Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2019 Sep 26;10:4374. doi: 10.1038/s41467-019-12355-7

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2019

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/.

PMC Copyright notice

Fig. 7 — Dnmt3b represses Wnt9b and Shh in vivo and in vitro. a Methylation of Wnt9b locus in E11.5 embryos of indicated genotypes determined by RRBS. b Wnt9b expression by RNA-seq (n = 3; *p < 0.05 by DESeq; left) and real-time qRT-PCR (n = 3; normalized to Gapdh; right) in E11.5 embryos. Horizontal line represents median, bounds of box—likely range of variation and whiskers—min and max values. c Immunoblot of Wnt9b and Hsc70 levels in E11.5 embryos. d Wnt9b expression by real-time qRT-PCR in fetal brain of E11.5 embryos (n = 3), normalized to Gapdh. e GSEA using RNA-seq data shows positive enrichment in Beta-catenin–TCF complex assembly in E11.5 Dnmt3b^−/− embryos (n = 3). Normalized enrichment scores (NES), false discovery rate (FDR) and p values are shown. f Left. Fgf8 expression by RNA-seq in Dnmt3b^+/+, Dnmt3b^−/−, and Dnmt3b^CI/CI E11.5 embryos (n = 3), *p < 0.05 (DESeq). Horizontal line represents median, bounds of box—likely range of variation and whiskers—min and max values. Right. Immunoblot analysis of Fgf8 levels in E11.5 embryos. g GSEA shows negative enrichment in developmental pathways in E11.5 Dnmt3b^−/− embryos relative to Dnmt3b^+/+ (n = 3). h Dnmt3b and Hsc70 expression in mouse Dnmt3b^−/− T cell lymphoma cells (^−/−) 72 h after transduction with lentiviruses expressing control vector (empty), Dnmt3b^WT (3b WT), Dnmt3b^CI (3b CI) as analyzed by immunoblot. i Real-time qRT-PCR analysis of Wnt9b and Fgf8 expression in samples prepared as in h. Average of two independent experiments normalized to β-actin. j Left: Schematic of mouse Wnt9b promoter with position of primers (arrows): P1: −763/−582 bp, P2: −603/−411 bp; P3: −342/−127 bp; P4: −143/ + 36 bp relative to TSS. Right: Real-time qRT-PCR on DNA immunoprecipitated with anti-FLAG antibody from Dnmt3b^−/− lymphoma cells expressing FLAG tagged Dnmt3b^WT or empty vector. Data are shown as averaged fold enrichment over empty vector (n = 2), *p < 0.001 (two-tailed Student’s t-test). Assay was performed in triplicates and normalized to the input DNA. k Shh expression by real-time qRT-PCR in samples described in h. Averaged data of two independent experiments were normalized to β-actin, *p < 0.001 (two-tailed Student’s t-test). Data in figures b, d, i, j, and k are presented as means ± SEM. Source data are provided as a Source Data file