel) express detectable Ifnb1 transcript (top panels). Actb amplification serves as a loading control. ( b ) Ifnb mRNA quantification by qRT-PCR after increasing amounts of poly(I:C) in Dgcr8 -/- and Dicer -/- mESCs. Data show the average (n = 3)±s.d., normalized to mock, (*) p-value<0.05 by t-test. ( c ) IFN-β ELISA analyses from mock and poly(I:C) stimulated wild-type and miRNA-deficient ESCs. Data show the average (n = 3)±s.d., (*) p-value<0.05 by two-tailed T test ( d ) TMEV vRNA quantification in wild-type and miRNA-deficient cell lines after mock or Ruxolitinib treatment (JAK1/2 inhibitor). Data show the average (n = 3)±s.d., (*) p value < 0.05 by Student’s t-test, n.s. non-significant ( e ) TCID50 quantification after TMEV infection of Dgcr8 (left) and Dicer (right) rescued cell lines.

    Article Snippet: Dgcr8 knockout ( Dgcr8 -/- ) mouse ESCs were purchased from Novus Biologicals (NBA1-19349) and the parental strain, v6.5 ( Dgcr8 +/+ , also named in the text ESC1) from ThermoFisher (MES1402).

    Techniques: Reverse Transcription Polymerase Chain Reaction, Amplification, Control, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Infection

    ( a, b ) Susceptibility of Dgcr8 -/- , Dicer -/- and parental cells to TMEV infection after inhibition of IRF3 (BX795) ( a ) and Nf-κB (BMS345541) ( b ), normalized to mock-treated cells. ( c ) Heat map of significantly differentially expressed proteins (p<0.05) in the absence ( Dgcr8 -/- ) or presence ( Dgcr8 resc ) of miRNAs identified by STRING analysis. ( d ) Western blot analysis of MAVS expression in miRNA-deficient cells ( Dgcr8 -/- and Dicer -/- , lanes 2 and 5), wild-type counterparts ( Dgcr8 +/+ and Dicer +/+ , lanes 1 and 4) and respective rescued ESCs lines ( Dgcr8 resc and Dicer resc , lanes 3 and 6). MAVS quantification normalized to Tubulin and relative to wild-type levels is shown at the top of the panel. 10.7554/eLife.44171.010 Figure 3—source data 1. Source data of mass spectrometry results.

    Journal: eLife

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    doi: 10.7554/eLife.44171

    Figure Lengend Snippet: ( a, b ) Susceptibility of Dgcr8 -/- , Dicer -/- and parental cells to TMEV infection after inhibition of IRF3 (BX795) ( a ) and Nf-κB (BMS345541) ( b ), normalized to mock-treated cells. ( c ) Heat map of significantly differentially expressed proteins (p<0.05) in the absence ( Dgcr8 -/- ) or presence ( Dgcr8 resc ) of miRNAs identified by STRING analysis. ( d ) Western blot analysis of MAVS expression in miRNA-deficient cells ( Dgcr8 -/- and Dicer -/- , lanes 2 and 5), wild-type counterparts ( Dgcr8 +/+ and Dicer +/+ , lanes 1 and 4) and respective rescued ESCs lines ( Dgcr8 resc and Dicer resc , lanes 3 and 6). MAVS quantification normalized to Tubulin and relative to wild-type levels is shown at the top of the panel. 10.7554/eLife.44171.010 Figure 3—source data 1. Source data of mass spectrometry results.

    Article Snippet: Dgcr8 knockout ( Dgcr8 -/- ) mouse ESCs were purchased from Novus Biologicals (NBA1-19349) and the parental strain, v6.5 ( Dgcr8 +/+ , also named in the text ESC1) from ThermoFisher (MES1402).

