POMC-related products in the intermediate pituitary of the amphibian, Bufo marinus: differential subcellular processing in the Golgi and secretory granules.
In the intermediate pituitary of the anuran amphibian, Bufo marinus, the N-acetylation of ACTH(1-13)-NH2 to yield alpha-MSH occurs as a cosecretory processing event, whereas the N-acetylation of beta-endorphin occurs as a posttranslational processing event. To understand how these two N-acetylation reactions are segregated, B. marinus intermediate pituitary cells were analyzed by immunogold labeling electron microscopy, and by using an ultracentrifugation procedure. The immunogold labeling studies indicated that ACTH(1-13)-NH2-related immunoreactivity was colocalized with N-acetylated beta-endorphin-related immunoreactivity in secretory granules. Furthermore, ACTH(1-13)-NH2-related immunoreactivity was not detected in either the ER or the Golgi. N-Acetylated beta-endorphin-related immunoreactivity, however, was detected in the Golgi. Ultracentrifugation analysis revealed that in an ER/microsomal fraction, beta-LPH-sized and nonacetylated beta-endorphin-sized immunoreactive material were present in a molar ratio of 1:2. No N-acetylated forms of beta-endorphin were detected in the ER/microsomal fraction. In a Golgi/secretory granule fraction, the molar ratio of beta-LPH to beta-endorphin was 1:9 with 58% of the beta-endorphin being N-acetylated. Collectively, these data support the following hypotheses. The proteolytic cleavage of ACTH (1-39) to yield ACTH (1-13)-NH2 is a late processing event occurring in secretory granules. The cleavage of beta-LPH to yield nonacetylated beta-endorphin is an early processing event that may occur in the ER or the Golgi. Because N-acetylated beta-endorphin and nonacetylated ACTH(1-13)-NH2 are colocalized in secretory granules, it appears, therefore, that the N-acetylation of beta-endorphin is completed prior to loading into secretory granules. Thus, there is a spatial and temporal separation of the posttranslational processing events associated with the beta-LPH portion and ACTH portion of the POMC biosynthetic pathway in amphibian intermediate pituitary cells.
Steveson
T C
TC
Johns Hopkins School of Medicine, Department of Neuroscience, Baltimore, MD 21205, USA.
Dores
R M
RM
D000107:Acetylation; D000818:Animals; D002024:Bufo marinus / Q000378:metabolism*; D003593:Cytoplasm / Q000737:chemistry* / Q000648:ultrastructure; D003594:Cytoplasmic Granules / Q000737:chemistry / Q000648:ultrastructure; D006056:Golgi Apparatus / Q000737:chemistry / Q000648:ultrastructure; D016253:Microscopy, Immunoelectron; D010446:Peptide Fragments / Q000302:isolation & purification*; D010902:Pituitary Gland / Q000737:chemistry / Q000378:metabolism*; D011333:Pro-Opiomelanocortin / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D013347:Subcellular Fractions / Q000737:chemistry; D000521:alpha-MSH / Q000031:analogs & derivatives* / Q000302:isolation & purification; D001615:beta-Endorphin / Q000031:analogs & derivatives* / Q000302:isolation & purification
D010446:Peptide Fragments; D000521:alpha-MSH; D001615:beta-Endorphin; D011333:Pro-Opiomelanocortin; C035101:N-acetyl-beta-endorphin