Biosynthesis and Secretion of α1‐Antitrypsin in Primary Cultures of Rat Hepatocytes

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Abstract
The biosynthesis and secretion of α1-antitrypsin was studied in rat hepatocyte primary cultures. After labeling with [35S]methionine an α1-antitrypsin with an apparent molecular weight of 49000 estimated by sodium dodecyl sulfate/polyacrylamide slab gel electrophoresis was immunoprecipitated from the cell homogenate. This intracellular form of α1-antitrypsin could be deglycosylated by endoglycosidase H treatment indicating that its oligosaccharide chains were of the high-mannose type. Pulse-chase experiments showed that about 30 min after its synthesis the transformation of the 49000-Mrα1-antitrypsin to a protein with an apparent molecular weight of 54000 began. Only this 54000-Mr protein was secreted by the hepatocytes. The 54000-Mrα1-antitrypsin was not sensitive to endoglycosidase H, but sensitive to neuraminidase, and it incorporated [3H]galactose and [3H]fucose indicating that its oligosaccharide chains were of the complex type. In the presence of tunicamycin, which blocks the formation of N-asparagine-linked oligosaccharide chains, an unglycosylated α1-antitrypsin with an apparent molecular weight of 41000 was found in the cells as well as in the medium. However, tunicamycin decreased the secretion of α1-antitrypsin by 60–70%, whereas the secretion of albumin remained unaffected. In the presence of colchicine the secretion of both α1-antitrypsin and albumin was impaired. The results demonstrate the importance of glycosylation for the secretion of α1-antitrypsin.