    Techniques: Infection, Inhibition, Western Blot, Expressing, Mass Spectrometry

    ( a ) Treatment of mESCs with inhibitors of IRF3 and NF-κB activation improves viral replication to a much greater extend in the Dgcr8 -/- and Dicer -/- cells compared to the parental cells, suggesting that the inhibitory effect that miRNAs have on the IFN response is upstream of these two transcription factors. ( b ) Western blot analyses of phospho-IRF3 upon stimulation of Dgcr8 -/- ESCs with poly(I:C) in the presence or absence of BX759 (inhibitor IRF3 phosphorylation). Tubulin serves as a loading control ( c ) Quantification of Tnf mRNA upon poly(I:C) stimulation in Dgcr8 +/+ and Dgcr8 -/- in the presence or absence of BMS345541 (inhibitor of NFkB activation). Data show the average (n = 2) normalized to Gapdh ( d ) Quantification of oxidative phosphorylation activity by Rhodamine 123 uptake assay in Dgcr8 (left) and Dicer (right) cell lines. ( e ) qRT-PCR quantification of Mavs mRNA. Mavs expression is higher in the absence of miRNAs ( Dgcr8 -/- and Dicer -/- ), when compared to parental cell lines ( Dgcr8 +/+ and Dicer +/+ ). Data show the average (n = 3)±s.d, (*) denotes p<0.05, t-test. ( f ) Western blot analyses for MDA5, RIG-I and PKR for wild-type cell lines (+/+), Dgcr8 and Dicer KO (-/-), and the respective rescued cell lines (resc). Quantification of the signal normalized to Tubulin and relative to wild-type is shown at the top of each panel.

    Journal: eLife

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    doi: 10.7554/eLife.44171

    Figure Lengend Snippet: ( a ) Treatment of mESCs with inhibitors of IRF3 and NF-κB activation improves viral replication to a much greater extend in the Dgcr8 -/- and Dicer -/- cells compared to the parental cells, suggesting that the inhibitory effect that miRNAs have on the IFN response is upstream of these two transcription factors. ( b ) Western blot analyses of phospho-IRF3 upon stimulation of Dgcr8 -/- ESCs with poly(I:C) in the presence or absence of BX759 (inhibitor IRF3 phosphorylation). Tubulin serves as a loading control ( c ) Quantification of Tnf mRNA upon poly(I:C) stimulation in Dgcr8 +/+ and Dgcr8 -/- in the presence or absence of BMS345541 (inhibitor of NFkB activation). Data show the average (n = 2) normalized to Gapdh ( d ) Quantification of oxidative phosphorylation activity by Rhodamine 123 uptake assay in Dgcr8 (left) and Dicer (right) cell lines. ( e ) qRT-PCR quantification of Mavs mRNA. Mavs expression is higher in the absence of miRNAs ( Dgcr8 -/- and Dicer -/- ), when compared to parental cell lines ( Dgcr8 +/+ and Dicer +/+ ). Data show the average (n = 3)±s.d, (*) denotes p<0.05, t-test. ( f ) Western blot analyses for MDA5, RIG-I and PKR for wild-type cell lines (+/+), Dgcr8 and Dicer KO (-/-), and the respective rescued cell lines (resc). Quantification of the signal normalized to Tubulin and relative to wild-type is shown at the top of each panel.

    Article Snippet: Dgcr8 knockout ( Dgcr8 -/- ) mouse ESCs were purchased from Novus Biologicals (NBA1-19349) and the parental strain, v6.5 ( Dgcr8 +/+ , also named in the text ESC1) from ThermoFisher (MES1402).

    Techniques: Activation Assay, Western Blot, Phospho-proteomics, Control, Activity Assay, Quantitative RT-PCR, Expressing

    ( a ) Dual luciferase assay with Mavs , Rig-I and Mda5 3’UTRs in miRNA-deficient cells lines ( Dgcr8 -/- and Dicer -/- ). Data show the average (n = 3)±s.d normalized to Renilla and relative to the parental lines, (*) p-value<0.05 by t-test ( b ) Western blot of cell line overexpressing MAVS lacking the 3’UTR in Dgcr8 +/+ cells (lane 3). MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( c ) Susceptibility (TCID 50 /ml) of same cells lines as in ( b ) to TMEV infection (left panel) and quantification of viral RNA after TMEV infections in the same cell lines (right panel). Data show the average (n = 5)±s.d. (*) p-value<0.05 by t-test ( d ) Ifnb mRNA expression after poly(I:C) transfection of the same cell lines as in ( b ), average is represented (n = 3)±s.d, normalized to Dgcr8 +/+ cell line, (*) p-value<0.05 by t-test.

