The Role of Cholesterol in Cell Signaling
thesisposted on 2015-10-21, 00:00 authored by Ewa M. Stec
Membrane lipids regulate a wide range of biological processes, including cell signaling. Cholesterol is a major lipid component of the mammalian plasma membrane. Although the metabolism and transport of cholesterol and its role in cardiovascular disease have been extensively investigated, the exact cellular function of cholesterol is yet to be fully explored. Cholesterol plays a pivotal role in the stability and architecture of the plasma membrane, most notably the formation of cholesterol-rich membrane microdomains, and structural and functional modulation of integral proteins embedded in plasma membrane. The role of cholesterol in the plasma membranes extends beyond the modulation of the fluidity and permeability of the bilayer. Accumulating evidence suggests that changes in cellular cholesterol levels in the plasma membrane modulate functionality of proteins involved in signaling pathways. However, the direct involvement of cholesterol in cellular activities through specific interactions with cellular proteins has remained unclear. Also, quantitative information about its cellular spatiotemporal dynamics and its local concentration changes under pathophysiological conditions is surprisingly scarce, making it difficult to elucidate how potential changes in the local concentrations of cholesterol may affect cellular processes in health and disease. Our laboratory has recently made two breakthroughs that should greatly help address these important questions. First, we discovered that cholesterol specifically interacts with various cytosolic scaffold proteins containing PDZ (PSD95, DLG1, and ZO1) domains and regulate their diverse cellular signaling activities. This important finding not only demonstrates that cholesterol can directly interact with major cellular regulatory proteins but also offers excellent systems to investigate the direct correlation between membrane cholesterol levels and cell cellular activities. Second, we develop a new fluorescence imaging technology for accurate in situ quantification of cholesterol in a spatiotemporally resolved manner under physiological and patho-physiological conditions. The specific and sensitive cholesterol quantification was achieved by engineered sensor derived from the D4 domain of perfringolysin O toxin and favorable spectral and membrane-binding properties of DAN and Nile Red 3 probes. This simultaneous quantification of cholesterol provides us with physiologically important data and new insight unattainable by conventional methodologies. Collectively, our discovery of a new class of cholesterol binding proteins and our new quantitative cholesterol imaging technology represent an important technical advance toward understanding of complex cholesterol-mediated cell regulation.