Journal: bioRxiv
Article Title: ZCCHC4 Promotes Translation of Replication-dependent Histone mRNAs by Recruiting Cytoplasmic eIF3 complex
doi: 10.1101/2025.06.21.660898
Figure Lengend Snippet: a, Volcano plot depicting down-regulated (blue) and up-regulated (red) genes identified by ATAC-seq in ZCCHC4 KO cells compared with WT cells. b, Accessible chromatin profiling of HeLa WT and ZCCHC4 KO cells by ATAC-Seq. Peak distributions are presented as a profile plot (upper panel) and heatmap (low panel) over ± 3 kb centered on the TSSs. c, Western blot of CDKN1A in HeLa WT and ZCCHC4 KO cells. GAPDH was used as a loading control. d, WT and ZCCHC4 knockout cells were subjected to a double thymidine block to arrest cells at the G1/S checkpoint (G1/S). Cells were released from the block for either 4 or 8h and immunoblotted with the cyclinB1 antibodies to determine the expression of cyclin proteins. Async, asynchronous cells. GAPDH was used as a loading control. Blots shown are representative of three independent experiments. e, Representative images (left) and quantification (right) of cell cycle progression of ZCCHC4 KO versus WT HeLa cells. f, Propidium iodide staining detected by flow cytometry showing the distribution of each cell cycle stage (G1, S, and G2/M phase) after ZCCHC4 knockout at 0, 4, 6, 8, 10, 12, 14, 16 and 22h after cell synchronization and unsynchronized cells. The flow cytometry profiles (left) and quantitative statistical curves (right) are presented. Note: e, Two-tailed unpaired Student’s t tests. Data were presented as mean ± SD (n=3). * P <0.05, ns: not significant.
Article Snippet: The corresponding antibodies included anti-ZCCHC4 antibody with 1:1000 dilution (Abcam, ab209901), anti-RPS6 antibody with 1:1000 dilution (Proteintech, 66886-1-Ig), anti-RPL14 antibody with 1:1000 dilution (Proteintech, 14991-1-AP), anti-CDKN1A antibody with 1:1000 dilution (Proteintech, 10355-1-AP), anti-puromycin antibody with 1:1000 dilution (ABclonal, A21205), anti-H2B antibody with 1:2000 dilution (PTM-1007), anti-H2A antibody with 1:2000 dilution (ABclonal, A3692), anti-H2AZ antibody with 1:2000 dilution (Proteintech, 16441-1-AP), anti-HistoneH1.2 antibody with 1:1000 dilution (Proteintech, 19649-1-AP), anti HistoneH3 antibody with 1:2000 dilution (ABclonal, A22348), anti-α-Tubulin antibody with 1:3000 dilution (Proteintech, 80762-1-RR), anti-EIF3H antibody with 1:1000 dilution (Proteintech, 11310-1-AP), anti-EIF3A antibody with 1:1000 dilution (Proteintech, 26178-1-AP), anti-PDCD4 antibody with 1:1000 dilution (Proteintech, 12587-1-AP), anti-CyclinB1 antibody with 1:1000 dilution (ABclonal, A19037), anti-GAPDH antibody with 1:5000 dilution (Proteintech, 10494-1-AP), anti-β-actin antibody with 1:3000 dilution (SAB Signalway Antibody, 52901), and anti-Flag antibody with 1:2000 dilution (Sigma, F1804).
Techniques: Western Blot, Control, Knock-Out, Blocking Assay, Expressing, Staining, Flow Cytometry, Two Tailed Test
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