cAMP‐dependent protein phosphorylation of membrane proteins in the parotid gland, platelets and liver

Abstract

Stimulation of secretion in exocrine secretory glands leads to the phosphorylation of a 22-kDa membrane protein (protein 111) whose function is still unknown [Jahn et al. (1980) Eur. J. Biochem. 112, 345-352;Jahn & Soling (1980) Proc. Nut1 Acad. Sci. USA 78, 6903 -69061. This report describes the comparison of this protein with phosphorylated membrane proteins of similar molecular mass in platelets and liver. Incubation of platelets with agents which raise the intracellular cAMP concentration results in the phosphorylation of a 22-kDa protein which is also phosphorylated in membrane preparations by endogenous kinases or by exogenous CAMP-dependent protein kinase. It is shown that this protein is distinct from protein I11 although both proteins have the same molecular mass and are substrates of CAMP-dependent protein kinase. In contrast to platelets, protein I11 could be demonstrated in liver microsomes. This indicates that the function of protein 111 is not exclusively linked to the stimulus-secretion coupling in exocrine cells.Stimulation of secretion in exocrine glands by agonists involving cAMP as second messenger leads to the phosphorylation of the ribosomal protein S6 (protein I) and two other particulate proteins with apparent molecular masses of 24 kDa (protein 11) and 22 kDa (protein 111) [I -91. Recently, we published the purification and characterization of protein 111 [lo] showing that this protein belongs to a group of very hydrophobic membrane proteins whose functions are still unknown. Spearman et al. [Ill and Quissel[9] questioned the idea of a direct relationship between an increased phosphorylation state of protein I11 and the secretory process in exocrine secretory glands because the protein is located in the endoplasmic reticulum [9, 11, 121 and shows a slow rate of phosphate turnover. Plewe et al. [12], who found protein 111 in a microsomal subfraction that exhibits the highest ATPdependent calcium transport activity, discussed a possible role of protein 111 for the regulation of calcium sequestration by the endoplasmic reticulum. This hypothesis was adopted in analogy to the function of phospholamban, the putative regulator of the cardiac Ca2+ pump, which was described previously as a 22-kDa phosphoprotein In contrast to the parotid gland, where a rise in intracelluIar cAMP concentration induces exocytosis, cAMP inhibits aggregation and secretion in platelets. This effect of cAMP in platelets is associated with the phosphorylation of various proteins suggesting a coupling between the inhibitory role of cAMP on exocytosis and protein phosphorylation. One of these proteins is a 22-kDa membrane protein which is claimed to play a role in the CAMP-stimulated active transport of Ca2+ out of the platelet cytosol [26-361. This protein was found in Ca'+-accumulating vesicles, which led to the assumption of an analogy to phospholamban ('thrombolamban' [36]), although experimental data supporting this assumption have not been presented [37, 381. The 22-kDa protein which became phosphorylated by an en...