(A) Mouse model 1 (Med12^fl/+ Med12R^mt/+ Amhr2-Cre). A subset of cells express the Med12 c.131G>A variant from the autosomal ROSA locus, while X chromosome–derived Med12 will either be conditionally excised at 1 locus, or silenced by X chromosome inactivation at the other locus. Transcription from the mutant autosome (A^mt/+) is indicated with an arrow, and the promoter region is depicted in green. The blue star indicates the Med12 c.131G>A variant. The pink star indicates the floxed Med12 allele on the X chromosome (X^cko), which, in the presence of Amhr2-Cre, will lose exons 1–7. In the cells in which the Med12 floxed allele is inactivated, WT Med12 will be expressed. In cells with an active Med12 floxed allele, only mutant Med12 will be expressed. The red chromosome indicates the inactivated X chromosome. (B) cDNA sequencing from tumor tissue showing the presence of the mutant c.131G>A variant (green chromatogram peak, black arrow) in the absence of the WT Med12 allele, consistent with the monoclonal composition of tumor cells. (C) Eighteen-week-old multiparous Med12^fl/+ Med12R^mt/+ Amhr2-Cre uterus revealed a 4-mm tumorous lesion (outlined by white dotted lines). (D) Histological examination confirmed the presence of a large leiomyoma nodule growing from the smooth muscle layer of the uterus. A higher-magnification image of the black-boxed neoplastic area appears in E and shows the presence of fascicles with plump spindle cells, eosinophilic cytoplasm, and ECM deposits. (F) Twenty-four-week-old Med12^fl/+ Med12R^mt/+ Amhr2-Cre multiparous female uterus showing multiple nodules (white arrows). (G) Multiple leiomyoma nodules are outlined by black dotted lines, and the black box, shown at higher magnification in H, highlights fibrosis and ECM deposition. Approximately 80% (16 of 20 females) of the uteri exhibited leiomyoma-like lesions and hyperplasia. LM, leiomyoma; ES, endometrial stroma; MY, myometrium. Scale bars: 2,000 μm (C and F), 1,000 μm (D), 500 μm (G), 100 μm (H), 50 μm (E).