Regulation of integrin-mediated myeloid cell adhesion to fibronectin: influence of disulfide reducing agents, divalent cations and phorbol ester.
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.
Davis
G E
GE
Department of Pathology, Texas A&M University Health Sciences Center, College Station 77843-1114.
Camarillo
C W
CW
D002118:Calcium / Q000494:pharmacology; D002448:Cell Adhesion / Q000187:drug effects / Q000276:immunology; D002460:Cell Line; D004229:Dithiothreitol / Q000494:pharmacology; D005353:Fibronectins / Q000378:metabolism*; D006801:Humans; D016023:Integrins / Q000378:metabolism*; D007962:Leukocytes / Q000166:cytology* / Q000187:drug effects / Q000276:immunology*; D008345:Manganese / Q000494:pharmacology; D010446:Peptide Fragments / Q000378:metabolism; D017469:Receptors, Fibronectin; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D005353:Fibronectins; D016023:Integrins; D010446:Peptide Fragments; D017469:Receptors, Fibronectin; D008345:Manganese; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D004229:Dithiothreitol