Carbachol, Substance P, and Phorbol Ester Promote the Tyrosine Phosphorylation of Protein Kinase C in Salivary Gland Epithelial Cells

doi:10.1074/jbc.270.22.13490

Carbachol, Substance P, and Phorbol Ester Promote the Tyrosine Phosphorylation of Protein Kinase C in Salivary Gland Epithelial Cells (*)

Stephen P. Soltoff(§) and
Alex Toker

From the (1) Department of Medicine, Division of Signal Transduction, Beth Israel Hospital, Boston, Massachusetts 02115

^§ To whom correspondence should be addressed:
Div. of Signal Transduction, Harvard Medical School, Alpert Bldg., 200 Longwood Ave., Boston, MA 02115.
Tel.: 617-278-3093; Fax: 617-278-3131; E-mail: ssoltoff{at}mercury.bih.harvard.edu.

Abstract

The initiation of saliva formation by parotid acinar cells, which comprise the majority of cells in this salivary gland, is initiated by the release of neurotransmitters (acetylcholine, substance P) from parasympathetic nerves. In response to substance P and the muscarinic agonist carbachol, two ligands that activate phospholipase C-linked receptors, which stimulate fluid secretion, PKC was phosphorylated on tyrosine residues. The maximal agonist-dependent tyrosine phosphorylation occurred within seconds of the addition of either agonist and then returned rapidly to a smaller increased level. Phorbol ester also caused a rapid increase in tyrosine phosphorylation, which reached a maximal level 5 min after the addition of phorbol 12-myristate 13-acetate. The increase in tyrosine phosphorylation of PKC was blocked by tyrosine kinase inhibitors genistein and staurosporine. Ionophore-mediated elevation of [Ca] or activation of the β-adrenergic receptor, epidermal growth factor receptor, or insulin receptor did not promote the tyrosine phosphorylation of PKC. These results indicate that tyrosine phosphorylation plays a role in early signal transduction events promoted by the activation of muscarinic and substance P receptors and suggests that the tyrosine phosphorylation of PKC has a role in the activation of fluid secretion by neurotransmitters binding to phospholipase C-linked receptors.

Footnotes

↵* This work was supported in part by National Institute of Health Grant DE10877 (to S. P. S.) and GM41890 (to A. T., and L. C. Cantley). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

PKC

protein kinase C

anti-P-Tyr

anti-phosphotyrosine

PMSF

phenylmethylsufonyl fluoride

EGF

epidermal growth factor

PMA

phorbol 12-myristate 13-acetate

PAGE

polyacrylamide gel electrophoresis.
↵²S. P. Soltoff, unpublished results.