    Journal: eLife

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    doi: 10.7554/eLife.44171

    Figure Lengend Snippet: ( a ) Dual luciferase assay with Mavs , Rig-I and Mda5 3’UTRs in miRNA-deficient cells lines ( Dgcr8 -/- and Dicer -/- ). Data show the average (n = 3)±s.d normalized to Renilla and relative to the parental lines, (*) p-value<0.05 by t-test ( b ) Western blot of cell line overexpressing MAVS lacking the 3’UTR in Dgcr8 +/+ cells (lane 3). MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( c ) Susceptibility (TCID 50 /ml) of same cells lines as in ( b ) to TMEV infection (left panel) and quantification of viral RNA after TMEV infections in the same cell lines (right panel). Data show the average (n = 5)±s.d. (*) p-value<0.05 by t-test ( d ) Ifnb mRNA expression after poly(I:C) transfection of the same cell lines as in ( b ), average is represented (n = 3)±s.d, normalized to Dgcr8 +/+ cell line, (*) p-value<0.05 by t-test.

    Article Snippet: Dgcr8 knockout ( Dgcr8 -/- ) mouse ESCs were purchased from Novus Biologicals (NBA1-19349) and the parental strain, v6.5 ( Dgcr8 +/+ , also named in the text ESC1) from ThermoFisher (MES1402).

    Techniques: Luciferase, Western Blot, Infection, Expressing, Transfection

    ( a ) Transfection of miRNA mimics miR-125a-5p, miR-125b-5p, miR-185–5p and miR-673–5p in Dgcr8 -/- cells followed by MAVS western blot. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( b ) Quantification of TMEV replication by qRT-PCR in the same cell lines as in ( a ) (n = 3) ( c ) Western blot analysis of MAVS expression in Dgcr8 +/+ cells transfected with antagomirs against miR-125a-5p, miR-125b-5p and miR-673–5p. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( d ) Western blot analysis of MAVS expression in miR-673 -/- cell lines. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( e ) Quantification of TMEV vRNA in miR-673 CRISPR knock-out cell lines in a Dgcr8 +/+ background. Data show the average (n = 3)±s.d (*) p-value<0.05 by t-test. ( f ) Quantification of TMEV vRNA shown as fold-change compared to mock upon JAK1/2 inhibition by Ruxolitinib treatment. Data show the average (n = 3)±s.d ( g ) qRT-PCR quantification of mmu-miR-673–5p expression after retinoic acid (RA) differentiation of ESC. Data show the average (n = 2)±s.e.m, normalized to U6 snRNA, (*) p-val <0.05 by t-test.

    Journal: eLife

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    doi: 10.7554/eLife.44171

    Figure Lengend Snippet: ( a ) Transfection of miRNA mimics miR-125a-5p, miR-125b-5p, miR-185–5p and miR-673–5p in Dgcr8 -/- cells followed by MAVS western blot. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( b ) Quantification of TMEV replication by qRT-PCR in the same cell lines as in ( a ) (n = 3) ( c ) Western blot analysis of MAVS expression in Dgcr8 +/+ cells transfected with antagomirs against miR-125a-5p, miR-125b-5p and miR-673–5p. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( d ) Western blot analysis of MAVS expression in miR-673 -/- cell lines. MAVS quantification normalized to Tubulin and relative to wild-type is shown at the top ( e ) Quantification of TMEV vRNA in miR-673 CRISPR knock-out cell lines in a Dgcr8 +/+ background. Data show the average (n = 3)±s.d (*) p-value<0.05 by t-test. ( f ) Quantification of TMEV vRNA shown as fold-change compared to mock upon JAK1/2 inhibition by Ruxolitinib treatment. Data show the average (n = 3)±s.d ( g ) qRT-PCR quantification of mmu-miR-673–5p expression after retinoic acid (RA) differentiation of ESC. Data show the average (n = 2)±s.e.m, normalized to U6 snRNA, (*) p-val <0.05 by t-test.

    Article Snippet: Dgcr8 knockout ( Dgcr8 -/- ) mouse ESCs were purchased from Novus Biologicals (NBA1-19349) and the parental strain, v6.5 ( Dgcr8 +/+ , also named in the text ESC1) from ThermoFisher (MES1402).

