Intracellular transport of secretory and membrane proteins in hepatoma cells infected by vesicular stomatitis virus

Abstract

Rat hepatoma cells which secrete several serum proteins have been infected with a temperature sensitive mutant of Vesicular Stomatitis Virus, which is defective in RNA replication and transcription. These infected cells, grown at 39.5°C, continue normal secretion of albumin and transferrin, so that synthesis and maturation of secretory and membrane proteins, including the VSV G protein, can be studied in the same cells. Serum proteins are not secreted from the cells at the same rate; half of the newly synthesized albumin appears in the medium after 23 minutes, while half of the glycoprotein transferrin is secreted only after about 40 minutes. By contrast, two membrane proteins—the VSV G glycoprotein and a high molecular weight hepatoma surface protein—are detected at the cell surface approximately 23 minutes after their synthesis. The differences in the rate of externalization among the secretory and membrane glycoproteins are due to differences in the rate of transport from the rough endoplasmic reticulum (ER) into or through the Golgi region, as judged by the time required for maturation of the asn-linked oligosaccharides. Unglycosylated VSV G does not reach the cell surface, while unglycosylated transferrin is secreted at the same rate as the glycosylated species. Thus neither the synthesis nor the presence of the asn-linked oligosaccharide chain are determinants in the rate of transferrin secretion. The monovalent ionophore monensin inhibits both secretion of serum proteins and migration of membrane proteins to the cell surface. However, the intracellular transport of transferrin is blocked in a stage where the molecule is still sensitive to endo-β-N-acetylglucosaminidase H, while VSV G reaches a stage in which it is resistant to the enzyme. These results indicate that at least part of the intracellular pathway may be different for membrane and secretory proteins and glycoproteins.