权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

The Identity of the Notch Ankyrin Repeat Domain as a Determinant of Isoform-Specific Notch Signaling

Notch锚蛋白重复结构域作为异构体特异性Notch信号传导决定因素的身份

基本信息

批准号：
10152114
负责人：
Kristen Michelle Ramsey
金额：
$ 6.64万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-04-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10152114
关键词：
Animal Organ Ankyrin Repeat Architecture Binding Binding Sites Biochemical Biological Assay Biophysical Process Biophysics Breast Breast Cancer Cell C-terminal Calorimetry Cell Differentiation process Cell Nucleus Cell physiology Cells Characteristics Complex DNA DNA Binding DNA-Binding Proteins Data Development Dimerization Disease Engineering Event Foundations Genetic Transcription Head Hela Cells In Vitro Individual Integral Membrane Protein Left Ligand Binding Domain Link Literature Luciferases MCF7 cell Maps Matrigel Invasion Assay Measures Mediating Mediator of activation protein Membrane Monitor Mutagenesis N-terminal NMR Spectroscopy Nature Neoplasm Metastasis Notch Signaling Pathway Oncogenic Outcome Pathway interactions Play Process Protein Isoforms Reporting Research Resistance Role Series Side Signal Pathway Signal Transduction Site Source Structure Surface Testing Therapeutic Thermodynamics Titrations Transactivation Transcriptional Activation Transforming Growth Factor beta Tumor Cell Invasion Variant Work analytical ultracentrifugation animal tissue biophysical properties defined contribution design dimer epithelial to mesenchymal transition extracellular inhibitor/antagonist malignant breast neoplasm mutant notch protein promoter protein protein interaction receptor recruit targeted treatment therapeutic target tumor tumor progression

项目摘要

Project Summary/Abstract: The Notch signaling pathway is a crucial regulator of development and cell differentiation in almost all animal organs and tissues. The Notch receptor is a 300-kDa transmembrane protein consisting of an extracellular ligand-binding domain and an intracellular domain (NICD). The mammalian Notch pathways consists of four receptor isoforms which share a conserved architecture and overlapping but non-redundant functions. In fact, each isoform plays such a critical role in Notch function that congenital diseases due to severely deficient signaling by one isoform are exceedingly rare. The complexity encoded in the Notch pathway by the four receptor isoforms can be divided into two components: 1) isoform-specific differences in canonical Notch signaling outcomes, and 2) isoform-specific crosstalk with other signaling pathways. Upon receiving a signal, a series of proteolytic cleavages release the NICD from the membrane which, in the canonical pathway, translocates to the nucleus where it binds DNA-binding protein CSL and recruits co-activator Mastermind (MAML) to form the Notch transcriptional activation complex (NTC). Two domains of the NICD are responsible for forming the ternary NTC, the N-terminal disordered RAM linker and the ankyrin repeat domain (ANK) which forms a cleft with CSL for MAML binding. It has been suggested that isoform-specific outcomes in canonical signaling are mediated by variations in the ANK domains which are additionally responsible for mediating NTC dimerization on specialized promoters containing paired head-to-head CSL binding sites. However, these reports, to date, have been qualitative in nature and have offered inconsistent conclusions. Additionally, these studies did not investigate the role of the ANK domains in facilitating crosstalk with other pathways. Because ANK domains are ubiquitous protein-protein interaction motifs, it is unsurprising that many non-canonical NICD interactions are also mediated by the ANK domains. Two of the most common Notch-intersecting pathways are Wnt and TGFβ, and though this crosstalk is isoform-specific, the synergistic effects of Notch, Wnt, and TGFβ signaling have been implicated in numerous disease states, particularly in metastatic events and increased tumor invasion in breast cancer. Despite numerous attempts to therapeutically target Notch signaling, a lack of data on the mechanism governing isoform-specific Notch signaling has left these efforts unsuccessful. I propose three Aims to fill this void: I will 1) quantify the stability of each isoform’s NTC and their dimerization capacities, 2) characterize the binding of each ANK isoform with key non-canonical binding partners that crosstalk with Wnt and TGFβ, and 3) define the role of variations in the ANK domains on Wnt and TGFβ crosstalk in the cellular context of breast cancer. This work will integrate in vitro biophysical characterizations with functional analyses in cells to define the biophysical mechanism(s) and sequence basis of isoform-specific Notch signaling. These data represent a holistic view of Notch signaling that will serve as a critical foundation for designing more targeted, isoform-specific Notch therapeutics.

项目总结/文摘: