Additional file 1 of Transient Polycomb activity represses developmental genes in growing oocytes
Creators: name: Jarred, Ellen G.
name: Qu, Zhipeng
name: Tsai, Tesha
name: Oberin, Ruby
name: Petautschnig, Sigrid
name: Bildsoe, Heidi
name: Pederson, Stephen
name: Zhang, Qing-hua
name: Stringer, Jessica M.
name: Carroll, John
name: Gardner, David K.
name: Van den Buuse, Maarten
name: Sims, Natalie A.
name: Gibson, William T.
name: Adelson, David L.
name: Western, Patrick S.
Publisher: name: figshare
Year: 2023
Subjects: subject: Genetics
subject: FOS: Biological sciences

description: Additional file 1: Fig. S1: Zp3Cre-mediated Eed deletion results in loss of EED in oocytes from the primary follicle stage in Eed-hom females. Representative images of EED (red) immunofluorescence analysis in primary (top), secondary (middle) and antral (bottom) follicles in females producing Eed-wt and Eed-hom oocytes. a. shows widefield follicle images and b. shows magnified images of the area inside the white squares containing the oocyte nucleus. Lamin B1 (green) marks the nuclear lamina and delineates the edge of the oocyte nucleus in b. DAPI (blue) shows DNA in somatic cells. Images are representative of two ovaries from three biological replicates. Scale bars: 20 μm. Fig. S2: Deletion of Eed in oocytes moderately increased the rate of Surrounded Nucleolus (SN) GV oocytes compared to Non-Surrounded Nucleolus (NSN) GV oocytes. Percentage of SN (left) and NSN (right) GV oocytes obtained from Eed-wt, Eed-het, and Eed-hom females during oocyte collections. *P &lt; 0.05, one-way ANOVA plus Tukey’s multiple comparisons test, N = 7 Eed-wt, 5 Eed-het and 7 Eed-hom females. Error bars represent mean ± standard deviation. Fig. S3: Loss of Eed in growing oocytes did not impact the transcription of genes encoding other core PRC2 subunits, PRC1 core components or DNMTs. Expression of core (a) PRC2 and (b) PRC1 components, and (c) DNMTs in Eed-wt, Eed-het and Eed-hom oocytes. Data represent the mean transcripts per million reads (TPM) for each gene from N = 5 Eed-wt, 4 Eed-het and 6 Eed-hom females. FDR &lt; 0.05 for Eed only, error bars represent mean ± standard deviation. Fig. S4: Eed deletion results in up-regulation of a subset of imprinted and X-linked genes in Eed-hom GV oocytes. Expression of core (a) putative H3K27me3-imprinted (b) classically imprinted, and (c) X-linked Eed oocyte DEGs in Eed-het and Eed-hom oocytes. Data represent the mean transcripts per million reads (TPM) for each gene from N = 5 Eed-wt, 4 Eed-het and 6 Eed-hom females. For classically imprinted and X-linked genes, expression levels were highly varied across some genes and have therefore been graphed for genes with low (TPM &lt; 1), medium (TPM 1 to 5) or high (TPM &gt; 5) expression. For all genes FDR &lt; 0.05 in Eed-hom vs Eed-het oocytes, Error bars represent mean ± standard deviation. Fig. S5: Loss of Eed in growing oocytes did not impact expression of LINE-1 transposons. (a) Percentages of total input reads which aligned to a LINE-1 (L1) element. (b) Number of reads which map to unique and multiple L1s. (c) Proportions of reads according to the number of sites mapped to per read, to a maximum of 20. For (a-c), data represent individual replicates from Eed-wt (n = 5), Eed-wt Cre (n = 2), Eed-het (n = 4) and Eed-hom (n = 6) females. Fig. S6: Eed oocyte DEGs were not dysregulated in pre-implantation embryos. Venn Diagram comparing Eed oocyte DEGs against DEGs identified in Eed maternal null morula and blastocyst embryos. Six genes (Plxnd1, Tceal8, Rap2c, Bbx, Xlr3c and Trm2b) were common in oocytes and morula embryos, five genes (Chrdl1, Lonrf2, Trim6, Cyp1b1 and Ccbe1) were common in oocytes and blastocyst embryos, and two genes (Tspan6 and Gk) were common in morula and blastocyst embryos. For full DEG lists see Tables S1, S6 and S7. Morula and Blastocyst data sets were generated by analysis of published raw data sets (Manuscript References [33, 34).