Activation of store-operated Ca2+ entry in RBL cells without the contribution of protein kinases.
The role of protein kinases on store-operated Ca2+ entry in rat basophilic leukaemia cells (RBL) has been studied using the whole-cell configuration of the patch-clamp technique and Ca2+ imaging with fura-2. Specific inhibitors of tyrosine kinase (lavendustin A), mitogen-activated protein (MAP) kinase (SB 203580, PD 98059), Ca2+/calmodulin-dependent kinase (CaMK, KN-62, KN-93) and protein kinase C (PKC, bisindolylmaleimide I) had no significant effect on peak current amplitude and time constant of activation. Likewise, the broad spectrum kinase blockers H-7 and staurosporine did not alter Ca2+ entry compared to control recordings. Store-mediated Ca2+ entry was unaffected if intracellular ATP was substituted by either adenosine 5'-O-(2-thiodiphosphate) (ADPbetaS) or adenylyl-imidodiphosphate (AMP-PNP). Similarly, buffering intracellular Mg2+, an essential cofactor for protein kinases, had no effect on Ca2+ influx. These results indicate that protein phosphorylation by various kinases is not required for the activation of the store-operated Ca2+ current in RBL cells.
Bödding
M
M
University Laboratory of Physiology, Oxford, UK.
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D015471:Leukemia, Basophilic, Acute / Q000201:enzymology / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D010636:Phenols / Q000494:pharmacology; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000502:physiology*; D051381:Rats; D019311:Staurosporine / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D004791:Enzyme Inhibitors; D010636:Phenols; D047428:Protein Kinase Inhibitors; C062898:lavendustin A; D011494:Protein Kinases; D019311:Staurosporine; D002118:Calcium