Probing Supramolecular Structure, Stoichiometry, and Trafficking in Live Cells of Oligomers of G-Protein Coupled receptors

University Of Wisconsin-Milwaukee, Milwaukee WI

Investigators

Abstract

Members of the large class of membrane receptor proteins called G-protein-coupled receptors (GPCRs) recognize and respond to a wide variety of signals that may arrive in the form of small ligand molecules such as neurotransmitters and hormones, and physical stimuli such as light and mechanical stress. The relatively recent observation that GPCRs form oligomeric complexes in vivo has led to many interesting questions related to the structure and functional role of the oligomers, including their size, distribution within the cell, and response to ligand binding. In this project, the Principal Investigator will develop new methodology based on advanced laser-scanning microscopy for measurements, in single live cells, of spectrally resolved Foerster Resonance Energy Transfer (FRET) a non-radiative transfer of energy from an excited fluorescent molecule to a non-excited one that resides nearby. The experimental procedures thus established will be used for probing non-invasively the temporal changes in the structure and cellular localization of oligomeric complexes of fluorescently-tagged GPCRs in the presence and absence of natural and artificial ligands. The methods developed as part of the proposed research will allow the PI to contribute to the understanding of the biology of GPCR-mediated signaling and its dysfunction in diseases or other medical disorders. The outcomes of this research are therefore expected to not only dramatically advance research but also to provide foundations for developing more efficient pharmaceutical agents. The research will be housed by the Physics Department and the Department of Biological Sciences at the University of Wisconsin-Milwaukee, and it will create an excellent environment for graduate and undergraduate students to practice an interdisciplinary approach to science, by using techniques and methods from a variety of sub-disciplines seamlessly integrated to serve a well-defined goal. Some of the experimental set-ups in the PIs laboratory will be used for demonstrations in his undergraduate- and graduate-level classes. The PI will also collaborate with a high-school teacher to stimulate the interest in science of high school students through small research projects conducted in the PI's laboratory as well as through the research projects brought by the teacher back to her class. This project is being supported jointly by the Physics of Living Systems Program in the Physics Division and by Biomolecular Dynamics, Structure and Function in MCB.