TY - JOUR
T1 - Somatostatin-14, somatostatin-28, and prosomatostatin([1-10]) are independently and efficiently processed from prosomatostatin in the constitutive secretory pathway in islet somatostatin tumor cells (1027B2)
AU - Patel, Yogesh C.
AU - Galanopoulou, Aristea S.
AU - Rabbani, Shahida N.
AU - Liu, Jun Li
AU - Ravazzola, Mariella
AU - Amherdt, Mylene
N1 - Funding Information:
This work was supported by grants from the Canadian Medical Research Council (MT-6196) and the Swiss NSF (31-34088.92).
PY - 1997/8/8
Y1 - 1997/8/8
N2 - We have characterized the biosynthetic origin of somatostatin-14 (SS-14), SS-28, and pro-SS([1-10]) from pro-SS (PSS) in 1027B2 rat islet tumor cells. Because these cells lack regulated secretion and show unresponsiveness of the SS gene to cAMP, we have additionally carried out morphological and functional studies to elucidate the molecular defect in cAMP signalling and to localize the sites of PSS maturation along the secretory pathway. Cell extracts and secretion media were analysed by high performance liquid chromatography and specific C- and N-terminal radioimmunoassays. Electron microscopic sampling of 1027B2 cell cultures showed that most cells had very few dense core secretory granules for heterogeneous sizes. The cells expressed the endoproteases furin, PC1, and PC2 and contained large quantities of fully processed SS-14 and SS-28 with very little unprocessed PSS (ratio SS - 14:SS - 28:PSS = 39:51:10%). They secreted high concentrations of SS-14, SS-28, and PSS([1-10]) constitutively along with PC1 and PC2. Pulse-chase studies demonstrated that PSS is rapidly (within 15 min), and efficiently processed to SS - 14, SS - 28, and PSS([1-10]) via separate biosynthetic pathways: PSS → SS-14 + 8 kDa; PSS → SS-28 + 7 kDa; PSS → PSS([1-10]). Monensin reduced intracellular SS-like immunoreactivity without altering processing efficiency. Transfection with the catalytic subunit of protein kinase A (PKA-C) activated SS promoter-CAT activiting indicating that the defect in cAMP-dependent signaling in 1027B2 cells lies at the level of PKA-C. PKA-C overexpression failed to alter the ratio of processed SS - 14 and SS - 28. These results demonstrate that SS - 14, SS - 28, and PSS([1-10]) are independently synthesized from PSS and that efficient precursor processing can occur within the constitutive secretory pathway in the relative absence of dense core secretory vesicles.
AB - We have characterized the biosynthetic origin of somatostatin-14 (SS-14), SS-28, and pro-SS([1-10]) from pro-SS (PSS) in 1027B2 rat islet tumor cells. Because these cells lack regulated secretion and show unresponsiveness of the SS gene to cAMP, we have additionally carried out morphological and functional studies to elucidate the molecular defect in cAMP signalling and to localize the sites of PSS maturation along the secretory pathway. Cell extracts and secretion media were analysed by high performance liquid chromatography and specific C- and N-terminal radioimmunoassays. Electron microscopic sampling of 1027B2 cell cultures showed that most cells had very few dense core secretory granules for heterogeneous sizes. The cells expressed the endoproteases furin, PC1, and PC2 and contained large quantities of fully processed SS-14 and SS-28 with very little unprocessed PSS (ratio SS - 14:SS - 28:PSS = 39:51:10%). They secreted high concentrations of SS-14, SS-28, and PSS([1-10]) constitutively along with PC1 and PC2. Pulse-chase studies demonstrated that PSS is rapidly (within 15 min), and efficiently processed to SS - 14, SS - 28, and PSS([1-10]) via separate biosynthetic pathways: PSS → SS-14 + 8 kDa; PSS → SS-28 + 7 kDa; PSS → PSS([1-10]). Monensin reduced intracellular SS-like immunoreactivity without altering processing efficiency. Transfection with the catalytic subunit of protein kinase A (PKA-C) activated SS promoter-CAT activiting indicating that the defect in cAMP-dependent signaling in 1027B2 cells lies at the level of PKA-C. PKA-C overexpression failed to alter the ratio of processed SS - 14 and SS - 28. These results demonstrate that SS - 14, SS - 28, and PSS([1-10]) are independently synthesized from PSS and that efficient precursor processing can occur within the constitutive secretory pathway in the relative absence of dense core secretory vesicles.
KW - Processing
KW - Secretory pathways
KW - Somatostatin
KW - Tumor cells
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U2 - 10.1016/S0303-7207(97)00107-X
DO - 10.1016/S0303-7207(97)00107-X
M3 - Article
C2 - 9296377
AN - SCOPUS:0030921526
SN - 0303-7207
VL - 131
SP - 183
EP - 194
JO - Molecular and Cellular Endocrinology
JF - Molecular and Cellular Endocrinology
IS - 2
ER -