Stem cell reprogramming during oncogenesis and development

项目总结： 组织干细胞对死亡细胞的补充和伤口修复至关重要。上一首 研究表明，许多癌症可能是由干细胞功能障碍引起的， 要么是通过突变的积累，要么是最近表观遗传学中的异常 风景。Sox9是一种转录因子，在细胞的发育和发育过程中发挥着重要作用。 许多干细胞室的维护。在皮肤中，Sox9对毛囊是必不可少的 而表皮干细胞的特点是缺乏Sox9的表达。 有趣的是，基底细胞癌(BCC)过表达SOX9，但已被证明 来源于表皮干细胞，而非毛囊干细胞。此外，Sox9对于BCC AS至关重要 Sox9的基因缺失完全消除了体内肿瘤的形成。它们的作用机制 异位Sox9参与了BCC和其他癌症的形成，目前尚不清楚。 然而，在HFs中，Sox9结合并调节关键的开放染色质结构域(超级增强子) 它编排了干细胞基因，我的初步研究表明，异位表达 在成人表皮中的Sox9通过激活先前沉默的染色质来改变染色质景观 可能对基底细胞癌转化起重要作用的基因。因此，我假设Sox9会引发 通过直接结合和重塑KEY染色质改变表皮干细胞的命运 增强剂。利用新开发的表达可诱导的Sox9的转基因小鼠 表皮干细胞，我将绘制并注释染色质和转录变化 在Sox9诱导后发生，以精确定位Sox9阴性表皮和Sox9+的偏离 毛囊干细胞。利用核小体结合实验的组合， 免疫沉淀和质谱学，我会确定Sox9是否可以直接 识别和重塑异染色质，或者它是否通过 其他蛋白质。最后，使用福克斯实验室开发的一种强大的宫内慢病毒方法，我 将在以下背景下对使用CRISPR/CAS确定的候选人的结果进行功能测试 肿瘤的形成和发展。我的研究将直接加深我们对Sox9的了解 介导的染色质重塑和随后的致癌细胞和干细胞的激活 转录途径。确定Sox9的关键靶基因以及涉及的蛋白质 在表皮到基底细胞的重新编程将导致新疗法的发展，用于 将肿瘤的表观遗传学图景“重置”为非恶性形态。

Project Summary: Tissue stem cells are critical for the replenishment of dying cells and for wound-repair. Previous studies have demonstrated that many cancers can arise from the dysfunction of stem cells, either through accumulation of mutations, or more recently, aberrancies in the epigenetic landscape. Sox9, a transcription factor, plays an important role in the development and maintenance of many stem cell compartments. In the skin, Sox9 is essential for hair follicle stem cells, while epidermal stem cells are characterized by lack of Sox9 expression. Interestingly, basal cell carcinomas (BCC) overexpress Sox9, yet have been demonstrated to arise from epidermal and not hair follicle stem cells. Moreover, Sox9 is critical for BCC as genetic loss of Sox9 completely abolishes tumor formation in vivo. The mechanisms by which ectopic Sox9 contributes to tumor formation for BCC and other cancers remain to be elucidated. However, in HFs, Sox9 binds and regulates the key open chromatin domains (super-enhancers) that choreograph stem cell genes, and my preliminary studies indicate that ectopic expression of Sox9 in the adult epidermis shifts the chromatin landscape by activating previously silenced genes that may be important for BCC transformation. Therefore, I hypothesize that Sox9 elicits a fate switch in Epidermal stem cells by directly binding and remodeling chromatin at key enhancers. Utilizing a newly developed transgenic mouse that expresses an inducible Sox9 in the epidermal stem cells, I will map and annotate the chromatin and transcriptional changes that occur after induction of Sox9 to pinpoint deviations from Sox9-negative epidermal and Sox9+ hair follicle stem cells. Exploiting a combination of nucleosome binding experiments, immunoprecipitations and mass spectrometry, I will determine whether Sox9 can directly recognize and remodel heterochromatin, or whether it achieves this through the interactions of other proteins. Finally, using a powerful in utero lentiviral approach developed in the Fuchs lab, I will functionally test the results of candidates identified using CRISPR/CAS in the context of tumor formation and progression. My studies will directly further our knowledge regarding Sox9 mediated chromatin remodeling and subsequent activation of oncogenic and stem cell transcriptional pathways. Identifying the key Sox9 target genes as well as the proteins involved in epidermal to BCC reprogramming will lead to the development of novel therapeutics used to “reset” the tumor epigenetic landscape into a non-malignant form.