Novel markers for constitutive secretion used to show that tissue plasminogen activator is sorted to the regulated pathway in transfected PC12 cells.
The rat pheochromocytoma cell line PC12 contains two distinct pathways of protein secretion. Proteins secreted via the regulated pathway are stored in secretory vesicles and exocytosed only in response to a specific signal, whereas proteins secreted via the constitutive pathway are exported continuously. Analysis of regulated secretion of a heterologous protein in this system often relies on comparison of secretion rates with those of endogenous proteins known to be secreted via the constitutive route. In order to improve these controls, we have evaluated a number of secreted enzymes, selected for the sensitivity and convenience of their assays, as transgenic markers for the constitutive pathway. We show that both human-secreted placental alkaline phosphatase (SEAP) and bacterial beta-lactamase operate in this way in transfected PC12 cells. In contrast, transfected human tissue plasminogen activator (tPA) is shown to be sorted to the regulated pathway.
Harrison
T M
TM
Department of Biochemistry, University of Leicester, U.K.
Chidgey
M A
MA
Uff
S
S
D000469:Alkaline Phosphatase / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001358:Avian Sarcoma Viruses / Q000235:genetics; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D015415:Biomarkers; D002217:Carbachol / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D015870:Gene Expression / Q000502:physiology; D006801:Humans; D016716:PC12 Cells / Q000201:enzymology / Q000378:metabolism*; D051381:Rats; D010959:Tissue Plasminogen Activator / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology; D014162:Transfection; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism
D015415:Biomarkers; D002217:Carbachol; D000469:Alkaline Phosphatase; D010959:Tissue Plasminogen Activator; D001618:beta-Lactamases