Journal: BioEssays : news and reviews in molecular, cellular and developmental biology

Article Title: Cytoplasmic mRNPs revisited: Singletons and condensates.

doi: 10.1002/bies.202000097

Figure Lengend Snippet: FIGURE 1 Schematic overview of the different cytoplasmic mRNP particles present in eukaryotic cells and their interrelationships. Protein exchange or remodeling steps are indicated between the different mRNP states: pre-translational mRNPs, active translating mRNPs, stress granules and P-bodies. (a) Mature messenger RNAs coated with RNA-binding proteins exit the nucleus through the nuclear pore. At this stage, three characteristic RNA-binding complexes are present in pre-translational mRNPs; namely CBP20/80 (forming the cap-binding complex CBC), the exon-junction complex (EJC), and PABPN1 (together with PABPC1). In the cytoplasm, pre-translational mRNPs are transported in a

Article Snippet: Antibodies against CBP80 (E-7, Santa Cruz Biotechnology), YBX1 (ab12148, Abcam) and PABPC1 (HPA067156, Atlas Antibodies) were directly conjugated to Alexa Fluor 488, 568 and 647 dyes, respectively.

Techniques: RNA Binding Assay, Binding Assay

Journal: BioEssays : news and reviews in molecular, cellular and developmental biology

Article Title: Cytoplasmic mRNPs revisited: Singletons and condensates.

doi: 10.1002/bies.202000097

Figure Lengend Snippet: FIGURE 2 Super-resolution fluorescence microscopy images acquired by Structured Illumination Microscopy of pre-translational mRNPs, P-bodies and stress granules in HeLa cells. Upper panel—Direct immunofluorescent staining of a linear pre-translational mRNP unit with three of its protein constituents: CBP80 (green), YBX1 (red), and PABPC1 (cyan). CBP80 is at the 5′ terminus of the mRNP (5′-cap), YBX1 is coating the mRNA body, and PABPC1 is binding to the poly(A) tail at the 3′ terminus. Antibodies against CBP80 (E-7, Santa Cruz Biotechnology), YBX1 (ab12148, Abcam) and PABPC1 (HPA067156, Atlas Antibodies) were directly conjugated to Alexa Fluor 488, 568 and 647 dyes, respectively. Scale bars = 5 μm (whole cell image) and 0.1 μm (zoomed-in image). Lower panels—Image of a whole cell with its P-bodies visualized by the presence of DCP1a-GFP and a zoomed-in view of one P-body (left). Indirect immunofluorescence staining of G3BP in a cell stressed with 1 mM sodium arsenite for 20 minutes showing stress granules in the cytoplasm and a detailed view of two stress granules (right). A mouse monoclonal anti-G3BP (ab56574, Abcam) primary antibody and a secondary antibody anti-mouse Alexa Fluor 488 were used. Scale bars = 5 μm (whole cell images) and 0.5 μm (zoomed-in images).

Article Snippet: Antibodies against CBP80 (E-7, Santa Cruz Biotechnology), YBX1 (ab12148, Abcam) and PABPC1 (HPA067156, Atlas Antibodies) were directly conjugated to Alexa Fluor 488, 568 and 647 dyes, respectively.

Techniques: Fluorescence, Microscopy, Staining, Binding Assay, Immunofluorescence

Journal: Journal of experimental & clinical cancer research : CR

Article Title: EIF4A3-mediated biogenesis of circSTX6 promotes bladder cancer metastasis and cisplatin resistance.

doi: 10.1186/s13046-023-02932-6

Figure Lengend Snippet: Fig. 9 Schematic diagram describes the mechanism by which circSTX6 promotes metastasis and chemoresistance of BCa through rehulation of SUZ12 by sponging miR-515-3p and interacting with PABPC1, respectively

Article Snippet: Short hairpin RNAs targeting circSTX6, SUZ12 and PABPC1 were synthesized by BIOMED, and were cloned into the pLKO.1 vector (addgene, USA).

Techniques:

Journal: Genes & Development

Article Title: tRNA synthetase activity is required for stress granule and P-body assembly

doi: 10.1101/gad.353535.125

Figure Lengend Snippet: Inhibition of tRNA charging activates the ISR without inducing RNP granules. ( A ) U-2 OS cells were treated with 0.2% DMSO carrier, 20 µM halofuginone (HF), 250 µM sodium arsenite (As), or 1.25 µM thapsigargin (Tg) for 1 h, and Western blotting for phosphorylated and total eIF2α was done. Total protein ( n = 3 independent replicates) and molecular weights (in kilodaltons) are shown. ( B ) Translation activity was measured using bioorthogonal noncanonical amino acid tagging in cells treated as in A . Methionine or azidohomoalanine (AHA) was added 10 min prior to collection. The ratio of nascent to total protein is shown from n = 3 independent replicates. ( C ) U-2 OS cells stably expressing GFP-G3BP1 (green) were treated as in A and fixed, and nuclei were stained with Hoechst (blue). The percentage of cells with SGs was quantified from n = 3 independent experiments; ≥305 cells were counted in each condition. Scale bars, 10 µm. ( D ) Cells were treated as in C , and immunofluorescence and fluorescence in situ hybridization were done to detect UBAP2L (magenta), PABPC1 (cyan), and poly(A) RNA with oligo(dT) probes (yellow) ( n = 2 independent experiments). ( E ) Cells were treated with 0.2% DMSO, 20 µM HF plus 0.2% DMSO, 250 µM arsenite, or 1.25 µM thapsigargin for 1 h, and the percentage of cells with SGs was quantified ( n = 3 independent experiments); ≥287 cells were counted per treatment. ( F ) Cells were treated, and immunofluorescence microscopy was done as in D to detect the P-body marker DCP1A. Quantification of the percentage of cells with P-bodies (PBs) from n = 3 independent experiments is shown; ≥364 cells were counted per treatment. ( G ) Cells were treated as in C to detect XRN1 (magenta) and EDC4 (yellow) by immunofluorescence microscopy ( n = 2 independent experiments). ( H ) U-2 OS cells were treated as in E , and immunofluorescence microscopy was done to detect DCP1A. The percentage of cells with PBs from n = 3 independent experiments is shown; ≥351 cells were counted per treatment across all replicates. Representative images are shown with the average ± SEM for each experiment, and green, gray, and pink points represent the average of each replicate. Statistical significance was assessed with an ordinary one-way ANOVA followed by Tukey's multiple comparisons test. (*) P < 0.05, (**) P < 0.01, (***) P < 0.005, (****) P < 0.001.

Article Snippet: PABPC1 , Proteintech , 66809 , 1:100 (immunofluorescence).

Techniques: Inhibition, Western Blot, Activity Assay, Stable Transfection, Expressing, Staining, Immunofluorescence, Fluorescence, In Situ Hybridization, Microscopy, Marker

Journal: Stem Cell Research & Therapy

Article Title: Deer thymosin beta 10 functions as a novel factor for angiogenesis and chondrogenesis during antler growth and regeneration

doi: 10.1186/s13287-018-0917-y

Figure Lengend Snippet: Antibodies

Article Snippet: Vascular endothelial growth factor-D (VEGFD) , Rabbit , Polyclonal antibody , Elabscience, China , E-AB-33213 , WB: 1 μg/ml (1:1000).

Techniques: Control, Labeling