Function and Mechanism of the Hippo-PI5P4K Axis for Growth Control

项目摘要 我们的长期目标是破译PI 5 P4 K驱动生长的分子机制，并阐明 它们的失调是如何导致乳腺肿瘤发生的。在本建议中，我们将着手提供一个 全面了解一个新的信号网络，这是至关重要的整合代谢信息， 确定相关蛋白质和途径如何相互作用以维持细胞内稳态。 磷脂酰肌醇-5-磷酸4-激酶（PI 5 P4 Ks）是一类非经典的磷酸肌醇激酶 我们已经证明这对保持养分供应和燃料生长至关重要，但它们是如何被调节的 他们自己在很大程度上是未知的。通过全面的体外激酶筛选，我们发现了 Hippo途径的激酶，MST 1和MST 2作为PI 5 P4 Ks以及脂质的直接负调节剂， PI-5-P与Hippo衔接蛋白MOB 1相互作用，进而影响活性 河马下游效应子雅普。众所周知，河马通路对发育、生长 和器官发生，并且该途径的失调导致肿瘤进展。 虽然PI 5 P4 Ks与肿瘤生长有关，但人们对PI 5 P4 Ks的调控机制知之甚少。 此外，PI 5 P4 Ks如何在生理条件下实际上命令细胞生长， 病理学背景如乳腺癌是不完整的。为了回答这些重要的问题，我们将 以下目标： 在目标1中，我们将使用非常可行的生物物理学和结构生物学来确定 PI-5-P和MOB 1之间界面以及定量PI-5-P如何调节MOB 1与其它Hippo途径的结合 体外成分。 在目的2中，我们将研究PI 5 P4 Ks的活性如何控制MOB-LATS复合物的活化 MST 1/2下游，探索PI 5 P4 K致瘤特性的一种潜在机制。 在目标3中，我们将使用一组P15 P4 K-Hippo轴来确定乳腺癌中P15 P4 K-Hippo轴的生物学意义。 TNBC细胞系、新型小鼠乳腺肿瘤模型和患者乳腺肿瘤样品。 该项目旨在将联合收割机定量结构生物学与分子细胞遗传学相结合，以破译PI-5- P调节乳腺癌细胞生长的Hippo途径。鉴于PI 5 P4 K是令人兴奋的“可药物化”目标， 对于乳腺癌，表征PI 5 P4 Ks如何连接到Hippo通路揭示了一种可能的策略， 未来的抗癌疗法

PROJECT SUMMARY Our long-term goal is to decipher the molecular mechanisms underlying PI5P4K-driven growth and to elucidate how their dysregulation contributes to breast tumorigenesis. In this proposal, we will set out to provide a comprehensive understanding of a new signaling network that is critical to integrate metabolic information and determine how the related proteins and pathways interact to maintain cellular homeostasis. Phosphatidylinositol-5-phosphate 4-kinases (PI5P4Ks) are a family of non-canonical phosphoinositide kinases which we have shown to be critical for preserving nutrient availability and fuel growth yet how they are regulated themselves is largely unknown. Using a comprehensive in vitro kinase screen, we have discovered the core kinases of the Hippo pathway, MST1 and MST2 as direct negative regulators of the PI5P4Ks as well as the lipid substrate of the PI5P4Ks, PI-5-P to interact with the Hippo adaptor protein MOB1 which in turn effects the activity of the Hippo downstream effector YAP. The Hippo pathway is known to be important for development, growth and organogenesis, and dysregulation of this pathway leads to tumor progression. While PI5P4Ks have been linked to tumor growth, exactly how the PI5P4Ks are regulated is poorly understood. Furthermore, how the PI5P4Ks actually command cell growth under both physiological conditions and pathological contexts such as breast cancer is incomplete. To answer these important questions, we will perform the following Aims: In Aim 1 we will use very feasible biophysical and structural biology to determine atomic-resolution details of the interface between PI-5-P and MOB1 and quantitate how PI-5-P regulates MOB1 binding to other Hippo pathway components in vitro. In Aim 2 we will investigate how the activity of the PI5P4Ks control the activation of the MOB-LATS complex downstream of MST1/2, exploring one potential mechanism for PI5P4K tumorigenic properties. In Aim 3 we will establish the biological significance of the PI5P4K-Hippo axis in breast cancer using a panel of TNBC cell lines, novel mouse breast tumor models, and patient breast tumor samples. This project seeks to combine quantitative structural biology with molecular cell genetics to decipher how PI-5- P regulates the Hippo pathway for breast cancer cell growth. Given that PI5P4Ks are exciting ‘druggable’ targets for breast cancer, characterizing how PI5P4Ks connect to the Hippo pathway reveals a possible strategy for future anti-cancer therapies.