Calcium-G protein interactions in the regulation of macrophage secretion.
The interplay between activated G proteins and intracellular calcium ([Ca(2+)](i)) in the regulation of secretion was studied in the macrophage, coupling membrane capacitance with calcium-sensitive microfluorimetry. Intracellular elevation of either the nonhydrolyzable analogue of GTP, guanosine-5'-O-(3-thiotriphosphate) (GTP gamma S), or [Ca(2+)](i) enhanced the amplitude and shortened the time course of stimulus-induced secretion in a dose-dependent manner. Both the ionophore- and the stimulus-induced secretory response were abolished in the presence of guanosine-5'-O-(2-thiodiphosphate) (GDP beta S). The K(d) of Ca(2+)-driven secretion was independent of GTP gamma S concentration, whereas the K(d) of the GTP gamma S-driven response decreased from 63 to 31 microM in the presence of saturating concentrations of [Ca(2+)](i). The time course of stimulus-induced secretion was dependent upon the concentration of [Ca(2+)](i). The time course of GTP gamma S-driven secretion was concentration-independent at high levels of [Ca(2+)](i), suggesting that a calcium-dependent translocation/binding step was rate-limiting. Our data strongly support a model in which [Ca(2+)](i) and activated G proteins act independently of one another in the sequential regulation of macrophage secretion. [Ca(2+)](i) appears to play a role in the recruitment and priming of vesicles from reserve intracellular pools at a step that is upstream of G protein activation. While activated, G proteins appear to play a key role in fusion of docked vesicles. Thus, secretion can result either from activating more G proteins or from elevating [Ca(2+)](i) at basal levels of G protein activation.
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Department of Neurobiology, Pharmacology and Physiology, The University of Chicago, Chicago, Illinois 60637, USA.
Krupa
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Nelson
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DJ
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D005089:Exocytosis / Q000502:physiology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D007074:Immunoglobulin G / Q000276:immunology; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D010587:Phagocytosis / Q000502:physiology; D013696:Temperature
D007074:Immunoglobulin G; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019204:GTP-Binding Proteins; D002118:Calcium