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Prohormone Convertases 1 and 2 Process ProPACAP and Generate Matured, Bioactive PACAP38 and PACAP27 in Transfected Rat Pituitary GH4C1 Cells
- Source :
- Neuroendocrinology. 69:217-226
- Publication Year :
- 1999
- Publisher :
- S. Karger AG, 1999.
-
Abstract
- Pituitary adenylate cyclase-activating polypetide (PACAP) exists in two amidated forms, PACAP38 and PACAP27, which are expressed in the magnocellular and parvocellular neurons of the paraventricular nucleus (PVN) and the magnocellular neurons of the supraoptic nucleus (SON) of the hypothalamus. The prohormone convertases PC1 and PC2, subtilisin-like PCs of the Kex2 family, are expressed in neuroendocrine cells. Immunocytochemistry and in situ hybridization of PC1 and PC2 in the hypothalamus have shown that PC1 and PC2 are also present in the PVN and SON. Therefore, it is possible that the precursor of PACAP is processed by PC1 and/or PC2 in the hypothalamic nuclei and then converted to its mature forms. To test this hypothesis, rat pituitary GH4C1 cells were supertransfected with human PACAP cDNA and either rat PC1 or PC2 cDNA. The acid extracts of these cells were analyzed by reversed-phase HPLC for proPACAP, PACAP38 and/or PACAP27 radioimmunoassays using three antibodies with different recognition sites, and then bioassayed for the ability to stimulate adenylate cyclase. The cells transfected with PACAP cDNA alone yielded PACAP-like immunoreactivity (PACAP-li) corresponding to molecular weights between 15 and 20 kDa without PACAP bioactivity. Cotransfection of these cells with PC1 or PC2 generated PACAP-li, which coeluted with synthetic PACAP38 and PACAP27, respectively. Western blot also revealed 4.5- and 3.0-kDa PACAP-li bands, which correspond to the molecular weights of PACAP38 and PACAP27, respectively. The HPLC fractions containing PACAP-li, which were coeluted with synthetic PACAP38 and PACAP27, showed marked bioactivities. These findings suggest that the precursor of PACAP expressed in the PVN and SON of the hypothalamus could be efficiently processed by PC1 and PC2, and then converted to mature, bioactive PACAP38 and PACAP27.
- Subjects :
- endocrine system
medicine.medical_specialty
genetic structures
Endocrine and Autonomic Systems
Endocrinology, Diabetes and Metabolism
Adenylate kinase
Transfection
Biology
Molecular biology
ADCY10
Rat Pituitary
Cellular and Molecular Neuroscience
Endocrinology
nervous system
Parvocellular cell
Internal medicine
medicine
Prohormone Convertases
sense organs
hormones, hormone substitutes, and hormone antagonists
Subjects
Details
- ISSN :
- 14230194 and 00283835
- Volume :
- 69
- Database :
- OpenAIRE
- Journal :
- Neuroendocrinology
- Accession number :
- edsair.doi...........dd743f5e93a734aad3692cec73313509