Proteolytic processing of pro-opiomelanocortin occurs in acidifying secretory granules of AtT-20 cells.
Using antibodies specific for pro-opiomelanocortin (POMC), amidated joining peptide (JP), and the prohormone convertase PC1, we showed immunocytochemically that PC1 in a corticotrophic tumor cell line, AtT-20, was co-localized either with POMC or with amidated JP in secretory granules, and also confirmed that POMC was cleaved mainly in secretory granules. Analysis using DAMP (3- [2,4-dinitroanilino]-3'-amino-N-methyldipropylamine) as the pH probe suggested a correlation between POMC processing and acidic pH in the secretory granules. Bafilomycin A1, a specific inhibitor of vacuolar-type H(+)-ATPase, completely inhibited POMC processing and caused constitutive secretion of the unprocessed precursor. By contrast, chloroquine, a weak base that is known to neutralize acidic organelles, was unable to inhibit POMC processing. Electron microscopic analysis revealed that, in AtT-20 cells treated with bafilomycin A1, the trans-Golgi cisternae were dilated and few secretory granules were present in the cytoplasm. These observations suggest that acidic pH provides a favorable environment for proteolytic processing of POMC by PC1 but is not required, and that integrity of the trans-Golgi network and sorting of POMC into secretory granules are important for POMC processing.
Tanaka
S
S
Department of Cell Biology, Gunma University, Maebashi, Japan.
Yora
T
T
Nakayama
K
K
Inoue
K
K
Kurosumi
K
K
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D016282:Aspartic Acid Endopeptidases / Q000378:metabolism; D002738:Chloroquine / Q000494:pharmacology; D003594:Cytoplasmic Granules / Q000378:metabolism*; D010450:Endopeptidases / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018942:Macrolides; D051379:Mice; D015388:Organelles / Q000187:drug effects; D010446:Peptide Fragments / Q000378:metabolism; D011333:Pro-Opiomelanocortin / Q000378:metabolism*; D045664:Proprotein Convertase 1; D043484:Proprotein Convertases; D011499:Protein Processing, Post-Translational; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors; D014407:Tumor Cells, Cultured; D025262:Vacuolar Proton-Translocating ATPases
D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D018942:Macrolides; D010446:Peptide Fragments; C057620:bafilomycin A; D011333:Pro-Opiomelanocortin; C064114:pro-opiomelanocortin joining peptide(79-108); D002738:Chloroquine; D010450:Endopeptidases; D043484:Proprotein Convertases; C473656:Pcsk1 protein, mouse; D045664:Proprotein Convertase 1; D016282:Aspartic Acid Endopeptidases; D025262:Vacuolar Proton-Translocating ATPases; D006180:Proton-Translocating ATPases