Modulation of the carbohydrate moiety of thyroglobulin by thyrotropin and calcium in Fisher rat thyroid line-5 cells.
Thyroglobulin secreted in the medium by Fisher rat thyroid line-5 (FRTL-5) cells cultured in the presence of thyroid stimulating hormone (TSH) shows a slower electrophoretic mobility in sodium dodecyl sulfate-polyacrylamide gel electrophoresis and a higher density position in a CsCl gradient than thyroglobulin secreted by FRTL-5 cells cultured in the absence of TSH for 5-7 days. Such a TSH effect is much less or not evident when secreted thyroglobulin is digested with peptide N-glycohydrolase F or when intracellular thyroglobulin is compared. Intracellular thyroglobulin migrates faster than thyroglobulin secreted either in the presence or in the absence of TSH. Evaluation of the mannose and galactose content of thyroglobulin demonstrates that intracellular thyroglobulin has more mannose and less galactose than extracellular thyroglobulin; it also shows that TSH decreases the mannose content of thyroglobulin while increasing its galactose content. Bio-Gel P6 chromatography shows that TSH increases the complex type carbohydrate chains while decreasing the high mannose chains in the secreted thyroglobulin. High mannose type oligosaccharides were characterized by fast atom bombardment-mass spectrometry analysis. Treatment with the calcium ionophore A23187 (5 microM) of FRTL-5 cells cultured with or without TSH causes the appearance of a "fast" migrating form of thyroglobulinin in the culture medium. Bio-Gel P6 chromatography shows that A23187 causes a dramatic decrease of the complex carbohydrate chains of the secreted thyroglobulin.
Centro di Endocrinologia e Oncologia Sperimentale del Consiglio Nazionale delle Ricerche, Napoli, Italy.
B
B
Di Jeso
D
D
Liguoro
P
P
Ferranti
M
M
Marinaccio
R
R
Acquaviva
S
S
Formisano
E
E
Consiglio
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000818:Animals; D001345:Autoradiography; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology*; D002460:Cell Line; D002499:Centrifugation, Density Gradient; D005690:Galactose / Q000378:metabolism*; D006031:Glycosylation; D008358:Mannose / Q000378:metabolism*; D008715:Methionine / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D051381:Rats; D011916:Rats, Inbred F344; D016339:Spectrometry, Mass, Fast Atom Bombardment; D013462:Sulfur Radioisotopes; D017026:Swainsonine / Q000494:pharmacology; D013954:Thyroglobulin / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D013961:Thyroid Gland / Q000187:drug effects / Q000378:metabolism*; D013972:Thyrotropin / Q000494:pharmacology*; D014316:Tritium
D013462:Sulfur Radioisotopes; D014316:Tritium; D000001:Calcimycin; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D013972:Thyrotropin; D013954:Thyroglobulin; D008715:Methionine; D008358:Mannose; D017026:Swainsonine; D002118:Calcium; D005690:Galactose