Journal: bioRxiv

Article Title: GC content shapes mRNA decay and storage in human cells

doi: 10.1101/373498

Figure Lengend Snippet: (A) mRNA stabilization after XRN1 silencing in HeLa and HCT116 cells (in brown) applies to GC-rich mRNAs. The analysis was performed as in . The GC content distribution for all mRNAs is presented for comparison (in grey). (B) mRNA stabilization after PAT1B silencing in HEK293 cells (in peach) applies to AU-rich mRNAs. The analysis was performed as in (A). (C) Read coverage of XRN1 targets (FC>1, n=333) and non-targets (FC<-1, n=139), as defined in the siXRN1 dataset. Their average read coverage was analyzed in control cells (upper panel) and after XRN1 silencing (lower panel), and normalized as described in the Methods. (D) Read coverage of PAT1B targets (FC>0.6, n=616) and non-targets (FC<-0.6, n=493), as defined in the siPAT1B dataset. The data were analyzed as in (C). (E,F) Clustering analysis of mRNAs depending on their GC content, their differential expression after silencing DDX6, XRN1 or PAT1B, and their enrichment in PBs. Raw GC content and log2 transformed ratio of the other datasets were used for the clustering of both transcripts (lines) and datasets (columns). The values were color-coded as indicated on the right scale, and the Spearman correlation matrix is presented in F (all p-values <10-48). The heatmap highlights the distinct fate of GC-rich and AU-rich mRNAs. See also Figures S4 and S6.

Article Snippet: Primary antibodies were: rabbit polyclonal anti-DDX6 (1:15000; Novus Biological), rabbit polyclonal anti-ribosomal S6 (1:5000; Cell Signaling Technology), anti-XRN1 (1:1000, Bethyl Laboratories), anti-XRN1 (1:5000 Novus Bioscience), anti-Pol II (1:100, Santa Cruz).

Techniques: Expressing, Transformation Assay

Journal: bioRxiv

Article Title: A role for DIS3L2 over human nonsense-mediated mRNA decay targets

doi: 10.1101/722702

Figure Lengend Snippet: Schematic representation of GADD45A mRNA uridylation frequency in LUC, DIS3L2 and DIS3L2+XRN1-depleted HeLa cells. Each blue circle represents the GADD45A mRNA 3’ end. The negative values upstream of the blue circle indicate the 3’ end position in the 3’UTR if the mRNA was trimmed, considering that the point zero is the polyadenylation site of GADD45A mRNA. Red squares indicate the length of the poly(A) tail added after trimming. Non-templated uridine residues detected at the 3’-end are indicated by green Us, the non-templated cytidine residues by blue Cs, the non-templated guanosine residues by dark red Gs, and non-templated adenosine residues by red As. The 3’ ends detected also revealed the existence of heterogeneous non-templated nucleotide additions.

Article Snippet: The membrane was incubated overnight (O/N) at 4°C with mouse anti-α-tubulin (Roche, loading control) at 1:4000, rabbit anti-DIS3L2 (Novus Biologicals) at 1:200, rabbit anti-XRN1 (Novus Biologicals) at 1:500 or mouse anti-FLAG M2 (Sigma) at 1:1000 dilution in blocking buffer.

Techniques:

Journal: eLife

Article Title: Cytoplasmic mRNA decay represses RNA polymerase II transcription during early apoptosis

doi: 10.7554/eLife.58342

Figure Lengend Snippet:

Article Snippet: Antibody , Anti-XRN1 (rabbit polyclonal) , Bethyl Laboratories , Cat#A300-433A; RRID: AB_2219047 , WB: (1:1000).

Techniques: Binding Assay, Recombinant, Virus, Reverse Transcription, Modification, Western Blot, TUNEL Assay, Cell Fractionation, Cloning, Expressing, Transfection, Sequencing, Control, Software

Journal: bioRxiv

Article Title: GC content shapes mRNA decay and storage in human cells

doi: 10.1101/373498

Figure Lengend Snippet: (A) mRNA stabilization after DDX6 silencing in HEK293 and K562 cells applies to GC-rich mRNAs. The fold-changes (FC) in mRNA accumulation (in green) were analyzed as in . (B) mRNA translation derepression after DDX6 silencing in HEK293 cells applies to AU-rich mRNAs. The fold-changes in translation rate (in orange) were analyzed as in (A). (C) GC-rich mRNAs are particularly enriched in the DDX6 CLIP experiment (in dark green). See also Figures S3-5.

Article Snippet: Primary antibodies were: rabbit polyclonal anti-DDX6 (1:15000; Novus Biological), rabbit polyclonal anti-ribosomal S6 (1:5000; Cell Signaling Technology), anti-XRN1 (1:1000, Bethyl Laboratories), anti-XRN1 (1:5000 Novus Bioscience), anti-Pol II (1:100, Santa Cruz).

Techniques:

Journal: bioRxiv

Article Title: GC content shapes mRNA decay and storage in human cells

doi: 10.1101/373498

Figure Lengend Snippet: (A) mRNA stabilization after XRN1 silencing in HeLa and HCT116 cells (in brown) applies to GC-rich mRNAs. The analysis was performed as in . The GC content distribution for all mRNAs is presented for comparison (in grey). (B) mRNA stabilization after PAT1B silencing in HEK293 cells (in peach) applies to AU-rich mRNAs. The analysis was performed as in (A). (C) Read coverage of XRN1 targets (FC>1, n=333) and non-targets (FC<-1, n=139), as defined in the siXRN1 dataset. Their average read coverage was analyzed in control cells (upper panel) and after XRN1 silencing (lower panel), and normalized as described in the Methods. (D) Read coverage of PAT1B targets (FC>0.6, n=616) and non-targets (FC<-0.6, n=493), as defined in the siPAT1B dataset. The data were analyzed as in (C). (E,F) Clustering analysis of mRNAs depending on their GC content, their differential expression after silencing DDX6, XRN1 or PAT1B, and their enrichment in PBs. Raw GC content and log2 transformed ratio of the other datasets were used for the clustering of both transcripts (lines) and datasets (columns). The values were color-coded as indicated on the right scale, and the Spearman correlation matrix is presented in F (all p-values <10-48). The heatmap highlights the distinct fate of GC-rich and AU-rich mRNAs. See also Figures S4 and S6.

Article Snippet: Primary antibodies were: rabbit polyclonal anti-DDX6 (1:15000; Novus Biological), rabbit polyclonal anti-ribosomal S6 (1:5000; Cell Signaling Technology), anti-XRN1 (1:1000, Bethyl Laboratories), anti-XRN1 (1:5000 Novus Bioscience), anti-Pol II (1:100, Santa Cruz).

Techniques: Comparison, Expressing, Transformation Assay