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Emplified by Fzd5 and Fzd8 (Figure 2B,C).

por Rudolph Schaefer (2020-04-01)

Emplified by Fzd5 and Fzd8 (Figure 2B,C). PubMed ID: Fzd5 methylation was 12.1 (range 6.8 to 21.3 ) in PU.1-wt animals and 33.4 (range 18.2 to 44.9 ), 40.9 (range 34 to 47.8 ) and 50.4 (range 39.8 to 92.8 ) in preleukemic, early leukemic and late leukemic stage animals, respectively. The stage-dependent increase of Fzd8 methylation was similar to that of Fzd5, starting with 22.8 (range 15.9 to 37.3 ) in PU.1-wt animals and increasing to 30.6 (range 24.3 to 44.7 ), 35.4 (range 28.1 to 41.9 ) and 44.8 (range 36.1 PubMed ID: to 67.2 ) in the preleukemic, early leukemic and late leukemic stage animals, respectively.Acute myeloid leukemia-specific methylation changesTo ensure that the observed changes in DNA methylation are not the result of tissue-specific methylation patterns and, thus, reflect differences in cell type composition, we analyzed Fzd5, Fzd8 and eight additional differentially methylated regions in four hematopoietic cell types, LSK (lineage-negative, c-Kit+, Sca-1+ cells), CMPs, GMPs andSonnet et al. Genome Medicine 2014, 6:34 7 ofAZfp574 Ptrf Lmna Spen Bsn Mdga Pde4d Robo3 Map3k6 Nfix Ltbp3 Mapk11 Il17re Myh14 Gm1679 Gabbr Chr10:012631455-012632051 2410137M14Rik Chr6:030888000-030889519 Fzd8 Fzd5 chr5:067705737-067707409 Mab21l1 1200009l06Rik Egr3 Drd4 Efcab4a Hic1 Il4il Prdm16 chr1:107998952-108000925 Cxcl14 Il17rc Zfp775 Cdh15 Stc2 Tcp11 Adam11 Fosb Chr8:125563887-DFzdDNA Methylation [ ]60 50 40 30 20 100100BFzdDNA Methylation [ ]100 80 60 40 20C** * *** *** Ro 31-8220 (mesylate) **** *100 80 60 40 20EFzdFzdDNA Methylation [ ]*** ** **DNA Methylation [ ] 40 30 20 10Figure 2 Validation of screening results by quantitative DNA methylation analysis (MassARRAY). (A) Heatmap showing the average methylation levels in 40 gene-specific amplicons (columns) and individual PU.1-kd and PU.1-wt animals (rows). Methylation levels range from 0 (light green) to 100 (dark blue). Grey boxes represent missing values. The bar to the left of the heatmap indicates different disease stages (black, preleukemic; blue, early leukemic; red, late leukemic; green, PU.1-wt). Unsupervised clustering discriminates PU.1-kd and PU.1-wt animals. (B,C) DNA methylation levels of Fzd5 (B) and Fzd8 (C). Average amplicon methylation is shown for different animals of the different stages. The black bar represents the median methylation within one stage. Mann-Whitney U test was used to test for differences between the different disease stages (*P < 0.05, **P < 0.01, ***P < 0.001). (D,E) Methylation levels (heatmaps on top, bar graphs below) of Fzd5 (D) and Fzd8 (E) amplicons in sorted cells of PU.1-wt animals and two groups of preleukemic PU.1-kd animals, KD1 and KD2. The sorted cells comprise LSKs (lineage-negative, Ska1-positive, c-kit-negative cells), CMPs (common myeloid progenitor cells), GMPs (granulocyte-macrophage progenitor cells) and MEPs (megakaryocyte-erythroid progenitor cells). The heatmaps display single CpG units (columns) of PU.1-kd or PU.1-wt animals (rows). Methylation values range from 0 (light green) to 100 (dark blue). The bar graphs show average methylation (y-axis) of the different amplicons. In (D), analysis of KD2-GMP failed, indicated by grey CpG units in the heatmap and missing value in the bar graph.MEPs, which were enriched from both PU.1-wt and PU.1kd BM, respectively, and which represent different stages of hematopoietic commitment. We found both genes to be similarly hypermethylated in all four cell types in PU.1kd BM (F.