Molecular Mechanisms of Cell Fate Decisions in Hair Follicle Stem Cells

描述(由申请人提供)：破译成体干细胞(SC)命运决定的分子机制对于了解疾病和组织再生治疗至关重要。这项建议的总体目标是利用毛囊和皮肤作为模型系统，在分子水平上解决转录因子和表观遗传调控在命运选择中的相互作用。 脊椎动物成体干细胞种群在其天然组织中的决定。因为头发 毛囊干细胞在小鼠皮肤中同步增殖并恢复静止， 这个系统允许机械性的洞察，否则就更难实现。在AIM (1)我们计划更深入地调查更多休眠与 毛囊干细胞利基中更活跃的细胞亚群，通过表征他们的 细胞培养和细胞移植实验中的形态发生和自我更新能力。 干细胞生境中这两种细胞亚群的存在似乎是 对于至少几种脊椎动物组织来说，这是一个相对常见的特征。它们之间的相互作用 维持组织内环境稳定对于准确描述干细胞很重要 行为和命运的决定。在AIM(2)中，我们将检查组蛋白标记的分布 通过染色质免疫沉淀跨越基因组中干细胞活动的两个阶段 和测序，并试图检查两个已知的成人组织之间的潜在联系 干细胞特征：静止性和可塑性。在AIM(3)中，我们将雇用两名 转录因子RUNX1和GATA6作为命运获得调控因子。我们将测试他们的 通过与组蛋白表观基因组的相互作用重塑干细胞组蛋白表观基因组的潜在意义 我们发现一组组蛋白修饰酶在毛囊中表达 相关模式。我们将在小鼠皮肤中诱导RUNX1和GATA6的急性丢失，并 检测特异组蛋白修饰酶的mRNA表达水平 相关毛囊细胞群。此外，我们还将进行免疫共沉淀 或者下拉实验来检查潜在的蛋白质-蛋白质直接相互作用 在RUNX1或GATA6和特定的组蛋白修饰酶之间。这将开始 解决脊椎动物成体干细胞种群命运获得的机制 并阐明了作为主控的转录因子之间的相互作用 调控组蛋白表观遗传标记的细胞命运和全球重塑。 与公共卫生相关：许多疾病是由于对细胞命运的错误调控导致的，成体组织干细胞在其活动期间获得了维持正常组织稳态的能力。最近，针对特定的组蛋白修饰酶的新型药物疗法的开发前景很大，其中一些已经在临床上出现。这项建议试图利用一个通用的脊椎动物模型系统，小鼠皮肤和毛囊，以更好地了解组蛋白表观遗传标记和控制成年脊椎动物干细胞细胞命运的转录因子之间的相互作用。

DESCRIPTION (provided by applicant): Deciphering the molecular mechanisms of adult stem cell (SC) fate decisions is essential for understanding disease and for tissue regenerative therapy. The overall goal of this proposal is to utilize hair follicles and skin as model systems to address at the molecular level the interplay of transcriptional factors and epigenetic regulation in fate choice decisions of a vertebrate adult stem cell population in its native tissue. Because hair follicle stem cells synchronously proliferate and return to quiescence in the mouse skin, this system allows mechanistic insight that is otherwise more difficult to achieve. In Aim (1) we plan to investigate in more depth the relationship between more dormant versus more active cell subpopulations in the hair follicle stem cell niche, by characterizing their morphogenetic and self-renewal ability in cell culture and cell transplantation assays. The existence of these two kinds of cell subpopulations in stem cell niches appears to be a relatively common feature for at least several vertebrate tissues. Their interplay in maintaining tissue homeostasis is important for accurate description of stem cell behavior and fate decisions. In Aim (2) we will examine the distribution of histone marks across the genome at two stages of stem cell activity by chromatin immuno-precipitation and sequencing, and attempt to examine a potential link between two known adult tissue stem cells features: quiescence and plasticity. In Aim (3) we will employ two transcription factors Runx1 and Gata6 as fate acquisition regulators. We will test their potential implication in remodeling the stem cell histone epigenome via interactions with a battery of histone modifying enzymes we found expressed in the hair follicle in a relevant pattern. We will induce acute loss of Runx1 and Gata6 in mouse skin and examine the mRNA expression level of specific histone modifying enzymes in the relevant hair follicle cell populations. In addition we will perform co-immunoprecipitation or pull-down experiments to examine a potential direct protein-protein interaction between Runx1 or Gata6 and specific histone modifying enzymes. This will begin to address the mechanisms of fate acquisition in a vertebrate adult stem cell population and shed light on the interplay between transcription factors that work as master regulator in orchestrating cell fate and global remodeling of histone epigenetic marks. PUBLIC HEALTH RELEVANCE: A large number of diseases are due to miss-regulation of cell fate acquisition of adult tissue stem cells during their activity to maintain normal tissue homeostasis. There has been recently great promise for developing novel drug therapeutics that target specific histone modifying enzymes, some already present in clinics. This proposal seeks to utilize a versatile vertebrate model system, mouse skin and hair follicles, to gain better understanding of the interplay between histone epigenetic marks and transcription factors that control cell fate decisions in adult vertebrate stem cells.