Enhancing integrin α1 inserted (I) domain affinity to ligand potentiates integrin α1β1-mediated down-regulation of collagen synthesis.

Shi M, Pedchenko V, Greer BH, Van Horn WD, Santoro SA, Sanders CR, Hudson BG, Eichman BF, Zent R, Pozzi A
J Biol Chem. 2012 287 (42): 35139-35152

PMID: 22888006 · PMCID: PMC3471707 · DOI:10.1074/jbc.M112.358648

Integrin α1β1 binding to collagen IV, which is mediated by the α1-inserted (I) domain, down-regulates collagen synthesis. When unligated, a salt bridge between Arg(287) and Glu(317) is thought to keep this domain in a low affinity conformation. Ligand binding opens the salt bridge leading to a high-affinity conformation. How modulating integrin α1β1 affinity alters collagen homeostasis is unknown. To address this question, we utilized a thermolysin-derived product of the α1α2α1 network of collagen IV (α1α2α1(IV) truncated protomer) that selectively binds integrin α1β1. We show that an E317A substitution enhanced binding to the truncated protomer, consistent with a previous finding that this substitution eliminates the salt bridge. Surprisingly, we show that an R287A substitution did not alter binding, whereas R287E/E317R substitutions enhanced binding to the truncated protomer. NMR spectroscopy and molecular modeling suggested that eliminating the Glu(317) negative charge is sufficient to induce a conformational change toward the open state. Thus, the role played by Glu(317) is largely independent of the salt bridge. We further show that cells expressing E317A or R287E/E317R substitutions have enhanced down-regulation of collagen IV synthesis, which is mediated by the ERK/MAPK pathway. In conclusion, we have demonstrated that modulating the affinity of the extracellular α1 I domain to collagen IV enhances outside-in signaling by potentiating ERK activation and enhancing the down-regulation of collagen synthesis.

MeSH Terms (20)

Amino Acid Substitution Animals Cell Line, Transformed Collagen Type IV Down-Regulation Enzyme Activation Extracellular Signal-Regulated MAP Kinases Humans Integrin alpha1beta1 Laminin Ligands MAP Kinase Signaling System Mice Mice, Mutant Strains Models, Molecular Mutation, Missense Nuclear Magnetic Resonance, Biomolecular Protein Binding Protein Biosynthesis Protein Structure, Tertiary

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