Project Summary/Abstract Overview of Research: The Van Horn lab primarily focuses on elucidating and understanding the molecular mechanisms that underlie membrane protein function in health and disease. To achieve these goals, the laboratory employs a modern state-of-the-art and interdisciplinary approach using biophysical, structural, computational, and functional techniques. We are experts in membrane protein biophysics and make use of advanced NMR studies combined with functional whole-cell patch-clamp electrophysiology. These orthogonal data are linked with Rosetta-based computational techniques to understand protein function. Our primary target is the TRPM8 ion channel which was initially identified as an oncogene that is prognostic for some types of cancer progression. More recently, it has become a focus for therapeutic intervention in pain and obesity. Complicating the potential application of TRPM8 therapies is that it is a molecular integrator that is activated by a number of diverse stimuli. For example, TRPM8 is the primary human cold sensor but is also activated by the chemical menthol from mint, both of which activate TRPM8 signaling networks. The ability to respond to several different stimuli in a polymodal manner makes TRPM8 studies crucial to delineate the independence and interdependence of molecular mechanisms that result in biological function and complicate its therapeutic targeting. In addition to direct stimulation by cold and menthol, TRPM8 is regulated by diverse proteins, including the membrane protein, PIRT, which in turn modulates TRPM8 activation by cold, menthol, and other stimuli. Beyond our research on direct activation of TRPM8 by cold and menthol, we focus on determining the mechanisms whereby PIRT modulates TRPM8 function. This has led to a number of contributions from our lab including, biophysical and structural characterization of TRPM8 and PIRT, TRPM8–PIRT complex stoichiometry, identification of species-dependent regulation, and central insight into molecular regulatory mechanisms. These efforts have led to strong scientific output, including publications, seminars, and patents. Five-year Goals: Broadly defined, we will identify how TRPM8 is directly activated by cold temperatures and other stimuli, map the allosteric networks that allow for polymodal function, and determine structures of TRPM8 and related membrane protein complexes of functional consequence. Research Vision: In the past 8 years, there has been an explosion of TRP channel structural biology with now ~100 discrete TRP channel structures. This represents tremendous development and output. Our research seeks to extend and complement the structural momentum to delineate fundamental mechanistic properties such as allostery, dynamics, and protein complex regulation that determines function. These TRPM8 outcomes are anticipated to have direct impacts on human health and disease but also to serve as a template that defines and identifies fundamen...