Expression of porcine pro-opiomelanocortin in mouse neuroblastoma (Neuro2A) cells: targeting of the foreign neuropeptide to dense-core vesicles.
Pro-opiomelanocortin (POMC) is the precursor to several pituitary hormones and neuropeptides including adrenocorticotropic hormone (ACTH) and beta-endorphin (beta-END). In neuroendocrine cells, peptide hormones and neuropeptides are targeted to the dense-core vesicles of the regulated secretory pathway. These vesicles are transported to the ends of cellular extensions where they are stored until they release their content upon external stimulation of the cell. In order to study the cellular mechanisms involved in targeting of neuropeptides, we have expressed POMC in Neuro2A cells, a cell line of neural origin. Using immunofluorescence labeling and immunoelectron microscopy we show that in Neuro2A cells POMC is packaged in dense-core vesicles which accumulate at the tips of cellular processes. Intracellular accumulation of POMC was not observed in NIH 3T3 fibroblasts. When a soluble form of an integral membrane protein, neutral endopeptidase (E.C. 3.4.24.11) (secNEP), was expressed in Neuro2A cells, the protein was found to be constitutively secreted without prior accumulation in dense-core vesicles. Our results suggest that in Neuro2A cells, targeting to the regulated secretory pathway is restricted to peptide hormones and neuropeptides and establish this cell line as a valid model for studying the molecular events involved in neuropeptide sorting into the regulated secretory pathway.
Chevrier
D
D
Département de Biochimie, Faculté de Médecine, Université de Montréal, Canada.
Fournier
H
H
Nault
C
C
Zollinger
M
M
Crine
P
P
Boileau
G
G
D000818:Animals; D000911:Antibodies, Monoclonal; D002460:Cell Line; D003594:Cytoplasmic Granules / Q000648:ultrastructure*; D005455:Fluorescent Antibody Technique; D015151:Immunoblotting; D051379:Mice; D008854:Microscopy, Electron; D008970:Molecular Weight; D009447:Neuroblastoma; D011333:Pro-Opiomelanocortin / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D013552:Swine; D014162:Transfection
D000911:Antibodies, Monoclonal; D011333:Pro-Opiomelanocortin