Structural analysis of integrin α5β1 and Tie2 activation and modulation

Abstract

The cardiovascular system constantly remodels to facilitate effective transport of nutrients, oxygen, and immune cells. Angiogenesis is the formation of new vasculature from preexisting vessels and is one of the major contributors to vessel expansion and homeostasis. Clinical and biochemical studies have significantly enhanced our understanding of angiogenic signaling pathways and the proteins involved in both healthy and diseased states such as chronic inflammation and cancer. This thesis investigated the structural mechanisms of integrin α5β1 and Tie2 receptors in angiogenic regulation. Integrins are heterodimeric receptors critical for cell adhesion and motility. Integrin α5β1 is involved in the migration of endothelial cells during angiogenesis. Dysregulation of integrin α5β1 is associated with tumor progression and metastasis. Despite numerous efforts, α5β1-targeting therapeutics have been unsuccessful due to poor efficacy and off-target effects. A contributing factor is our limited understanding of how integrin conformation influences interactions with therapeutics. Using cutting-edge cryogenic electron microscopy (cryoEM) techniques in combination with cell-based assays, I analyzed the impact of potential therapeutic molecules such as antibodies and de novo-designed proteins on integrin α5β1’s conformation. These insights highlight how therapeutic binding influences integrin α5β1 conformation and underscore the importance of considering these effects during the development of future integrin-targeted therapeutics. Tie2 is a receptor tyrosine kinase that facilitates tight cell-cell contacts, regulating vascular quiescence. It is regulated by a family of growth factors called angiopoietins, which cluster and activate Tie2. Though the angiogenic role of Tie2 is clear, little is known about the allosteric structural perturbations that Tie2 undergoes to propagate the angiopoietin-binding signal across the cell membrane. In addition, it has been demonstrated that integrin α5β1 can modulate the activity of Tie2 through receptor crosstalk. However, there is no defined mechanism for this interaction. I took advantage of modern structural tools to overcome these gaps using full-length integrin α5β1 and Tie2 proteins. These findings contribute to our understanding of the underlying mechanisms of receptor crosstalk between integrin α5β1 and Tie2.