    Techniques: Transfection, Western Blot, Quantitative RT-PCR, Expressing, CRISPR, Knock-Out, Inhibition

    ( a ) Dgcr8 -/- cells were transfected with miRNA mimics for 24 hr, followed by RNA extraction and Mavs mRNA quantification by qRT-PCR. Transfection of all miRNA mimics result in (non-significant) decrease of Mavs levels. ( b ) Dgcr8 -/- mESCs were first transfected with mimics, as in ( a ) followed by TMEV infections 36 hr later, and measurement of the TCID50. All mimics except for mir-185-5p resulted in higher TCID50 values compared to mock transfected cells, suggesting that in mESC cells targeting Mavs with these miRNAs leads to a higher susceptibility to viral infection.

    Journal: eLife

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    doi: 10.7554/eLife.44171

    Figure Lengend Snippet: ( a ) Dgcr8 -/- cells were transfected with miRNA mimics for 24 hr, followed by RNA extraction and Mavs mRNA quantification by qRT-PCR. Transfection of all miRNA mimics result in (non-significant) decrease of Mavs levels. ( b ) Dgcr8 -/- mESCs were first transfected with mimics, as in ( a ) followed by TMEV infections 36 hr later, and measurement of the TCID50. All mimics except for mir-185-5p resulted in higher TCID50 values compared to mock transfected cells, suggesting that in mESC cells targeting Mavs with these miRNAs leads to a higher susceptibility to viral infection.

    Article Snippet: Dgcr8 knockout ( Dgcr8 -/- ) mouse ESCs were purchased from Novus Biologicals (NBA1-19349) and the parental strain, v6.5 ( Dgcr8 +/+ , also named in the text ESC1) from ThermoFisher (MES1402).

    Techniques: Transfection, RNA Extraction, Quantitative RT-PCR, Infection

    ( a ) Genomic sequencing of miR-673 locus contained in chromosome 12. In WT, the position of the two miRNAs encoded by the locus, miR-673–5p and miR-673–3p are indicated by yellow boxes. For miR-673 -/- clones (1,2 and 3) both sequenced alleles are shown with the indicated deletions induced by CRISPR. ( b ) Quantification of mmu-miR-673–5p abundance by qRT-PCR. Data show the average (n = 3)±s.d, normalized to U6 snRNA. (n.d, not detected) ( c ) Quantification of Mavs mRNA levels by RT-qPCR. Data show the average of at least (n = 3)±s.d, normalized to Actb , and relative to Dgcr8 +/+ .

    Journal: eLife

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    doi: 10.7554/eLife.44171

    Figure Lengend Snippet: ( a ) Genomic sequencing of miR-673 locus contained in chromosome 12. In WT, the position of the two miRNAs encoded by the locus, miR-673–5p and miR-673–3p are indicated by yellow boxes. For miR-673 -/- clones (1,2 and 3) both sequenced alleles are shown with the indicated deletions induced by CRISPR. ( b ) Quantification of mmu-miR-673–5p abundance by qRT-PCR. Data show the average (n = 3)±s.d, normalized to U6 snRNA. (n.d, not detected) ( c ) Quantification of Mavs mRNA levels by RT-qPCR. Data show the average of at least (n = 3)±s.d, normalized to Actb , and relative to Dgcr8 +/+ .

    Article Snippet: Dgcr8 knockout ( Dgcr8 -/- ) mouse ESCs were purchased from Novus Biologicals (NBA1-19349) and the parental strain, v6.5 ( Dgcr8 +/+ , also named in the text ESC1) from ThermoFisher (MES1402).

    Techniques: Genomic Sequencing, Clone Assay, CRISPR, Quantitative RT-PCR

    ( a ) TMEV vRNA quantification in wild-type and miRNA-deficient cell lines after mock or Ruxolitinib treatment (JAK1/2 inhibitor). Data show the average (n = 3)±s.d., (*) p value < 0.05 by Student’s t-test, n.s. non-significant ( b ) Growth curves of ESCs. Data show the average (n = 3)±s.d. ( c ) No obvious morphological changes were observed between wild-type ( Dgcr8 +/+ ) and miR-673-deficient ESCs ( d ) Quantification of Nanog and Pouf5f1 pluripotent markers upon knocking out miR-673 by CRISPR in comparison to wild-type ESCs, data show the average (n = 3)±s.e.m, (n.s.) non-significant by t-test. NIH3T3 serves as a negative control for pluripotent maker expression ( e ) Quantification of Nanog and Pouf51 expression after reintroduction of miR-673 mimics and controls for 24 or 48 hr. Data show the average (n = 3)±s.e.m. (n.s.) non-significant by t-test.

    Journal: eLife

    Article Title: MicroRNA-deficient mouse embryonic stem cells acquire a functional interferon response

    doi: 10.7554/eLife.44171

    Figure Lengend Snippet: ( a ) TMEV vRNA quantification in wild-type and miRNA-deficient cell lines after mock or Ruxolitinib treatment (JAK1/2 inhibitor). Data show the average (n = 3)±s.d., (*) p value < 0.05 by Student’s t-test, n.s. non-significant ( b ) Growth curves of ESCs. Data show the average (n = 3)±s.d. ( c ) No obvious morphological changes were observed between wild-type ( Dgcr8 +/+ ) and miR-673-deficient ESCs ( d ) Quantification of Nanog and Pouf5f1 pluripotent markers upon knocking out miR-673 by CRISPR in comparison to wild-type ESCs, data show the average (n = 3)±s.e.m, (n.s.) non-significant by t-test. NIH3T3 serves as a negative control for pluripotent maker expression ( e ) Quantification of Nanog and Pouf51 expression after reintroduction of miR-673 mimics and controls for 24 or 48 hr. Data show the average (n = 3)±s.e.m. (n.s.) non-significant by t-test.

    Article Snippet: Dgcr8 knockout ( Dgcr8 -/- ) mouse ESCs were purchased from Novus Biologicals (NBA1-19349) and the parental strain, v6.5 ( Dgcr8 +/+ , also named in the text ESC1) from ThermoFisher (MES1402).

    Techniques: CRISPR, Comparison, Negative Control, Expressing

    ( A–D ) mRNA stability or translation level changes of ESCC miRNA targets versus all mRNAs. The p value was calculated with Mann-Whitney test. A/B) mRNA stability changes in Dgcr8 KO ( A ) or Ddx6 KO ( B ) cells. n = 3 for wild-type, n = 4 for Ddx6 KO (2 replicates of each Ddx6 KO line), n = 3 for Dgcr8 KO. C/D) Translation level changes in Dgcr8 KO ( C ) or Ddx6 KO ( D ) cells. n = 3 for each genotype. ( E ) Translation level changes of individual ESCC miRNA targets in Dgcr8 KO and Ddx6 KO ESCs. See also .

    Journal: eLife

    Article Title: Decoupling the impact of microRNAs on translational repression versus RNA degradation in embryonic stem cells

    doi: 10.7554/eLife.38014

    Figure Lengend Snippet: ( A–D ) mRNA stability or translation level changes of ESCC miRNA targets versus all mRNAs. The p value was calculated with Mann-Whitney test. A/B) mRNA stability changes in Dgcr8 KO ( A ) or Ddx6 KO ( B ) cells. n = 3 for wild-type, n = 4 for Ddx6 KO (2 replicates of each Ddx6 KO line), n = 3 for Dgcr8 KO. C/D) Translation level changes in Dgcr8 KO ( C ) or Ddx6 KO ( D ) cells. n = 3 for each genotype. ( E ) Translation level changes of individual ESCC miRNA targets in Dgcr8 KO and Ddx6 KO ESCs. See also .

    Article Snippet: Cell line (Mouse) , Dgcr8 KO embryonic stem cell , PMID: 17259983; Novus Biologicals , NBA1-19349 , Dgcr8 KO mouse embryonic stem cell line previously generated in the Blelloch lab. Also available commercially from Novus Biologicals..

