Prohormone Convertases 1 and 2 Process ProPACAP and Generate Matured, Bioactive PACAP38 and PACAP27 in Transfected Rat Pituitary GH4C1 Cells
| Autor: | Akira Arimura, Min Li, Anikó Somogyvári-Vigh, Yujin Shuto |
|---|---|
| Rok vydání: | 1999 |
| Předmět: |
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 |
| Zdroj: | Neuroendocrinology. 69:217-226 |
| ISSN: | 1423-0194 0028-3835 |
| DOI: | 10.1159/000054422 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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