Characterization of β2 (CD18) integrin phosphorylation in phorbol ester-activated T lymphocytes

Integrins are transmembrane proteins involved in cell-cell and cell-extracellular-matrix interactions. The affinity and avidity of integrins for their ligands change in response to cytoplasmic signals. This 'inside-out' activation has been reported to occur also with beta(2) integrins (CD18). The beta(2) integrin subunit has previously been shown to become phosphorylated in T lymphocytes on cytoplasmic serine and the functionally important threonine residues after treatment with phorbol esters or on triggering of T-cell receptors. We have now characterized the phosphorylation of beta(2) integrins in T-cells in more detail. When T-cells were activated by phorbol esters the phosphorylation was mainly on Ser(756). After inhibition of serine/threonine phosphatases, phosphorylation was also found in two of the threonine residues in the threonine triplet 758-760 of the beta(2) cytoplasmic domain. Activation of T-cells by phorbol esters resulted in phosphorylation in only approx. 10%, of the integrin molecules. Okadaic acid increased this phosphorylation to approx. 30% of the beta(2) molecules, assuming three phosphorylation sites. This indicates that a strong dynamic phosphorylation exists in serine and threonine residues of the beta(2) integrins.