AJP - Cell Physiology, Vol 258, Issue 2 C227-C233, Copyright © 1990 by American Physiological Society
Protein kinase C mediates cholinergically regulated protein phosphorylation in a Cl(-)-secreting epithelium
J. A. Cohn
Department of Medicine, Duke University, Durham, North Carolina 27710.
T84 cell monolayers were used to study the cholinergic regulation of
protein phosphorylation in epithelial cells. When T84 cell monolayers are
labeled with 32Pi and stimulated with carbachol, six proteins exhibit
altered phosphorylation. The most prominent response is a fivefold increase
in labeling of p83, an acidic protein of Mr 83,000. Increasing labeling of
p83 parallels stimulated secretion with respect to the onset of agonist
action, agonist potency, and antagonism by atropine. However, the p83 and
secretory responses differ in that the p83 response is more sustained. When
T84 cell fractions are incubated with [gamma-32P]ATP, Ca2(+)-phospholipid
stimulates p83 labeling. Phosphorylation of p83 also occurs when a T84 cell
extract is incubated with purified protein kinase C and when intact cells
are exposed to phorbol myristate acetate. p83 does not become
phosphorylated in cell fractions incubated with adenosine 3',5'-cyclic
monophosphate (cAMP) or in monolayers stimulated with agonists acting via
cAMP. Thus carbachol stimulates the phosphorylation of an endogenous
substrate for protein kinase C in T84 cells. The duration of this
phosphorylation response suggests that protein kinase C may mediate a
sustained response to carbachol, possibly acting to limit the duration of
stimulated secretion.
