Impaired secretion and fluid-phase endocytosis in the End4 mutant of Chinese hamster ovary cells.
Mutant V.24.1 defines the End4 complementation group of temperature-sensitive Chinese hamster ovary cell mutants selected for resistance to protein toxins. We investigated the secretory pathway in the mutant cells and found: 1) The hemagglutinin of influenza virus failed to reach the plasma membrane and was retained in a form sensitive to endoglycosidase H at the restrictive temperature. 2) Transferrin receptors synthesized at the restrictive temperature remained sensitive to endoglycosidase H. 3) Secretion of total soluble protein into the medium was strongly reduced at high temperature. These data indicate that V.24.1 cells are defective in secretion at the restrictive temperature. To see what effect the lesion had on the endocytic pathway, we measured the accumulation and recycling of the fluid-phase marker horseradish peroxidase. Accumulation was inhibited by 50% while recycling was barely affected, suggesting that the rate of fluid-phase endocytosis was reduced. We previously showed that the clathrin-coated pit pathway of endocytosis was not affected in the mutant, indicated by a normal transferrin cycle (Colbaugh, P. A., Stookey, M., and Draper, R. K. (1989) J. Cell Biol. 108, 2211-2219). Thus, the secretory lesion correlates with reduced fluid-phase endocytosis without impairing the clathrin-dependent pathway of receptor-mediated endocytosis. We also investigated the delivery of endocytosed material to lysosomes and found that delivery was partially, but not completely, impaired in the mutant. This suggests that endocytosed material can enter lysosomes, although slowly, in the absence of a functional secretory pathway.
Wang
R H
RH
Molecular and Cell Biology Program, University of Texas, Dallas, Richardson 75083-0688.
Colbaugh
P A
PA
Kao
C Y
CY
Rutledge
E A
EA
Draper
R K
RK
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D004705:Endocytosis; D005260:Female; D005816:Genetic Complementation Test; D006735:Horseradish Peroxidase / Q000378:metabolism; D007700:Kinetics; D008247:Lysosomes / Q000378:metabolism; D009154:Mutation; D010053:Ovary; D014176:Protein Biosynthesis; D011506:Proteins / Q000378:metabolism; D013696:Temperature; D001616:beta-Galactosidase / Q000378:metabolism
D011506:Proteins; D006735:Horseradish Peroxidase; D001616:beta-Galactosidase