    Techniques: MANN-WHITNEY

    ( A ) Comparison between mRNA stability changes in Dgcr8 KO versus Ddx6 KO cells. n = 3 for wild-type, n = 4 for Ddx6 KO (2 replicates of each Ddx6 KO line), n = 3 for Dgcr8 KO. ( B ) Comparison between translation level changes in Dgcr8 KO versus Ddx6 KO cells. n = 3 for each genotype. ( C ) Comparison between mRNA changes in Dgcr8 KO versus Ddx6 KO cells. The p value was calculated with correlation significance test. ( D ) Summary schematic comparing Dgcr8 KO cells to Ddx6 KO cells. Dgcr8 KO leads to the loss of both translational repression and mRNA destabilization of miRNA targets, while Ddx6 KO only leads to the loss of translational repression of miRNA targets. mRNA stability is measured as the ratio of mRNA/4sU reads, changes in translation level are measured as the ratio of polysome/monosome reads, protein level changes are not directly measured but are predicted based on mRNA stability and translation level changes. Changes in translation level alone in Ddx6 KO cells produce similar phenotypes and global molecular changes to Dgcr8 KO cells. See also .

    Journal: eLife

    Article Title: Decoupling the impact of microRNAs on translational repression versus RNA degradation in embryonic stem cells

    doi: 10.7554/eLife.38014

    Figure Lengend Snippet: ( A ) Comparison between mRNA stability changes in Dgcr8 KO versus Ddx6 KO cells. n = 3 for wild-type, n = 4 for Ddx6 KO (2 replicates of each Ddx6 KO line), n = 3 for Dgcr8 KO. ( B ) Comparison between translation level changes in Dgcr8 KO versus Ddx6 KO cells. n = 3 for each genotype. ( C ) Comparison between mRNA changes in Dgcr8 KO versus Ddx6 KO cells. The p value was calculated with correlation significance test. ( D ) Summary schematic comparing Dgcr8 KO cells to Ddx6 KO cells. Dgcr8 KO leads to the loss of both translational repression and mRNA destabilization of miRNA targets, while Ddx6 KO only leads to the loss of translational repression of miRNA targets. mRNA stability is measured as the ratio of mRNA/4sU reads, changes in translation level are measured as the ratio of polysome/monosome reads, protein level changes are not directly measured but are predicted based on mRNA stability and translation level changes. Changes in translation level alone in Ddx6 KO cells produce similar phenotypes and global molecular changes to Dgcr8 KO cells. See also .

    Article Snippet: Cell line (Mouse) , Dgcr8 KO embryonic stem cell , PMID: 17259983; Novus Biologicals , NBA1-19349 , Dgcr8 KO mouse embryonic stem cell line previously generated in the Blelloch lab. Also available commercially from Novus Biologicals..

    Techniques: Comparison

    ( A ) Comparison between changes in nascent transcription (4sU-Seq) in Dgcr8 KO versus Ddx6 KO cells. n = 3 for wild-type, n = 4 for Ddx6 KO (2 replicates of each Ddx6 KO line), n = 3 for Dgcr8 KO. The p value was calculated with correlation significance test.

    Journal: eLife

    Article Title: Decoupling the impact of microRNAs on translational repression versus RNA degradation in embryonic stem cells

    doi: 10.7554/eLife.38014

    Figure Lengend Snippet: ( A ) Comparison between changes in nascent transcription (4sU-Seq) in Dgcr8 KO versus Ddx6 KO cells. n = 3 for wild-type, n = 4 for Ddx6 KO (2 replicates of each Ddx6 KO line), n = 3 for Dgcr8 KO. The p value was calculated with correlation significance test.

    Article Snippet: Cell line (Mouse) , Dgcr8 KO embryonic stem cell , PMID: 17259983; Novus Biologicals , NBA1-19349 , Dgcr8 KO mouse embryonic stem cell line previously generated in the Blelloch lab. Also available commercially from Novus Biologicals..

    Techniques: Comparison

    Journal: eLife

    Article Title: Decoupling the impact of microRNAs on translational repression versus RNA degradation in embryonic stem cells

    doi: 10.7554/eLife.38014

    Figure Lengend Snippet:

    Article Snippet: Cell line (Mouse) , Dgcr8 KO embryonic stem cell , PMID: 17259983; Novus Biologicals , NBA1-19349 , Dgcr8 KO mouse embryonic stem cell line previously generated in the Blelloch lab. Also available commercially from Novus Biologicals..

    Techniques: Generated, CRISPR, Recombinant, Plasmid Preparation, Sequencing, Software