Threonine 788 in integrin subunit β1 regulates integrin activation

In the present study, the functional role of suggested phosphorylation of the conserved threonines in the cytoplasmic domain of integrin subunit β1 was investigated. Mutants mimicking phosphorylated and unphosphorylated forms of β1 were expressed in β1 deficient GD25 cells. T788 in β1 was identified as a site with major influence on integrin function. The mutation to A788 strongly reduced β1-dependent cell attachment and exposure of the extracellular 9EG7 epitope, whereas replacement of T789 with alanine did not interfere with the ligand-binding ability. Talin has been shown to mediate integrin activation, but the talin head domain bound equally well to the wild-type β1 and the mutants indicating that the T788A mutation caused defect integrin activation by another mechanism. The phosphorylation-mimicking mutation T788D was fully active in promoting cell adhesion. GD25 cells expressing β1T788D accumulated increased number of focal contacts and migrated slowly compared to GD25 β1 wild-type. An analogous phenotype is seen when focal adhesion kinase activation is abrogated. However, neither the β1T788D nor the β1T788A mutation failed to induce tyrosine phosphorylation of focal adhesion kinase. The results suggest that phosphorylation of T788 in integrin β1 promotes inside-out receptor activation, as well as focal contact accumulation.