AJP - Gastrointestinal and Liver Physiology, Vol 269, Issue 4 481-G489, Copyright © 1995 by American Physiological Society
A novel serine-specific kinase activity associated with exocrine secretory granules
L. H. Tang, I. M. Modlin, T. A. Caulfield and J. R. Goldenring
Department of Surgery, Yale University School of Medicine, New Haven, Connecticut 06510, USA.
Zymogen secretion from exocrine cells involves an exocytotic process that
is highly regulated by the modification of cytoplasmic components at
different cellular levels. In the present studies, purified secretory
granules were prepared from rabbit gastric chief cells, rat pancreatic
acinar cells, and parotid glands to characterize a Mg(2+)-dependent protein
kinase activity. In chief cell granules, endogenous pepsinogen, a
fortuitous substrate, was phosphorylated at optimal Mg2+ and K+
concentrations of 40 and 50 mM, respectively. Adenosine 3',5'-cyclic
monophosphate, Ca2+, and calmodulin had no significant effects on the
kinase activity. In contrast, Mn2+ or Zn2+ inhibited the kinase activity.
In addition to pepsinogen, the exogenous substrates casein, myelin basic
protein, and lysine-rich histone were also phosphorylated by the
granule-associated kinase. All substrates were exclusively phosphorylated
on serine residues. ATP, but not GTP, served as the donor in the
phosphorate transfer reaction. Casein kinase (CK) inhibitors CKI-7 and
dibromoribofuransylbenzimidazole at concentrations (10 microM) that
significantly inhibited CK activities in the tissue homogenate failed to
inhibit the granule-associated kinase activity. The kinase activity was
localized to the granule membrane and could be removed from the membrane
with either 5 mM EDTA or alkaline carbonate extraction. Furthermore,
protease digestion sensitivity revealed that the kinase was localized on
the cytoplasmic face of the granules. Our results therefore indicate that
the secretory granules of exocrine gastric chief cells, pancreatic acini,
and parotid acini possess a unique serine-specific protein kinase activity.
The cytoplasmic orientation of the kinase activity suggests a possible role
in vesicle processing or the exocytotic process.
