PC5-A-mediated processing of pro-neurotensin in early compartments of the regulated secretory pathway of PC5-transfected PC12 cells.
Among the members of the proprotein convertase (PC) family, PC1 and PC2 have well established roles as prohormone convertases. Another good candidate for this role is PC5-A that has been shown to be present in the regulated secretory pathway of certain neuroendocrine tissues, but evidence that it can process prohormones is lacking. To determine whether PC5-A could function as a prohormone convertase and to compare its cleavage specificity with that of PC1 and PC2, we stably transfected the rat pheochromocytoma PC12 cell line with PC5-A and analyzed the biosynthesis and subcellular localization of the enzyme, as well as its ability to process pro-neurotensin/neuromedin N (pro-NT/NN) into active peptides. Our data showed that in transfected PC12 cells, PC5-A was converted from its 126-kDa precursor form into a 117-kDa mature form and, to a lesser extent, into a C-terminally truncated 65-kDa form of the 117-kDa product. Metabolic and immunochemical studies showed that PC5-A was sorted to early compartments of the regulated secretory pathway where it colocalized with immunoreactive NT. Furthermore, pro-NT/NN was processed in these compartments according to a pattern that differed from that previously described in PC1- and PC2-transfected PC12 cells. This pattern resembled that previously reported for pro-NT/NN processing in the adrenal medulla, a tissue known to express high levels of PC5-A. Altogether, these data demonstrate for the first time the ability of PC5-A to function as a prohormone convertase in the regulated secretory pathway and suggest a role for this enzyme in the physiological processing of pro-NT/NN.
Barbero
P
P
Institut de Pharmacologie Moléculaire et Cellulaire du CNRS, Université de Nice-Sophia Antipolis, Sophia Antipolis, 660 Route des Lucioles, 06560 Valbonne, France.
Rovère
C
C
De Bie
I
I
Seidah
N
N
Beaudet
A
A
Kitabgi
P
P
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D008969:Molecular Sequence Data; D009496:Neurotensin / Q000378:metabolism*; D016716:PC12 Cells; D045702:Proprotein Convertase 5; D011498:Protein Precursors / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D051381:Rats; D012697:Serine Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D011498:Protein Precursors; D009496:Neurotensin; D045702:Proprotein Convertase 5; D012697:Serine Endopeptidases; C473807:propeptide convertase 5, rat