TY - JOUR
T1 - Staurosporine-inhibitable protein kinase activity associated with secretory granule membranes isolated from rat submandibular gland cells
AU - Seo, Su Ryeon
AU - Kim, Yeun Ju
AU - Moon, Seok Jun
AU - Sugiya, Hiroshi
AU - Shin, Dong Min
AU - Lee, Syng Ill
AU - Seo, Jeong Taeg
N1 - Funding Information:
This work was supported by the Research Fund from Yonsei University for 1999.
PY - 2003/8/1
Y1 - 2003/8/1
N2 - Protein kinases, such as protein kinase C, have been shown to be associated with secretory granules and to regulate the event of exocytosis in various tissues including parotid salivary acinar cells. However, in submandibular acinar cells that play an important role in the secretion of proteins into the oral cavity, kinase activity on the granule membrane has not been explored. Therefore, in the present study, we isolated the secretory granules from rat submandibular acinar cells and investigated the localisation of protein kinases on the granule membrane. Initially, we isolated and purified secretory granules from rat submandibular acinar cells. Addition of [γ-32P] ATP to granule-membrane lysate phosphorylated the granule-membrane-associated 26, 32, 55 and 58 kDa proteins, sussestins the presence of endosenous kinase activity on the membrane. Moreover, the phosphorylation of 26 and 32 kDa proteins was inhibited by staurosporine and K252a, both non-specific protein kinase C inhibitors. However, the phosphorylation of 26 and 32 kDa proteins was not inhibited by other protein kinase C inhibitors, such as calphostin C, GF109203X and chelerythrine, indicatins that protein kinase C was not responsible for the phosphorylation. In addition, H-89, ML-9, KN-62 and genistein did not appear to inhibit this phosphorylation, indicating that protein kinase A, myosin light chain kinase (MLCK), Ca2+/calmodulin-dependent protein kinase II (CAMKII) and tyrosine kinase were not involved in the phosphorylation of 26 and 32 kDa proteins. Moreover, Ca2+ had no effect on the kinase activity. Therefore, our results suggest that an unidentified, staurosporine-inhibitable protein kinase activity is associated with the secretory granule membrane of rat submandibular acinar cells.
AB - Protein kinases, such as protein kinase C, have been shown to be associated with secretory granules and to regulate the event of exocytosis in various tissues including parotid salivary acinar cells. However, in submandibular acinar cells that play an important role in the secretion of proteins into the oral cavity, kinase activity on the granule membrane has not been explored. Therefore, in the present study, we isolated the secretory granules from rat submandibular acinar cells and investigated the localisation of protein kinases on the granule membrane. Initially, we isolated and purified secretory granules from rat submandibular acinar cells. Addition of [γ-32P] ATP to granule-membrane lysate phosphorylated the granule-membrane-associated 26, 32, 55 and 58 kDa proteins, sussestins the presence of endosenous kinase activity on the membrane. Moreover, the phosphorylation of 26 and 32 kDa proteins was inhibited by staurosporine and K252a, both non-specific protein kinase C inhibitors. However, the phosphorylation of 26 and 32 kDa proteins was not inhibited by other protein kinase C inhibitors, such as calphostin C, GF109203X and chelerythrine, indicatins that protein kinase C was not responsible for the phosphorylation. In addition, H-89, ML-9, KN-62 and genistein did not appear to inhibit this phosphorylation, indicating that protein kinase A, myosin light chain kinase (MLCK), Ca2+/calmodulin-dependent protein kinase II (CAMKII) and tyrosine kinase were not involved in the phosphorylation of 26 and 32 kDa proteins. Moreover, Ca2+ had no effect on the kinase activity. Therefore, our results suggest that an unidentified, staurosporine-inhibitable protein kinase activity is associated with the secretory granule membrane of rat submandibular acinar cells.
KW - K252a
KW - Secretory sranule membrane
KW - Staurosporine
KW - Submandibular acinar cells
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U2 - 10.1016/S0003-9969(03)00097-9
DO - 10.1016/S0003-9969(03)00097-9
M3 - Article
C2 - 12828983
AN - SCOPUS:0037967986
SN - 0003-9969
VL - 48
SP - 553
EP - 558
JO - Archives of Oral Biology
JF - Archives of Oral Biology
IS - 8
ER -