title: Dissection of lymphocyte function-associated antigen 1-dependent adhesion and signal transduction in human natural killer cells shown by the use of cholera or pertussis toxin.
abstract: The effect of the guanosine triphosphate-binding protein (G-protein) inhibitors cholera toxin (Ctx) and pertussis toxin (Ptx) has been analyzed on lymphocyte function-associated antigen 1 (LFA-1)-dependent adhesion and signal transduction in human natural killer (NK) cells. Ctx, but not Ptx, inhibited the LFA-1-dependent adhesion of NK cells to tumor target cells which constitutively express the intercellular cell adhesion molecule-1 (ICAM-1) and to NIH/3T3 mouse fibroblasts stably transfected with human ICAM-1. This effect was detectable only by the use of the entire Ctx but not of the Ctx B subunit. In addition, Ctx could inhibit both NK cell binding and spreading to purified ICAM-1 protein. NK cell treatment with Ctx modified neither the surface expression of LFA-1 nor its Mg2+ binding site. These findings, together with the absence of any detectable effect of Ctx on the constitutive phosphorylation of LFA-1 alpha, suggests that this toxin modifies the avidity of LFA-1 for ICAM-1 by acting on LFA-1-cytoskeletal protein association. Unlike Ctx, Ptx did not affect NK cell adhesion. The effects of Ctx and Ptx are unlikely to depend on intracellular levels of cyclic adenosine 3',5'-monophosphate (cAMP), since a strong increase of cAMP was induced by both toxins. Moreover, this was confirmed by the observation that the LFA-1-dependent adhesion was not inhibited by the adenylate cyclase activator forskolin (FSK), the phosphodiesterase inhibitor isobutyl-1-methylxanthine (IBMX), or both, which increase intracellular cAMP levels. Unlike the differential effect on cell adhesion, both the intracellular calcium [Ca2+]i increase and phosphoinositide breakdown mediated via LFA-1 were consistently inhibited in a dose-dependent manner by both Ctx and Ptx. Also in this case, the inhibitory effect did not depend on an increase of intracellular cAMP as indicated by NK cell treatment with FSK, IBMX, or both. Further evidence of the involvement of G-proteins in LFA-1-mediated signal transduction was the inhibitory effect of the GDP analog guanosine-5'-O-2-thiodiphosphate (GDP beta S) on LFA-1-mediated calcium mobilization. Taken together, our data provide evidence that the LFA-1-mediated NK cell adhesion and signal transduction are partially independent phenomena which may be regulated by different G-proteins.
authors.affiliation: Laboratory of Immunopathology, Centro di Biotecnologie Avanzate, Genoa, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Poggi
authors.forename: F
authors.initials: F
authors.lastname: Spada
authors.forename: P
authors.initials: P
authors.lastname: Costa
authors.forename: E
authors.initials: E
authors.lastname: Tomasello
authors.forename: V
authors.initials: V
authors.lastname: Revello
authors.forename: N
authors.initials: N
authors.lastname: Pella
authors.forename: M R
authors.initials: MR
authors.lastname: Zocchi
authors.forename: L
authors.initials: L
authors.lastname: Moretta
mesh_terms: D037361:Adenylate Cyclase Toxin; D000818:Animals; D002448:Cell Adhesion / Q000187:drug effects / Q000276:immunology*; D002772:Cholera Toxin / Q000494:pharmacology*; D000242:Cyclic AMP / Q000096:biosynthesis; D003602:Cytotoxicity, Immunologic / Q000187:drug effects; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism; D007694:Killer Cells, Natural / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D016169:Lymphocyte Function-Associated Antigen-1 / Q000187:drug effects / Q000502:physiology*; D012515:Mast-Cell Sarcoma; D051379:Mice; D037342:Pertussis Toxin; D015398:Signal Transduction / Q000187:drug effects / Q000276:immunology*; D014407:Tumor Cells, Cultured; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
chemical_list: D037361:Adenylate Cyclase Toxin; D007295:Inositol Phosphates; D016169:Lymphocyte Function-Associated Antigen-1; D010566:Virulence Factors, Bordetella; D002772:Cholera Toxin; D000242:Cyclic AMP; D037342:Pertussis Toxin