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. 2023 Feb 9;14:726. doi: 10.1038/s41467-023-36236-2

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 5 — a Genomic snapshots illustrating ZC3H4 and RNA Pol II ChIP-seq signal at the promoter of the Spice1 gene (left panel) and an intergenic enhancer region (chr16:30,818,164-30,824,484) (right panel). b Metaplot analysis of ZC3H4 and RNA Pol II ChIP-seq at the TSSs of transcribed genes (n = 11,823, left panel) and enhancers (n = 4156, right panel). The read density of ZC3H4 is shown on the left axis and RNA Pol II is shown on the right axis. c Metaplot analysis of RNA Pol II and ZC3H4 ChIP-seq at the TSSs of transcribed SET1-independent (n = 9151) and SET1-dependent genes (n = 2633). The read density of RNA Pol II is shown on the left axis and read density of ZC3H4 on the right axis. d A schematic illustrating the ZC3H4-dTAG line and a representative western blot (n = 3) comparing ZC3H4 levels in wild type (WT) cells and the ZC3H4-dTAG line before (0 h) and following 2 h treatment with dTAG13. HDAC1 functions as a loading control. e Metaplot analysis of transcription (cTT-seq) in the ZC3H4-dTAG line that is either untreated (UNT, dark red) or treated with dTAG13 for 2 h (light red), showing upstream antisense transcription at all TSSs (left panel, n = 20,633) and enhancer transcription (right panels, n = 4156). f As in e, but for the dTAG-SET1A/B cell line. g An MA-plot showing log2 fold changes (Log2 FC) in transcription (cTT-seq) in the ZC3H4-dTAG line following 2 h treatment with dTAG13 (n = 20,633), determined using DESeq2. Significant changes in transcription (p-adj < 0.05 and >1.5-fold) are coloured red and the number of significantly changed genes is indicated. h A genomic snapshot of cTT-seq signal in the ZC3H4-dTAG line at the Mcat gene in untreated cells (UNT) or cells treated with dTAG13 for 2 h. i A box pot showing log2 fold change (Log2 FC) in transcription (cTT-seq) in the ZC3H4-dTAG line after 2 h dTAG13 treatment with all genes (n = 20,633) separated into deciles based on their transcription level in untreated cells. The boxes show interquartile range, centre line represents median, whiskers extend by 1.5× IQR or the most extreme point (whichever is closer to the median), while notches extend by 1.58× IQR/sqrt(n), giving a roughly 95% confidence interval for comparing medians. j Metaplots comparing CpG density and SET1A levels at the TSSs of CGI-associated genes that are increased in transcription after 2 h of ZC3H4 depletion (n = 1653) and those that are unchanged (n = 12,667).