Mechanisms of transmembrane signalling by tetraspanins
Lead Research Organisation:
University of Birmingham
Department Name: Medical Sciences - Medicine
Abstract
In this project the structures and interactions of the human tetraspanin proteins will be investigated by heteronuclear magnetic resonance spectroscopy. We have discovered that tetraspanin C-termini are bound by the tandem PDZ proteins of syntenin, and intend to elucidate the structural basis of these interactions in order to understand how they influence receptor signaling and endocytosis. The novel regulatory effects of PDZ domain phosphorylation and stabilizing terminal extensions will be investigated in order to build a model of the signaling mechanism. The extracellular domain has been expressed for NMR analysis of its lipid interaction sites and protein docking studies, providing a basis for defining its function as an cell surface receptor. Tetraspanins have been expressed as functionally intact full length proteins, and production will be scaled up to analyse their structural and binding properties in vitro. In particular, we seek to determine the oligomeric state and structural properties of the proteins in the free and ligand bound states using mixed micelles to solubilize the intact receptor. Together with collaborative in vivo cell biological studies this will provide a better understanding of the structural mechanism of tetraspanin signaling.
Technical Summary
Tetraspanins are an extensive family of under-characterized integral membrane proteins. Each family member contains four transmembrane regions, two extracellular domains, and variable cytoplasmic termini. Their total size is 23 ? 40 kDa. These structural domains determine the specific interactions of the tetraspanin with ligand proteins including immune receptors and viral envelope proteins, as well as with phospholipids, glycolipids and cholesterol within transmembrane-enriched microdomains. The structural basis of these interactions is unclear, limiting our mechanistic understanding of tetraspanin functions and precluding rational drug design.
Tetraspanins are found in mammals, plants, fungi and protozoa. Humans contain 33 tetraspanins, which are generally involved in cell adhesion, migration and fusion, and cellular activation and signaling. They are found in essentially all tissues, with each cell type expressing several tetraspanins. Biological roles of mammalian tetraspanins include contributions to kidney, skin and platelet function, sperm?egg fusion, nervous system development, monocyte fusion and immune cell proliferation, although their molecular mechanisms remain poorly understood.
In this project the structures and interactions of the human tetraspanins will be investigated by heteronuclear magnetic resonance spectroscopy. We have discovered that tetraspanin C-termini are directly and specifically bound by tandem PDZ proteins, and intend to elucidate the structural basis of these interactions in order to understand how they influence receptor signaling and endocytosis. The novel regulatory effects of PDZ domain phosphorylation and stabilizing terminal extensions will be investigated in order to build a model of the signaling mechanism. The extracellular domain has been expressed for NMR analysis of its lipid interaction sites and protein docking studies, providing a basis for defining its function as a cell surface receptor. Finally two tetraspanins have been expressed as functionally intact full length proteins, and production will be scaled up to analyse their structural and binding properties in vitro. In particular, we seek to determine the oligomeric state and structural properties of the proteins in the free and ligand bound states using mixed micelles to solubilize the intact receptor. Together with collaborative in vivo cell localization and endocytic studies this will provide a comprehensive structural mechanism of tetraspanin signaling.
Tetraspanins are found in mammals, plants, fungi and protozoa. Humans contain 33 tetraspanins, which are generally involved in cell adhesion, migration and fusion, and cellular activation and signaling. They are found in essentially all tissues, with each cell type expressing several tetraspanins. Biological roles of mammalian tetraspanins include contributions to kidney, skin and platelet function, sperm?egg fusion, nervous system development, monocyte fusion and immune cell proliferation, although their molecular mechanisms remain poorly understood.
In this project the structures and interactions of the human tetraspanins will be investigated by heteronuclear magnetic resonance spectroscopy. We have discovered that tetraspanin C-termini are directly and specifically bound by tandem PDZ proteins, and intend to elucidate the structural basis of these interactions in order to understand how they influence receptor signaling and endocytosis. The novel regulatory effects of PDZ domain phosphorylation and stabilizing terminal extensions will be investigated in order to build a model of the signaling mechanism. The extracellular domain has been expressed for NMR analysis of its lipid interaction sites and protein docking studies, providing a basis for defining its function as a cell surface receptor. Finally two tetraspanins have been expressed as functionally intact full length proteins, and production will be scaled up to analyse their structural and binding properties in vitro. In particular, we seek to determine the oligomeric state and structural properties of the proteins in the free and ligand bound states using mixed micelles to solubilize the intact receptor. Together with collaborative in vivo cell localization and endocytic studies this will provide a comprehensive structural mechanism of tetraspanin signaling.
