The relationship between intracellular UDP-N-acetyl hexosamine nucleotide pool and monoclonal antibody production in a mouse hybridoma.
The effect of intracellular UDP-N-acetyl hexosamine accumulation in a murine hybridoma was investigated using tunicamycin and ammonium chloride. The treatment of cells with tunicamycin resulted in the inhibition of glycosylation of the secreted monoclonal antibody (IgGl) and cell growth arrest. Tunicamycin concentrations of 0.01-0.1 microgram ml-1 resulted in the formation of both glycosylated and non-glycosylated heavy chains of the immunoglobulin, whereas complete inhibition of protein glycosylation was observed at higher concentrations of tunicamycin. Tunicamycin treatment also resulted in a dose dependent accumulation of UDP-N-acetyl hexosamine. It was concluded that the specific monoclonal antibody production rate (qMab) was not dependent on the extent of glycosylation. Treatment of cells with NH4Cl also resulted in dose dependent accumulation of UDP-N-acetyl hexosamine. Supplementation of cultures with 10 mM NH4Cl resulted in a 40% reduction in cell growth rate and a 36% increase in the qMab. The data suggest that the reduction in growth rate and not UDP-N-acetyl hexosamine accumulation was the cause of increased qMab in the ammonium supplemented cultures. Ammonium chloride did not affect the extent of protein glycosylation. We conclude that UDP-N-acetyl hexosamine does not act as a mediator of enhanced rates of monoclonal antibody synthesis in the hybridoma cell culture system.
Barnabé
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Department of Microbiology, University of Manitoba, Winnipeg, Canada.
Butler
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M
D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D006031:Glycosylation; D006825:Hybridomas / Q000276:immunology*; D007143:Immunoglobulin Heavy Chains / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D009711:Nucleotides / Q000378:metabolism*; D014415:Tunicamycin / Q000494:pharmacology; D014539:Uridine Diphosphate Sugars / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D007143:Immunoglobulin Heavy Chains; D009711:Nucleotides; D014539:Uridine Diphosphate Sugars; D000643:Ammonium Chloride; D014415:Tunicamycin