The Journal of Immunology, Vol 151, Issue 12 7138-7150, Copyright © 1993 by American Association of Immunologists

ARTICLES

Regulation of integrin-mediated myeloid cell adhesion to fibronectin: influence of disulfide reducing agents, divalent cations and phorbol ester

GE Davis and CW Camarillo
Department of Pathology, Texas A&M University Health Sciences Center, College Station 77843-1114.

Three different agents, dithiothreitol (DTT), Mn2+, and phorbol ester (TPA), were found to induce HL-60 cell adhesion to fibronectin through distinct mechanisms. The binding of HL-60 cells to fibronectin and a 120-kDa fibronectin fragment is completely dependent on the alpha 5 beta 1 integrin, the adhesion activators, and appropriate divalent cations such as Mg2+. Mn2+ alone was able to induce maximal adhesion in the absence of these other activators. With any of the three activators, Ca2+ inhibited adhesion to fibronectin substrates by inhibiting alpha 5 beta 1-fibronectin binding. DTT and Mn2+ were both found to enhance the binding of fibronectin to purified alpha 5 beta 1, which suggests that both agents can directly stimulate the integrin- ligand binding reaction. TPA acts by inducing intracellular phosphorylation whereas neither DTT nor Mn2+ induced protein phosphorylation. TPA-treated HL-60 cells adhere and spread on fibronectin substrates, whereas DTT- and Mn(2+)-treated cells adhere but do not spread. The actin cytoskeletal inhibitor, cytochalasin B, markedly blocks TPA-induced adhesion, has an intermediate effect on DTT- induced adhesion, and has a minimal effect on Mn(2+)-induced adhesion. Collectively, the data suggest that TPA seems to act by inducing phosphorylation events that lead to cytoskeletal changes and alpha 5 beta 1 integrin activation. In contrast, DTT and Mn2+ seem to act primarily by directly influencing the alpha 5 beta 1-fibronectin binding reaction. These studies characterize in detail a regulatory system for studying leukocyte alpha 5 beta 1-fibronectin adhesion and identify DTT as a novel activator of alpha 5 beta 1-fibronectin binding.

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