Understanding how receptor tyrosine kinase activation dynamics specify proliferative cellular responses

了解受体酪氨酸激酶激活动力学如何指定增殖细胞反应

• 批准号: 10400914
• 负责人: Mark A Lemmon
• 金额: $ 51.82万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10400914
• 关键词: Address; Affinity; Agonist; Biochemical; Biological Assay; Biophysics; Breast Cancer cell line; Cell Proliferation; Cells; Cessation of life; Chimera organism; Data; Dimerization; Endocytosis; Epidermal Growth Factor; Epidermal Growth Factor Receptor; Epiregulin; ErbB4 gene; Feedback; Future; G-Protein-Coupled Receptors; Goals; Image; Kinetics; Ligands; MAP Kinase Modules; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Methods; Microscopy; Motivation; Mutation; Nature; Nerve Growth Factors; Neuregulins; Neurotrophic Tyrosine Kinase Receptor Type 1; Oncogenic; Outcome; PC12 Cells; Pathway interactions; Pharmacology; Phosphorylation Site; Play; Property; Receptor Activation; Receptor Protein-Tyrosine Kinases; Receptor Signaling; Resistance; Role; Signal Pathway; Signal Transduction; Specific qualifier value; Specificity; T-Cell Receptor; Testing; Therapeutic; Transducers; Work; base; cancer cell; design; dimer; epigen; insight; molecular imaging; network models; receptor; response; single molecule; targeted treatment; therapeutic target; trafficking

Project Summary We propose to combine biophysical imaging and biochemical approaches to address two key unanswered questions in the field of signaling by receptor tyrosine kinases (RTKs) – a principal class of therapeutic targets in cancer where resistance to current therapies necessitates new pharmacological approaches: a. What defines the distinct set of responses, and cell fate, downstream of a particular RTK? b. How can different ligands for a given RTK promote distinct cellular responses? It is well known that epidermal growth factor (EGF) and nerve growth factor (NGF) promote proliferative and differentiative responses respectively in PC12 cells, through EGFR and TrkA, apparently using the same set of signaling pathways. Early studies showed that Erk activation kinetics plays a key role in determining the different cell fates induced by these ligands, with transient Erk activation being associated with proliferation and sustained Erk activation with differentiation. Rather than being defined solely by different feedback ‘wiring’ in the intracellular MAP kinase cascade, we and others have found that the RTK activation kinetics play a direct role in defining the dynamic properties of the signaling network. Several recent studies further argue that the strength of ligand-induced RTK dimers (and/or their lifetime) dictates signaling specificity, offering the possibility of dynamically-determined biased agonism in RTK signaling. To gain insight into the mechanistic basis for biased agonism in RTK signaling, we propose to elucidate how activating the same RTK intracellular region in different ways can result in dramatically different cellular responses (proliferation vs differentiation). Our previous structural, biophysical, and biochemical data suggest the hypothesis that signaling outcome is determined by RTK dimer stability and dynamics. Using intact and chimeric receptors, we will test this hypothesis by asking how altering RTK dimerization kinetics influences receptor endocytosis, post-endocytic trafficking, and dynamics of the downstream signaling network. We combine single-molecule imaging and microscopy studies of RTK trafficking with mass spectrometry and biochemical studies of downstream signaling networks to yield an integrated picture of this. Our primary motivation is to investigate how modifying RTK signaling dynamics (rather than simply inhibiting RTKs) might be used in future therapeutic approaches in cancer. Our Specific Aims address the following questions: 1 How can the same RTK intracellular region elicit orthogonal cellular responses depending on how it is activated? 2 What are the lifetimes of ErbB4/HER4 dimers induced by different neuregulin (NRG) ligands, and what is the basis for their sustained signaling? Our overall goal is to understand how dimerization dynamics can define signaling specificity, possibly through a kinetic proofreading similar to that seen in the T-cell receptor.

项目摘要 我们建议结合生物物理成像和生化方法来解决两个尚未回答的关键问题 受体酪氨酸激酶(RTK)信号传导领域的问题--一类主要的治疗靶点 在对现有疗法产生耐药性的癌症中，需要新的药理学方法： A.是什么定义了特定RTK下游的一组不同的反应和细胞命运？ B.特定RTK的不同配体如何促进不同的细胞反应？ 众所周知，表皮生长因子(EGF)和神经生长因子(NGF)促进增殖和 分别通过EGFR和TrkA在PC12细胞中的分化反应，显然使用相同的一组 信号通路。早期的研究表明，ERK的激活动力学在决定 这些配体诱导不同的细胞命运，瞬时的ERK激活与增殖相关 分化过程中ERK的持续激活。而不是仅仅由不同的反馈“连接”来定义 在细胞内的MAPK级联中，我们和其他人发现rtk激活动力学在 在定义信令网络的动态属性方面发挥直接作用。最近的几项研究进一步认为， 配体诱导的RTK二聚体的强度(和/或其寿命)决定了信号的特异性，提供了 RTK信号中动态确定的偏向激动型的可能性。 为了深入了解RTK信号通路中偏向激动化的机制基础，我们建议阐明 如何以不同的方式激活相同的RTK胞内区可以导致显著不同的细胞 反应(增殖与分化)。我们之前的结构、生物物理和生化数据表明 信号转导结果由RTK二聚体的稳定性和动力学决定的假设。使用完整的和 嵌合受体，我们将通过询问改变RTK二聚化动力学如何影响这一假设来检验这一假说 受体内吞作用、内吞后转运和下游信号网络的动力学。我们 将RTK转运的单分子成像和显微研究与质谱学和 下游信令网络的生物化学研究，以得出这一综合图景。我们的初选 动机是研究修改RTK信号动力学(而不是简单地抑制RTK)可能会如何 将在未来的癌症治疗方法中使用。我们的具体目标是解决以下问题： 1相同的RTK胞内区如何根据其方式引发正交的细胞反应 激活了吗？ 2不同的神经调节蛋白(NRG)配体诱导的ErbB4/HER4二聚体的寿命是多少？ 他们持续发出信号的基础是什么？ 我们的总体目标是了解二聚化动力学如何定义信号特异性，可能是通过 一种类似于T细胞受体的动态校对。