Processing and routing of a membrane-anchored form of proneuropeptide Y.
To investigate factors governing proteolytic processing and routing of biologically active peptides in the secretory pathway, cDNAs for preproneuropeptide Y (preproNPY) and preproneuropeptide Y fused to a membrane anchor were transfected into pituitary cells. The anchor was the transmembrane and COOH-terminal cytoplasmic domain of peptidylglycine alpha-amidating monooxygenase (PAM); these domains are essential for correct routing of integral membrane forms of PAM. Like proneuropeptide Y (proNPY), the integral membrane form of proNPY was a good substrate for the endogenous prohormone convertases, yielding soluble NPY stored in regulated secretory granules. Tethering of proNPY to the membrane resulted in only a small delay in the rate of cleavage to produce mature NPY and in the arrival of NPY in regulated secretory granules. In contrast, the COOH-terminal region of proNPY remained attached to the transmembrane/COOH-terminal domain of PAM and was rerouted to the vicinity of the trans-Golgi network, where integral membrane forms of PAM are concentrated. Thus, the COOH-terminal of proNPY cannot override the signals in the PAM membrane anchor.
Milgram
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Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205-2185, USA.
Chang
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Mains
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D000818:Animals; D001665:Binding Sites; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D007150:Immunohistochemistry; D051379:Mice; D006899:Mixed Function Oxygenases / Q000378:metabolism; D009097:Multienzyme Complexes; D009478:Neuropeptide Y / Q000276:immunology / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D010911:Pituitary Neoplasms / Q000188:drug therapy / Q000473:pathology; D011233:Precipitin Tests / Q000379:methods; D011498:Protein Precursors / Q000276:immunology / Q000378:metabolism*; D011817:Rabbits; D013347:Subcellular Fractions; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D009097:Multienzyme Complexes; D009478:Neuropeptide Y; D010446:Peptide Fragments; D011498:Protein Precursors; C081158:proneuropeptide Y; D006899:Mixed Function Oxygenases; C035189:peptidylglycine monooxygenase; D013755:Tetradecanoylphorbol Acetate