Figure 4. P300 Promotes Invasion and Survival of ARID1A Mutant Endometriotic Epithelia.

(A) Western blot analysis as indicated in 12Z cells, representative of 2 independent experiments.

(B) Invasion of 12Z following indicated treatments. Representative images and total invaded cell numbers are shown (scale bar, 500 μm). Means ± SDs, n = 3. Unpaired, 2-tailed t test.

(C) Survival of mice based on time until vaginal bleeding. LtfCre^0/+; (Gt)R26Pik3ca*^H1047R; Arid1a^fl/fl (n = 16) median (μ_1/2) 107 days. LtfCre^0/+; (Gt) R26Pik3ca*^H1047R; Arid1a^fl/fl; Ep300^fl/fl (n = 12) median 143 days (p = 0.0018, Mantel-Cox test). LtfCre^0/+; Ep300^fl/fl mice were aged to 187 days, and no phenotypes were observed (n = 6).

(D) Histology and IHC using indicated antibodies (n ≥ 2 mice) in endometrium (scale bar, 200 μm). KRT8 was a positive control for endometrial epithelium. The arrowheads indicate epithelia.

(E) Quantification of H3K27ac and H3K18ac IHC, ratio of H-scores of epithelia to stroma. Means ± SDs, n = 4–8 mice, unpaired, 2-tailed t test.

(F) Western blot of H3K27ac following A-485 treatment of 12Z for 24 h and densitometry of H3K27ac relative to H3, normalized to control (vehicle). Means ± SDs, n = 3–5 independent replicates per condition. Unpaired, 2-tailed t tests were performed in comparison to the vehicle treatment condition. Irrelevant lanes were removed from the image; see Figure S7.

(G) Viability assay for cells treated with A-485, normalized cell counts relative to vehicle control. Raw data are presented in Figure S4C. Half-maximal inhibitory concentration (IC₅₀) values were not significantly different between 12Z untreated and control shRNA, or between control shRNA and shARID1A (unpaired, 2-tailed t test). Means ± SDs, n = 4.

(H) Invasion of 12Z following indicated cell treatments. Representative images and total invaded cell numbers are shown (scale bar, 500 μm). Means ± SDs, n = 4. Unpaired, 2-tailed t tests performed in comparison to siARID1A + vehicle.

(I) Caspase 3/7 activity of indicated cell treatments. Means ± SDs, n = 3. Unpaired, 2-tailed t test.

(J) Ratio of dead to live cells after 16 h in Matrigel. Means ± SDs, n = 6. Unpaired, 2-tailed t test.

*p < 0.05, **p < 0.01, and ***p < 0.001.