Two Synergistic Activation Mechanisms of α2β1 Integrin-mediated Collagen Binding

Wendy L Connors,Johanna Jokinen,Daniel J White,J Santeri Puranen,Pasi Kankaanpaöaö,Paula Upla,Mira Tulla,Mark S Johnson,Jyrki Heino

doi:10.1074/jbc.m700759200

Wendy L Connors, Johanna Jokinen + Show 7 more

Open Access

https://doi.org/10.1074/jbc.m700759200

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Abstract

Activation of protein kinase C by 12-O-tetradecanoylphorbol-13-acetate (TPA) induces ligand-independent aggregation of a cell surface collagen receptor, alpha2beta1 integrin. Concomitantly, TPA increases the avidity of alpha2beta1 for collagen and the number of conformationally activated alpha2beta1 integrins. The structural change was shown using a monoclonal antibody 12F1 that recognizes the "open" (active) conformation of the inserted domain in the alpha2 subunit (alpha2I). Amino acid residue Glu-336 in alpha2 subunit is proposed to mediate the interaction between alpha2I domain and beta1 subunit. Glu-336 seems to regulate a switch between open and "closed" conformations, since the mutation alpha2E336A inhibited the TPA-related increase in the number of 12F1 positive integrins. E336A also reduced cell adhesion to collagen. However, E336A did not prevent the TPA-related increase in adhesion to collagen or alpha2beta1 aggregation. Thus, alpha2beta1 integrin avidity is regulated by two synergistic mechanisms, first an alpha2E336-dependent switch to the open alpha2I conformation, and second an alpha2E336-independent mechanism temporally associated with receptor aggregation.

Highlights

Integrin-type cell adhesion receptors undergo functional regulation via different conformational states [1]
In an inactive integrin ␣/␤ heterodimer, the ligand binding headpiece is bent toward the cell surface [2], while an active integrin stands tall but may still have multiple activation states
We report here that while TPA induces rapid (5–15 min) clustering of cell surface ␣2␤1 in the absence of ligand and increases ␣2␤1 avidity for collagen, it activates the integrin via a conformational mechanism involving ␣2E336 and intermolecular cross-talk between the ␣2I and ␤1I domains

Summary

Introduction

Integrin-type cell adhesion receptors undergo functional regulation via different conformational states [1]. We report here that while TPA induces rapid (5–15 min) clustering of cell surface ␣2␤1 in the absence of ligand and increases ␣2␤1 avidity for collagen, it activates the integrin via a conformational mechanism involving ␣2E336 and intermolecular cross-talk between the ␣2I and ␤1I domains.

Results

Conclusion