A certain step of proteolytic processing of proopiomelanocortin occurs during the transition between two distinct stages of secretory granule maturation in rat anterior pituitary corticotrophs.
To obtain antibodies specific for precursor POMC and its cleaved peptides, we prepared synthetic peptides corresponding to the cleavage site (ST-1) between ACTH and beta-lipotropic hormone moieties of POMC and to amidated joining peptide (JP; ST-3) and generated their polyclonal antibodies in rabbits. The anti-ST-1 serum immunoprecipitated only POMC in the extract of AtT-20 cells. In addition to a similar immunoprecipitation examination, an immunoabsorption test showed that anti-ST-3 serum recognizes cleaved peptides, 16K amino-terminal peptide, and JP, but not POMC precursor. Double immunolabeling with gold particles of different sizes for the precursor POMC and its cleaved peptides revealed that the former was localized in electron-dense secretory granules, whereas the latter was found in the electron-lucent secretory granules in the same ultrathin cryosections. Colocalization of POMC and its cleaved peptides in the same secretory granules was rarely observed. Only POMC immunolabeling was detected in the Golgi cisternae. These results suggest that a certain step of proteolytic processing of POMC occurs in the secretory granules during the transition between the two distinct stages of secretory granule maturation in the rat anterior pituitary corticotroph.
Tanaka
S
S
Department of Morphology, Gunma University, Maebashi, Japan.
Kurosumi
K
K
D000324:Adrenocorticotropic Hormone / Q000032:analysis / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D003594:Cytoplasmic Granules / Q000737:chemistry / Q000502:physiology*; D010450:Endopeptidases / Q000378:metabolism*; D006056:Golgi Apparatus / Q000737:chemistry; D007150:Immunohistochemistry; D007163:Immunosorbent Techniques; D008297:Male; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis / Q000378:metabolism; D010903:Pituitary Gland, Anterior / Q000737:chemistry / Q000648:ultrastructure*; D011333:Pro-Opiomelanocortin / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains
D010446:Peptide Fragments; D011333:Pro-Opiomelanocortin; D000324:Adrenocorticotropic Hormone; D010450:Endopeptidases