Effect of serum on phenylalanine hydroxylase levels in cultured hepatoma cells.
Continued high levels of phenylalanine hydroxylase in cultured H4-II-E-C3 rat hepatoma cells require either serum or glucocorticoids in the culture medium. Upon withdrawal of serum, cellular phenylalanine hydroxylase levels decay exponentially with a half-life of 22 hours for about 60 hours, after which time a low, constant enzyme content persists for at least 96 hours. This decline of phenylalanine hydroxylase is fully reversible; normal enzyme levels are restored in a time- and dosage-dependent fashion upon addition of serum  to basal cultures. The serum factor is nondialyzable and moderately heat-stable. The stimulation by serum of the phenylalanine hydroxylas content of basal cultures is blocked by 3-[2-(3,5-dimethyl-2-oxocyclohexyl)-2-hydroxyethyl]glutarimide and requires ongoing cellular protein synthesis. When added to the enzyme-assay mixture in vitro, serum does not alter the phenylalanine hydroxylase activity of extracts from basal cultures. Three lines of evidence suggest that serum contains a nonsteroidal phenylalanine hydroxylase stimulatory components(s): (a) glucocorticoid antagonists inhibit less than one-half of the biological activity of serum; (b) exhaustive extraction of endogenous serum glucocorticoids with charcoal reduces the activity of serum to about one-half of control values; and (c) the stimulatory effects of charcoal reduces the values; and (c) the stimulatory effects of charcoal-extracted serum and hydrocortisone are additive. The phenylalanine hydroxylase stimulatory activities of the charcoal-extracted sera from four mammalian species and from three stages in development in one mammalian species are comparable. A survey of partially purified preparations of a number of known hormones failed to reveal any one capable of elevating the phenylalanine hydroxylas levels of basal cultures in a manner comparable to that of charcoal-extracted serum.
D000818:Animals; D001769:Blood; D006528:Carcinoma, Hepatocellular / Q000201:enzymology*; D002417:Cattle; D002460:Cell Line; D003300:Copper / Q000494:pharmacology; D003470:Culture Media; D004789:Enzyme Activation / Q000187:drug effects; D005934:Glucagon / Q000494:pharmacology; D005938:Glucocorticoids / Q000494:pharmacology; D006854:Hydrocortisone / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology; D007501:Iron / Q000494:pharmacology; D008113:Liver Neoplasms; D009374:Neoplasms, Experimental / Q000201:enzymology; D010651:Phenylalanine Hydroxylase / Q000096:biosynthesis / Q000378:metabolism*; D051381:Rats; D013045:Species Specificity; D015032:Zinc / Q000494:pharmacology
D003470:Culture Media; D005938:Glucocorticoids; D007328:Insulin; D003300:Copper; D005934:Glucagon; D007501:Iron; D010651:Phenylalanine Hydroxylase; D015032:Zinc; D006854:Hydrocortisone