Molecular architecture of claudin-based tight junction strands and paracellular ion channels

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Tight junctions (TJ) regulate the paracellular permeability for solutes and water across epithelial and endothelial tissue barriers. The key structural determinants for this function are the tetra-helical transmembrane proteins of the claudin (CLDN) family. They form the backbone of intramembrane TJ strands and depending on the subtype either barriers or paracellular channels for small cations, anions or water. Progress in crystallization, structure prediction, MD simulations and functional analysis of claudins strongly advanced the understanding of TJ structure and function. However, the molecular architecture of TJ strands and paracellular channels is still under debate. Although claudin sequence determinants influencing assembly and function have been identified, the 3D-structural differences leading to the functional diversity of TJs (channels for cation, cation & water or anions or ion barriers) are unclear. Main goal of the project is to elucidate the detailed molecular architecture of TJ strands and paracellular channels including claudin subtype-dependent diversity. Previously, we established and successfully applied methods (a) to dissect the multistep process of TJ strand assembly that includes homo- and heterophilic, cis- and trans-interactions between claudins (b) to investigate the transport physiology of TJs and (c) to model and simulate the molecular dynamics of claudin oligomers in a membrane environment. Crucially for the TJ field, convergence of cell-physiological, nanoscopic and structural/simulation data on TJ strands is still largely missing. The general aim of this funding period is to achieve this convergence for claudins expressed in the gastrointestinal tract and/or the kidney. The specific aims are elucidation of the structural and mechanistic basis for (i) selectivity of CLDN10b/CLDN15-like cation channels and that of CLDN10a- and CLDN17 anion channels, (ii) barrier (CLDN1, -3, -5) versus channel formation, (iii) claudin intermixing and segregation into functionally distinct segments along TJ strands and (iv) disturbance of channel or barrier function of claudins by pathogenic mutations. The proposal bundles cell-physiological and structural bioinformatics expertises on classic claudins. Since TJ function is essential for all organs and disturbed in a multitude of barrier- and transport-related diseases affecting the intestine, kidney, glands or the brain, the project will provide key insights in the molecular physiology and pathophysiology of tissues barriers in general.