Additional file 2 of Immediate and durable effects of maternal tobacco consumption alter placental DNA methylation in enhancer and imprinted gene-containing regions
Creators: name: Rousseaux, Sophie
name: Seyve, Emie
name: Chuffart, Florent
name: Bourova-Flin, Ekaterina
name: Benmerad, Meriem
name: Charles, Marie-Aline
name: Forhan, Anne
name: Heude, Barbara
name: Siroux, Valérie
name: Slama, Remy
name: Tost, Jorg
name: Vaiman, Daniel
name: Khochbin, Saadi
name: Lepeule, Johanna
Publisher: name: figshare
Year: 2020
Subjects: subject: Genetics
subject: FOS: Biological sciences

description: Additional file 2: Supplementary Figure S1. cellular heterogeneity estimated by the RefFreeEWAS method on 668 placenta samples from the EDEN cohort (A) contribution (%) of each latent variable (B) Pearson correlation between the 6 latent variables. Supplementary Figure S2. Q-Q plot, genomic inflation factor (lambda) and Bayesian inflation factor (BIF) for the association between each methylation site of the EWAS (425,878 CpGs) and tobacco smoking during pregnancy (current, never, former smoker). Results were adjusted for child sex, parity, education level, season of conception, study center, maternal body mass index before pregnancy, maternal age at delivery, gestational duration, paternal smoking status at conception, batch, plate and chip and estimated cellular heterogeneity. Supplementary Figure S3. H3K4me3 (A), H3K4me1 (B) and H3K27ac (C) ChIPSeq signal of 3 placenta replicates. The top panels represent the mean signal values centered on our 203 DMRs +/-2kb (blue) and the mean signal values centered on the 420 random regions +/-2kb (green). The center panels represent heatmaps of the corresponding ChIP-Seq signals centered on our 203 DMRs +/-2kb (upper heatmaps) or on the 420 random regions+/-2kb (lower heatmaps). Supplementary Figure S4. Expression of genes in normal human tissues from RNA-seq data. The barplots show average expression levels of the 16 imprinted genes overlapping our DMRs in normal tissues and development stages (adult, embryonic and fetal). For the sake of clarity, the plots are separated in two panels of 8 genes. Vertical lines on the top of the barplots represent standard deviations. Barplots representing expressions in placenta are in green. RNA-seq data in normal tissues were provided by GTEx portal and NCBI Sequence Read Archive (datasets PRJNA280600, PRJEB4337, PRJEB2445, PRJNA270632, GSE70741, GSE53096). The expression levels are represented in log-transformed RPKM (Read per Kilobase Million) values after addition of a pseudo count of 1, i.e. log-transformed RPKM = log2(1+RPKM). MIR675 has no expression in any of the analyzed samples. Figure S5. The methylation levels of 7 imprinted loci are consistently modified following exposure to cigarette smoking. The left, center and right panels are respectively centered on the DMRs associated with the NNAT locus +/- 2kb, the SGCE/PEG10 locus +/- 2kb and the H19/MIR675 locus +/- 2kb. The top panels show heatmaps respectively corresponding to H3K4me3, H3K4me1 and H3k27ac enrichments in placenta around the center of the region of interest +/- 2 kilobases. For each mark we downloaded triplicates from ENCODE data (see Methods). We computed the enrichment matrices from bigWig files using the deepTools software and displayed it using a custom R script. The middle panels show genes and CpG islands from the Illumina Infinium Human Methylation 450K BeadChip annotations, ICR from Pervjakova et al. [72], the DMR we identified and the corresponding Illumina probes of the region of interest. The lower panels show the methylation changes in smokers (S) (resp. Former Smokers (FS)) in red (resp. blue) compared to nonsmokers. The points represent the beta of the linear model and the error bars correspond to 2 standard deviations. The Y-axis represents the distributions of the variations of the regression coefficient (i.e. the associated change in DNA methylation level between 0 and 1) for each CpG within the DMRs selected by our analysis.