Structural model and functional significance of pH-dependent talin-actin binding for focal adhesion remodeling

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 105; no. 38; pp. 14436 - 14441
• Main Authors: Srivastava, J, Barreiro, G, Groscurth, S, Gingras, A.R, Goult, B.T, Critchley, D.R, Kelly, M.J.S, Jacobson, M.P, Barber, D.L
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 23.09.2008; National Acad Sciences
• Subjects: Actins; Actins - chemistry; Actins - metabolism; Amino acids; Animals; Binding sites; Biological Sciences; Cell adhesion & migration; Cell Line; Cell motility; Chemical equilibrium; Computer Simulation; Focal adhesions; Focal Adhesions - metabolism; Hydrogen-Ion Concentration; Mice; Microfilaments; Models, Molecular; Molecular structure; Mutation; NMR; Nuclear magnetic resonance; Nuclear Magnetic Resonance, Biomolecular; Physiological regulation; Protein Binding; Protein Structure, Tertiary; Proteins; Renovations; Sensors; Talin - chemistry; Talin - genetics; Talin - metabolism
• DOI: 10.1073/pnas.0805163105
• ISSN: 0027-8424

Actin filament binding by the focal adhesion (FA)-associated protein talin stabilizes cell-substrate adhesions and is thought to be rate-limiting in cell migration. Although F-actin binding by talin is known to be pH-sensitive in vitro, with lower affinity at higher pH, the functional significance of this pH dependence is unknown. Because increased intracellular pH (pHi) promotes cell migration and is a hallmark of metastatic carcinomas, we asked whether it increases FA remodeling through lower-affinity talin-actin binding. Talin contains several actin binding sites, but we found that only the COOH-terminal USH-I/LWEQ module showed pH-dependent actin binding, with lower affinity and decreased maximal binding at higher pH. Molecular dynamics simulations and NMR of this module revealed a structural mechanism for pH-dependent actin binding. A cluster of titratable amino acids with upshifted pKa values, including His-2418, was identified at one end of the five-helix bundle distal from the actin binding site. Protonation of His-2418 induces changes in the conformation and dynamics of the remote actin binding site. Structural analyses of a mutant talin-H2418F at pH 6.0 and 8.0 suggested changes different from the WT protein, and we confirmed that actin binding by talin-H2418F was relatively pH-insensitive. In motile fibroblasts, increasing pHi decreased FA lifetime and increased the migratory rate. However, expression of talin-H2418F increased lifetime 2-fold and decreased the migratory rate. These data identify a molecular mechanism for pH-sensitive actin binding by talin and suggest that FA turnover is pH-dependent and in part mediated by pH-dependent affinity of talin for binding actin.