title: Cyclic AMP- and inositol phosphate-independent inhibition of Ca2+ influx by cholera toxin in CD3-stimulated Jurkat T cells. A study with a cholera toxin-resistant cell variant and the Ca2+ endoplasmic reticulum-ATPase inhibitor thapsigargin.
abstract: In this report, we describe a Jurkat cell variant, termed JCT8, the selection of which is based upon its resistance to cell-growth inhibition mediated by the holotoxin of Vibrio cholerae, cholera toxin (CT). JCT8 cells exhibit normal cAMP production in response to various cAMP inducers, including CT, together with conserved ADP ribosylation in vitro of G-protein Gs alpha by the A subunit of the toxin. However, after a 4-h pretreatment with CT, JCT8 cells have a conserved expression of cell-surface CD3 molecules. These effects are in contrast to those elicited by the toxin in long term PGE2-desensitized Jurkat cells, which remain as sensitive as the wild type to the inhibitory action of CT on cell growth and CD3 cell-surface expression, despite poor responsiveness to CT with regard to cAMP production. In JCT8 cells, Ca2+ mobilization induced via the CD3/TCR is maintained after CT treatment contrasting with its complete suppression in the wild-type and in the PGE2-desensitized cells. However, as in the other cell types, CT still suppresses Ca2+ influx in JCT8 cells. Increase in inositol phosphates by CD3 stimulation of JCT8 cells, including of inositol 1,4,5-triphosphate (I(1,4,5)P3), is only partially antagonized by CT. This suggests either that in JCT8 cells there is a different susceptibility of Ca2+ mobilization and influx to partial inhibition by CT of CD3-triggered phospholipase C (PLC)-induced phosphoinositide hydrolysis or that an additional and PLC-independent suppressive effect of the toxin on Ca2+ influx may exist. To investigate this particular point further, we use Thapsigargin, a Ca(2+)-endoplasmic reticulum ATPase inhibitor that can mobilize in human T lymphocytes I(1,4,5)P3-dependent intracellular Ca2+ pools by a PLC-independent pathway. We demonstrate that the Ca2+ influx triggered in the wild-type Jurkat cells or in JCT8 cells by Thapsigargin is antagonized by CT. The present data are therefore consistent with the idea that CT specifically impairs in the Jurkat T cell model the entry of Ca2+ from extracellular spaces by a mechanism independent not only from cAMP but also in part from inhibition by the toxin of phosphoinositide hydrolysis.
authors.affiliation: Laboratoire d'Immunologie Cellulaire et Tissulaire, CNRS UA 186, CH Pitié-Salpêtrière, CERVI, Paris, France.
authors.forename: H
authors.initials: H
authors.lastname: Gouy
authors.forename: D
authors.initials: D
authors.lastname: Cefai
authors.forename: S B
authors.initials: SB
authors.lastname: Christensen
authors.forename: P
authors.initials: P
authors.lastname: Debre
authors.forename: G
authors.initials: G
authors.lastname: Bismuth
mesh_terms: D000945:Antigens, Differentiation, T-Lymphocyte / Q000378:metabolism; D017252:CD3 Complex; D002118:Calcium / Q000493:pharmacokinetics*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002772:Cholera Toxin / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000096:biosynthesis; D015232:Dinoprostone / Q000494:pharmacology; D004336:Drug Antagonism; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D066298:In Vitro Techniques; D007295:Inositol Phosphates / Q000096:biosynthesis; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D013601:T-Lymphocytes / Q000187:drug effects* / Q000378:metabolism; D013729:Terpenes / Q000494:pharmacology*; D019284:Thapsigargin
chemical_list: D000945:Antigens, Differentiation, T-Lymphocyte; D017252:CD3 Complex; D007295:Inositol Phosphates; D011948:Receptors, Antigen, T-Cell; D013729:Terpenes; D005576:Colforsin; D019284:Thapsigargin; D002772:Cholera Toxin; D000242:Cyclic AMP; D000252:Calcium-Transporting ATPases; D015232:Dinoprostone; D002118:Calcium