Spatiotemporal Modulation of PIP[3]Signaling in Prostatic Tubulogenesis

描述（由申请人提供）：本提案描述了一名具有泌尿病理学临床专业知识的医生-科学家的五年职业发展计划，该医生-科学家致力于建立一个专注于前列腺上皮形态发生和迁移的独立实验室。她将通过发育生物学，细胞迁移和尖端活细胞成像技术的课程来增加她的科学培训。这项研究将在细胞动力学中心进行，这是约翰霍普金斯医学院最先进的多学科中心，汇集了研究活细胞中空间和时间调节分子事件的专家。拟议的研究项目将侧重于阐明驱动前列腺上皮形态发生的信号通路，并可能在前列腺生长异常的成人疾病中重新激活。该项目将汇集她的两位科学导师的优势，Denise Montell博士，上皮细胞迁移基础科学的国际专家，以及大卫伯曼博士，泌尿病理学家，实验室专注于前列腺发育和疾病。尽管PIP 3（磷脂酰肌醇[3，4，5]-三磷酸）信号在调节细胞极性中的作用高度保守，但PIP 3信号在哺乳动物上皮形态发生中的作用仍然未知。基于初步的数据，我们假设PIP 3信号的时空调制是至关重要的两个关键过程驱动前列腺小管：前列腺导管的伸长和apicobasal极性规范。利用我们实验室开发的3D前列腺上皮培养系统，以及体外前列腺器官培养和最先进的延时成像技术，我们将在三个相互关联的目的中检查PIP 3调节对前列腺上皮形态发生的影响：第一章我们将使用转基因小鼠建立前列腺导管延长过程中PIP 3信号传导的细胞内定位和需求。PIP 3生物传感器和亚型特异性PI 3 K（磷酸肌醇-3-激酶）抑制剂; 2）我们将通过检查PI 3 K抑制对前列腺上皮迁移、增殖和凋亡的特异性作用来确定PIP 3信号传导调节前列腺导管伸长的细胞机制; 3）我们将研究PIP 3通过调节PI 3 K和PTEN的活性和水平在调节顶基底极性中的作用。（磷酸酶和张力蛋白同源物）在体外腺泡形态发生和体内使用一种新的转基因小鼠系在我们的实验室建立。通过阐明在发育过程中驱动前列腺生长的信号网络，我们将更好地了解这些途径的失调如何导致成人前列腺疾病。

DESCRIPTION (provided by applicant): This proposal describes the five-year career development plan of a physician-scientist with clinical expertise in urologic pathology who is committed to establishing an independent laboratory focusing on prostatic epithelial morphogenesis and migration. She will add to her scientific training with coursework in developmental biology, cellular migration, and cutting-edge live-cell imaging techniques. The research will take place in the Center for Cell Dynamics, a state-of-the-art multidisciplinary center at the Johns Hopkins School of Medicine that brings together experts studying spatially and temporally regulated molecular events in living cells. The proposed research project will focus on elucidating the signaling pathways that drive prostatic epithelial morphogenesis and are potentially re-activated in adult diseases of abnormal prostate growth. This project will bring together the strengths of her two scientific mentors, Dr. Denise Montell, an international expert in the basic science of epithelial cell migration, and Dr. David Berman, a urologic pathologist with a laboratory focused on prostate development and disease. Despite the highly conserved role of PIP3 (phosphatidylinositol [3,4,5]- trisphosphate) signaling in regulating cellular polarity, the role of PIP3 signaling in mammalian epithelial morphogenesis remains unknown. Based on preliminary data, we hypothesize that spatiotemporal modulation of PIP3 signaling is critical for two key processes driving prostatic tubulogenesis: prostatic ductal elongation and apicobasal polarity specification. Taking advantage of a 3D prostate epithelial culture system developed in our laboratory, as well as in vitro prostate organ culture and state-of-the-art time-lapse imaging techniques, we will examine the effects of PIP3 modulation on prostatic epithelial morphogenesis in three inter-related Aims: 1)We will establish the intracellular localization of and requirement for PIP3 signaling during prostatic ductal elongation using mice transgenic for a PIP3 biosensor and isoform-specific PI3K (phosphoinositide-3-kinase) inhibitors; 2)We will define the cellular mechanism by which PIP3 signaling regulates prostatic ductal elongation by examining the specific effects of PI3K inhibition on prostatic epithelial migration, proliferation and apoptosis; 3)We will examine the role of PIP3 in regulating apicobasal polarity by modulating the activity and levels of PI3K and PTEN (phosphatase and tensin homologue) in vitro during acinar morphogenesis and in vivo using a novel transgenic mouse line established in our laboratory. By elucidating the signaling networks that drive prostatic growth during development, we will better understand how dysregulation of these pathways may lead to prostatic disease in the adult.