Fig. 3: TRIM28 and FOXL2 act together on chromatin to maintain the ovarian pathway.
From: TRIM28-dependent SUMOylation protects the adult ovary from activation of the testicular pathway
a TRIM28 and FOXL2 are co-expressed in the nucleus of most follicular granulosa cells in 4-month-old control ovaries and in cells with flat nucleus surrounding follicles (identified as steroidogenic theca cells). In Trim28cKO ovaries, only few cells expressed FOXL2. Scale bar: 10 µm. At least three independent biological replicates were analysed, and the images presented are representative of all replicates. b Overlap between TRIM28 and FOXL2 genomic localisation in the adult ovary. Heatmaps in blue represent FOXL2 ChIP-seq and inputs reads mapped on TRIM28 peaks (±1 kb from the centre). Red traces represent TRIM28 ChIP-seq and inputs reads mapped on FOXL2 peaks. The Venn diagram on the right shows that 32,097 of the 51,764 TRIM28 peaks (62%) and of the 58,581 FOXL2 peaks (55%) overlap in control ovaries. c Examples of TRIM28 and/or FOXL2 peaks in/around genes the expression of which is altered in Trim28cKO ovaries. Upper panel: ovarian-specific genes downregulated in Trim28cKO ovaries (see also Supplementary Fig. 11). The Foxl2 gene is represented with the co-regulated non-coding Foxl2os gene106. Lower panel: testicular-specific genes upregulated in Trim28cKO ovaries (see also Supplementary Fig. 12). Green rectangles in the Sox9 panel: open chromatin regions described in the embryonic gonads, 13 corresponds to Enhancer13 that is crucial for sex-determination32. Relevant ChIP-seq peaks are highlighted in light blue (TRIM28) and light red (FOXL2). Yellow arrows indicate the gene orientation. d Pie charts showing up- and downregulated genes in Trim28cKO ovaries that are bound by TRIM28 and/or FOXL2. Genes are listed in Data S7. e Enrichment for binding motifs of transcription factors8 involved in granulosa cell fate maintenance (FOXL2, RUNX1 and ESR1/2) in reads of TRIM28 and FOXL2 ChIP-seq of adult control ovaries (this study), and TRIM28 ChIP-seq of bone marrow33 and of thymus34. n = 3 independent computational analyses. Bars are the mean ± SD. Ordinary one-way ANOVA with Tukey’s multiple comparisons test. Adj P Val: for all motifs ****<0.0001. RUNX1: *=0.0186; **=0.0037; ***=0.0003. ESR1: ***=0.00020,0324 ESR2: ***=0.0005. More statistical data are in Source data file. Source data are provided as a Source Data file. f Left, plot showing enriched proteins, ranked by relative abundance, identified by FOXL2 ChIP-SICAP. Only significant proteins (>2-fold enrichment over No-antibody control, n = 2) are shown. TRIM28 was identified amongst the top 20 proteins found to interact with FOXL2. Pie-chart (right) shows the percentage of the relative intensities of FOXL2 chromatin partners, normalised to the total abundance of the enriched proteins.
