Characteristics of the free cytosolic calcium timelag following IgE-mediated stimulation of human basophils: significance for the nonreleasing basophil phenotype.
These studies examine characteristics of the quiescent period (timelag) of the free cytosolic calcium ([Ca++]i) elevation that follows stimulation of human basophils through the IgE receptor. Previous studies established that the [Ca++]i timelag was sensitive to the rate of ligand binding, but little else is known about this response characteristic. The [Ca++]i timelag could be lengthened using antigenic stimulation that is rapid but only weakly induces secretion: tenfold differences in the "strength" of the stimulus, as assessed by histamine release, are associated with threefold differences in the timelag. Inhibiting p53/56lyn kinase with low concentrations of the specific inhibitor, PP1, lengthened the [Ca++]i timelag dramatically. PP1 was also found to delay the onset of syk phosphorylation and histamine release. Staurosporine and genistein, which are known to inhibit early tyrosine kinases, had, at best, only modest effects on the [Ca++]i timelag. Specific inhibitors of protein kinase C (PKC) had no effect on the [Ca++]i timelag, and direct activation of PKC with PMA had only very modest effects on the timelag. Contrary to expectations, basophils with the so-called nonreleasing phenotype demonstrated an IgE-mediated [Ca++]i response at the single-cell level. However, the length of [Ca++]i timelag in nonreleasing basophils was threefold longer than normally found in releasing basophils. Furthermore, the [Ca++]i response was significantly more asynchronous than in releasing basophils and lacking in a sustained [Ca++]i elevation. These studies indicate that the [Ca++]i timelag following stimulation through the IgE receptor is sensitive to inhibition of lyn kinase but not other agents that have been demonstrated to inhibit early tyrosine kinases previously. However, only one characteristic of the [Ca++]i response phenotype of nonreleasing basophils--the [Ca++]i timelag but not the absence of a sustained [Ca++]i elevation--could be mimicked by inhibition of lyn kinase with PP1.
MacGlashan
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Johns Hopkins University, Asthma & Allergy Center, Baltimore, Maryland 21224, USA. dmacglashan@welch.jhu.edu
Lavens-Phillips
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D000328:Adult; D001491:Basophils / Q000187:drug effects / Q000201:enzymology / Q000276:immunology* / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000276:immunology; D002469:Cell Separation; D002868:Chromosomal Proteins, Non-Histone; D003600:Cytosol / Q000201:enzymology / Q000276:immunology / Q000378:metabolism*; D004268:DNA-Binding Proteins; D004789:Enzyme Activation / Q000276:immunology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004792:Enzyme Precursors / Q000378:metabolism; D006636:Histamine Release / Q000187:drug effects / Q000276:immunology*; D056488:Histone Chaperones; D006801:Humans; D007073:Immunoglobulin E / Q000502:physiology*; D016130:Immunophenotyping; D007424:Intracellular Fluid / Q000201:enzymology / Q000276:immunology / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D007700:Kinetics; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011506:Proteins / Q000494:pharmacology; D017455:Receptors, IgE / Q000502:physiology; D000072377:Syk Kinase; D014157:Transcription Factors; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D004792:Enzyme Precursors; D056488:Histone Chaperones; D047908:Intracellular Signaling Peptides and Proteins; D011506:Proteins; D017455:Receptors, IgE; C513473:SET protein, human; D014157:Transcription Factors; C073882:phosphoprotein phosphatase inhibitor 1; D007073:Immunoglobulin E; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D010749:Phosphoprotein Phosphatases; D002118:Calcium