Acute inhibition of carboxypeptidase E expression in AtT-20 cells does not affect regulated secretion of ACTH.
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.
Kemppainen
Robert J
RJ
Departments of Anatomy, Physiology, and Pharmacology, College of Veterinary Medicine, Auburn University, Auburn, AL 36849, USA.
Behrend
Ellen N
EN
D000324:Adrenocorticotropic Hormone / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D043423:Carboxypeptidase H / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D052680:Corticotrophs / Q000201:enzymology* / Q000378:metabolism*; D051379:Mice; D016133:Polymerase Chain Reaction; D034741:RNA, Small Interfering
D034741:RNA, Small Interfering; D000324:Adrenocorticotropic Hormone; D043423:Carboxypeptidase H