Epigenome, MED12 and Progesterone Action in Uterine Leiomyomas

子宫肌瘤(LM，肌瘤)是女性最常见的肿瘤。没有长期的医疗治疗是 可用。每个LM似乎起源于单个突变的LM干细胞(LSC)在 肌层(MYO)。LSC占肿瘤质量的5%，并分化为中间细胞群 (LIC，7%)，然后成为终末分化细胞(LDC)，占肿瘤体积的88%。司机 介体复合体12亚单位(mut-MED12)突变发生在70%的LM中。孕酮(P4)及其受体 受体PR对于LM的生长是必不可少的。富含PR的LIC通过旁分泌将P4信号转导到PR缺乏的LSC 激活它们扩散的因素。PR选择性P4拮抗剂醋酸利普利酯(UPA)收缩Lm和 减轻了其症状，但由于有肝脏损伤的风险，它的使用被停止了。我们的总体目标是定义 全基因组P4作用在MUT-MED12相关的LM肿瘤发生中的作用及新发现 治疗靶点。我们发现MUT-MED12在物理上与PR和全基因组PR-染色质相互作用 与正常MYO组织相比，表达MUT-MED12的LM中相互作用景观显著失调 携带野生型MED12。MUT-MED12增强P4靶基因顺式调控元件的PR募集 编码旁分泌生长因子、细胞因子和细胞外基质蛋白对LSC增殖和 LM肿瘤发生学。此外，MED12中具有与人类等同的功能获得突变的小鼠的子宫 发展对P4的反应的LM，而MED12基因敲除阻断了小鼠子宫中的P4/PR信号转导。我们 假设MUT-MED12改变PR-染色质相互作用特征并增强LIC中的P4作用， 为LSC的生存和扩散提供支持。使用Chip-seq、RNA-seq、starr-seq和 CRISPR/Cas9-基因编辑策略，以及体内PDX和mut-MED12敲入小鼠模型，我们提出了 目的如下：(1)确定PR-染色质相互作用基因座是否与MUT-MUT-R特异相关。 MED12在不同的干细胞支持细胞群(LIC)中刺激P4活动，然后发送肿瘤 旁分泌信号促进LSC活性和肿瘤生长。我们将检验MUT-MED12的假设 与PR相互作用并改变其与染色质的相互作用，从而增强LIC对P4的反应性 激活支持邻近LSC功能的基因转录和旁分泌信号。(2)界定 MUT-MED12的致瘤活性是否通过改变PR-染色质相互作用而介导 等效人MUT-MED12小鼠模型。我们将检验MUT-MED12破坏染色质的假设 PR结合位点周围的特征，从而支持对LM至关重要的基因转录和途径 肿瘤发生，而uPA通过改变PR-染色质-表观基因组相互作用和 逆转疾病相关基因的不适当表达。破译全基因组机制，请访问 一个明确的细胞群体水平将帮助我们确定与mut-med12相关的特定于基因型的新靶点。 用于治疗LM的药物基因组学和精确医学。

Uterine leiomyoma (LM, fibroid) is the most common tumor in women. No long-term medical treatment is available. Each LM seems to originate from the clonal expansion of a single mutated LM stem cell (LSC) in the myometrium (MYO). LSC comprise 5% of tumor mass and differentiate into an intermediate cell population (LIC, 7%), which then become terminally differentiated cells (LDC) comprising 88% of the tumor bulk. Driver mutations of mediator complex subunit 12 (mut-MED12) occur in 70% of all LM. Progesterone (P4) and its receptor PR are essential for LM growth. PR-rich LIC transduce P4 signaling to PR-deficient LSC via paracrine factors to activate their proliferation. Ulipristal acetate (UPA), a PR-selective P4 antagonist, shrank LM and reduced its symptoms, but its use was halted because of risk of liver injury. Our overall goal is to define the role of genomewide P4 action in the etiology of mut-MED12‒associated LM tumorigenesis and identify novel therapeutic targets. We found that mut-MED12 physically interacts with PR and genomewide PR-chromatin interaction landscapes are dramatically dysregulated in LM expressing mut-MED12 vs. normal MYO tissue carrying wild type MED12. Mut-MED12 enhances PR recruitment to cis-regulatory elements of P4 target genes encoding paracrine growth factors, cytokines and extracellular matrix proteins critical for LSC proliferation and LM tumorigenesis. Furthermore, the uteri of mice with a human-equivalent gain-of-function mutation in Med12 develop LM in response to P4, whereas Med12 knockout blocks P4/PR signaling in mouse uterus. We hypothesize that mut-MED12 alters PR-chromatin interaction signatures and enhances P4 action in LIC, providing a support niche for LSC survival and proliferation. Using ChIP-seq, RNA-seq, STARR-seq, and CRISPR/Cas9-gene editing strategies, and in vivo PDX and mut-Med12 knock-in mouse models, we propose the following Aims: (1) Determine whether PR-chromatin interaction loci specifically associated with mut- MED12 stimulate P4 action in a distinct stem-support cell population (LIC), which then send tumorigenic paracrine signals to increase LSC activity and tumor growth. We will test the hypothesis that mut-MED12 interacts with PR and alters its interaction with chromatin, thereby enhancing P4 responsiveness of LICs to activate gene transcription and paracrine signaling that support the function of adjacent LSC. (2) Define whether tumorigenic activity of mut-Med12 is mediated via altering PR-chromatin interaction landscapes in a human-equivalent mut-Med12 mouse model. We will test the hypothesis that mut-Med12 disrupts chromatin features surrounding PR-binding sites, thereby supporting gene transcription and pathways critical for LM tumorigenesis, whereas UPA shrinks LM via altering the aberrant PR-chromatin-epigenomic interactions and reversing inappropriate expression of disease-associated genes. Deciphering the genomewide mechanisms at a defined cell population level will help us identify genotype-specific novel targets associated with mut-MED12 for pharmacogenomics and precision medicine in the treatment of LM.