Acute inhibition of carboxypeptidase E expression in AtT-20 cells does not affect regulated secretion of ACTH

• Published in: Regulatory peptides 2010-12, Vol.165 (2), p.174-179
• Main Authors: Kemppainen, Robert J., Behrend, Ellen N.
• Format: Article
• Language: English
• Subjects: ACTH; Adrenocorticotropic Hormone - secretion; Animals; AtT-20 cells; Biological and medical sciences; Blotting, Western; Carboxypeptidase E; Carboxypeptidase H - genetics; Carboxypeptidase H - metabolism; Cell Line; Corticotrophs - enzymology; Corticotrophs - metabolism; Corticotrophs - secretion; Fundamental and applied biological sciences. Psychology; Mice; Polymerase Chain Reaction; Proopiomelanocortin; RNA, Small Interfering; Secretion; Sorting; Vertebrates: endocrinology
• ISSN: 0167-0115; 1873-1686
• DOI: 10.1016/j.regpep.2010.07.162

Carboxypeptidase E (CPE) is an exopeptidase that removes C-terminal basic amino acids from a variety of bioactive peptides. In addition to this role, data obtained in recent years has supported a potential function for CPE as a sorting receptor, helping direct peptides destined for regulated secretion from the trans-Golgi to granules in preparation for release. This possible sorting function was assessed using mouse AtT-20 cells, a well-established corticotroph cell line that synthesizes and releases POMC/ACTH in regulated fashion. Cells that were treated with siRNA to Cpe effectively suppressed CPE expression. ACTH was released in a regulated fashion from CPE-depleted cells in response to two secretagogues, 8-bromo-cyclic AMP and corticotrophin-releasing hormone. POMC/ACTH content of CPE-depleted cells was higher than that of control cells, but both released a similar percentage of ACTH content in response to secretagogue addition. Cells depleted of CPE generally secreted more high-molecular weight forms of POMC/ACTH under basal conditions than control cells; however, the CPE-depleted cells responded to a secretagogue by releasing newly synthesized ACTH 1-39 in a manner similar to controls. These results, whereby RNAi was used to acutely suppress CPE, do not support a role for this protein as necessary for or central to sorting of POMC/ACTH to the regulated secretory pathway in AtT-20 cells. ► Carboxypeptidase E (CPE) has been proposed to act as a sorting receptor ► Suppression of CPE in AtT-20 cells did not affect regulated release of ACTH ► Constitutive release of precursor forms of ACTH was increased by CPE suppression