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Identification of Exo2 as the catalytic subunit of protein kinase A reveals a role for cyclic AMP in Ca(2+)‐dependent exocytosis in chromaffin cells.
Author(s) -
Morgan A.,
Wilkinson M.,
Burgoyne R.D.
Publication year - 1993
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1002/j.1460-2075.1993.tb06052.x
Subject(s) - exocytosis , biology , secretion , protein subunit , biochemistry , gene
Digitonin‐permeabilized chromaffin cells secrete catecholamines by exocytosis in response to micromolar Ca2+ concentrations, but lose the ability to secrete in response to Ca2+ as the cells lose soluble proteins through the plasma membrane pores. We have previously shown [Morgan and Burgoyne (1992) Nature, 355, 833–836] that cytosol can retard this loss of secretory competence and that two distinct stimulatory activities (Exo1 and Exo2) are present in cytosol. Here we report that Exo2 behaved as a single peak of activity through purification on hydroxyapatite, ammonium sulfate precipitation and gel filtration and the activity correlated with a single polypeptide of approximately 44 kDa on SDS gels. Protein sequencing of this band revealed it to be the catalytic subunit of cyclic AMP‐dependent protein kinase (PKA). Both cyclic AMP and the commercially available catalytic subunit of PKA stimulated exocytosis in a dose‐dependent manner which was absolutely dependent on the presence of micromolar Ca2+. These data show that PKA (Exo2) regulates Ca(2+)‐dependent exocytosis in bovine adrenal chromaffin cells.