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Facilitation of Ca 2+ ‐dependent exocytosis by Rac1‐GTPase in bovine chromaffin cells
Author(s) -
Li Quanwen,
Ho Chi S.,
Marinescu Vlad,
Bhatti Humaa,
Bokoch Gary M.,
Ernst Stephen A.,
Holz Ronald W.,
Stuenkel Edward L.
Publication year - 2003
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.2003.039073
Subject(s) - rac1 , microbiology and biotechnology , pak1 , secretion , gtpase , biology , rac gtp binding proteins , guanine nucleotide exchange factor , cytoskeleton , signal transduction , cell , biochemistry
Rho family GTPases are primary mediators of cytoskeletal reorganization, although they have also been reported to regulate cell secretion. Yet, the extent to which Rho family GTPases are activated by secretory stimuli in neural and neuroendocrine cells remains unknown. In bovine adrenal chromaffin cells, we found Rac1, but not Cdc42, to be rapidly and selectively activated by secretory stimuli using an assay selective for the activated GTPases. To examine effects of activated Rac1 on secretion, constitutively active mutants of Rac1 (Rac1‐V12, Rac1‐L61) were transiently expressed in adrenal chromaffin cells. These mutants facilitated secretory responses elicited from populations of intact and digitonin‐permeabilized cells as well as from cells under whole cell patch clamp. A dominant negative Rac1 mutant (Rac1‐N17) produced no effect on secretion. Expression of RhoGDI, a negative regulator of Rac1, inhibited secretory responses while overexpression of effectors of Rac1, notably, p21‐activated kinase (Pak1) and actin depolymerization factor (ADF) promoted evoked secretion. In addition, expression of effector domain mutants of Rac1‐V12 that exhibit reduced activation of the cytoskeletal regulators Pak1 and Partner of Rac1 (POR1) resulted in a loss of Rac1‐V12‐mediated enhancement of evoked secretion. These findings suggest that Rac1, in part, functions to modulate secretion through actions on the cytoskeleton. Consistent with this hypothesis, the actin modifying drugs phalloidin and jasplakinolide enhanced secretion, while latrunculin‐A inhibited secretion and eliminated the secretory effects of Rac1‐V12. In summary, Rac1 was activated by secretory stimuli and modulated the secretory pathway downstream of Ca 2+ influx, partly through regulation of cytoskeletal organization.