β Integrin Tyrosine Phosphorylation Is a Conserved Mechanism for Regulating Talin-induced Integrin Activation

Integrins are large membrane-spanning receptors fundamental to cell adhesion and migration. Integrin adhesiveness for the extracellular matrix is activated by the cytoskeletal protein talin via direct binding of its phosphotyrosine-binding-like F3 domain to the cytoplasmic tail of the β integrin sub...

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Bibliographic Details
Published in:The Journal of biological chemistry 2009-12, Vol.284 (52), p.36700-36710
Main Authors: Anthis, Nicholas J., Haling, Jacob R., Oxley, Camilla L., Memo, Massimiliano, Wegener, Kate L., Lim, Chinten J., Ginsberg, Mark H., Campbell, Iain D.
Format: Article
Language:English
Subjects:
Amino Acid Motifs - physiology
Amino Acid Substitution
Animals
Cell Line
DNA-Binding Proteins - chemistry
DNA-Binding Proteins - genetics
DNA-Binding Proteins - metabolism
Integrin beta Chains - chemistry
Integrin beta Chains - genetics
Integrin beta Chains - metabolism
Mice
Mutation, Missense
Nuclear Magnetic Resonance, Biomolecular
Phosphoproteins - chemistry
Phosphoproteins - genetics
Phosphoproteins - metabolism
Phosphorylation - physiology
Protein Binding - physiology
Protein Structure, Tertiary - physiology
Protein Synthesis, Post-Translational Modification, and Degradation
RNA-Binding Proteins - chemistry
RNA-Binding Proteins - genetics
RNA-Binding Proteins - metabolism
Talin - chemistry
Talin - genetics
Talin - metabolism
Tyrosine - chemistry
Tyrosine - genetics
Tyrosine - metabolism
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Summary:Integrins are large membrane-spanning receptors fundamental to cell adhesion and migration. Integrin adhesiveness for the extracellular matrix is activated by the cytoskeletal protein talin via direct binding of its phosphotyrosine-binding-like F3 domain to the cytoplasmic tail of the β integrin subunit. The phosphotyrosine-binding domain of the signaling protein Dok1, on the other hand, has an inactivating effect on integrins, a phenomenon that is modulated by integrin tyrosine phosphorylation. Using full-length tyrosine-phosphorylated 15N-labeled β3, β1A, and β7 integrin tails and an NMR-based protein-protein interaction assay, we show that talin1 binds to the NPXY motif and the membrane-proximal portion of β3, β1A, and β7 tails, and that the affinity of this interaction is decreased by integrin tyrosine phosphorylation. Dok1 only interacts weakly with unphosphorylated tails, but its affinity is greatly increased by integrin tyrosine phosphorylation. The Dok1 interaction remains restricted to the integrin NPXY region, thus phosphorylation inhibits integrin activation by increasing the affinity of β integrin tails for a talin competitor that does not form activating membrane-proximal interactions with the integrin. Key residues governing these specificities were identified by detailed structural analysis, and talin1 was engineered to bind preferentially to phosphorylated integrins by introducing the mutation D372R. As predicted, this mutation affects talin1 localization in live cells in an integrin phosphorylation-specific manner. Together, these results indicate that tyrosine phosphorylation is a common mechanism for regulating integrin activation, despite subtle differences in how these integrins interact with their binding proteins.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.M109.061275