Interchangeable c ~ Chain Cytoplasmic Domains Play a Positive Role in Control of Cell Adhesion Mediated by VLA - 4 , a 81 Integrin

Integrins can exist in a range of functional states, depending on the cell type and its state of activation. Although the mechanism that controls activity is unknown, it has been suggested that for some integrins, ot chain cytoplasmic domains may exert either a negative effect or no effect on adhesion function. To address this issue for VLA-4 (an ot431 heterodimer), we constructed an & cytoplasmic deletion mutant and chimeric ot chains composed of the extracellular domains of oe 4 and the cytoplasmic domains of c~ 2, ot 4, or ots. Upon stable transfection of wild-type c~ 4, VLA-4 heterodimer was obtained that mediated (a) poor adhesion to CS1 peptide, fibronectin, or vascular cell adhesion molecule 1 (VCAM-1) (in K562 cells); (b) poor adhesion to CS1 peptide but moderate adhesion to VCAM-1 (in MIP101 cells); and (c) moderate adhesion to both CS1 peptide and VCAM-1 (in PMWK cells). Chimeric o~ 4 constructs and wild-type ot 4 yielded similar results in these cell lines. In contrast, truncation of the c~ 4 cytoplasmic domain (after the conserved GFFKR motif) caused an almost complete loss of adhesive activity in all three cell lines. Thus, several interchangeable ot chain cytoplasmic domains play a fundamentally positive role in determining the state of constitutive activity for VLA-4. The ot chain cytoplasmic domain is also required for agonist-stimulated adhesion, since phorbol ester stimulated the cell adhesion mediated by wild-type and chimeric ol chains, but not by the cytoplasmic deletion mutant. The inactivity of both wild-type VLA-4 (in K562 cells), and truncated VLA-4 (in all three cell lines) was overcome by the addition of a stimulatory anti31 monoclonal antibody. Thus, the ol cytoplasmic domain-dependent cellular mechanism controlling both constitutive and agonist-stimulated VLA-4 activity could be bypassed by external manipulation of the integrin.