Clonal dynamics and single cell transcriptomics of prostate luminal epithelial cells in homeostasis and aging

项目总结 最常见的前列腺疾病、良性前列腺肥大和前列腺癌与 衰老，归因于荷尔蒙失衡和腺体动态平衡的丧失。考虑到对前列腺的重大影响 病理对男性的健康及其对医疗系统的重大负担，重要的是进一步 阐明正常动态平衡对非调控克隆生长造成的障碍。到目前为止，大多数研究都集中在 关于阐明不同干细胞前列腺癌群体的身份并验证其干细胞潜能 培养化验。然而，对成人的细胞和种群动态的精确理解 前列腺动态平衡，以及它的精细平衡如何随着年龄的增长而变化，并对年龄相关的细胞做出贡献 过度增殖，到目前为止还没有出现。这一领域的进展将对确定 老年组织中去调节的前列腺群和设计预防性治疗。在初步研究中， 利用体内谱系追踪和单细胞转录组学，我发现了一种“腔中间”转录。 具有独特的Wnt/p63信号的状态(LUMI)，它构成了癌细胞群体的很大一部分 在肿瘤形成的早期阶段扩张。 在F99阶段的提案中，我将研究LUMI细胞状态作为癌症的首选起源细胞 由Wnt信号和p63控制的模型和潜在的生长计划。具体地说，我将聘用 用功能器法研究Wnt信号在体外和体外促进LUMI细胞生长中的作用 说明体内消除Wnt/p63信号的作用。在提案的K00阶段，我将利用 我的方法和生成新的小鼠模型来描绘细胞水平的异质性和生长模式 并在成人和老年前列腺腔内层探讨其潜在的分子调控机制。 具体地说，我将利用创新的数学建模来整合从基因获得的克隆数据 谱系追踪以描述前列腺细胞所采用的成年和衰老的动态平衡生长模式。这就做 还建立了一种新的小鼠模型：Nkx3.1CreERT2/+-pletti-Carlin小鼠，用于体内同步谱系 跟踪和条形码，这将提供更高的世系祖先分辨率。单细胞RNAseq和单细胞 细胞ATACseq将被用来评估细胞状态并揭示前列腺的动态分子异质性 血统追踪小鼠模型中的管腔细胞。K00 Aim将提供一个全面的功能图 成人和老年前列腺中的管腔前列腺细胞及其相关的谱系和MASTER调节基因 用流明克隆人。 综上所述，我的研究将提供一个全面的了解人口增长的动态 前列腺组织的管腔层，并可能导致识别与年龄相关的新的治疗靶点 前列腺增生性疾病。

PROJECT SUMMARY The most common prostate disorders, benign prostate enlargement and prostate cancer are associated with aging and attributed to hormonal imbalances and loss of glandular homeostasis. Given the major impact prostate pathologies have on men’s health and their significant burden on healthcare systems, it is important to further elucidate the barriers posed by normal homeostasis to deregulated clonal growth. To date, most studies focused on elucidating the identity of various stem cell prostate populations and on validating their stem cell potential in culture assays. However, a precise understanding of the cellular and population dynamics at play in adult prostate homeostasis, and how its fine balances change with aging and contribute to age-related cellular hyperproliferations, has not emerged so far. Advances in this area would be extremely beneficial for identifying deregulated prostate populations in aged tissues and for designing prophylactic therapies. In preliminary studies, using in vivo lineage-tracing and single cell transcriptomics, I uncovered a “luminal intermediate” transcriptional state (LumI) with unique Wnt/p63 signaling which constitutes a great portion of the cancer cell populations expanding in early stages of tumorigenesis. In the F99 phase of the proposal, I will investigate the LumI cell state as the preferred cell of origin in cancer models and the underlying growth program controlled by Wnt signaling and p63. Specifically, I will employ functional organoid assays to investigate the role of Wnt signaling in promoting LumI cell growth in vitro and delineate the effects of eliminating the Wnt/p63 signaling in vivo. In the K00 phase of the proposal, I will leverage my methods and generate new mouse models to delineate the cellular level heterogeneity and growth patterns in adult and aged prostate luminal layer and explore the underlying molecular regulatory mechanisms. Specifically, I will utilize innovative mathematical modeling to integrate the clonal data obtained from genetic lineage tracing to characterize the modes of adult and aging homeostatic growth adopted by prostate cells. I will also generate a novel mouse model: Nkx3.1CreERT2/+-Confetti-CARLIN mouse for simultaneous in vivo lineage tracing and barcoding which will provide increased lineage ancestry resolution. Single cell RNAseq and Single cell ATACseq will be used to assess cellular states and uncover the dynamic molecular heterogeneity of prostate luminal cells in the lineage tracing mouse model. The K00 Aim will provide a comprehensive functional map of luminal prostate cells in adult and aging prostate and identify lineages and master regulator genes associated with luminal clones. Taken together, my studies will provide a comprehensive understanding of population growth dynamics in the luminal layer of prostate tissue and may lead to identification of novel therapeutic targets for age-related prostate hyperproliferative disorders.