Kinetics of ?2-macroglobulin endocytosis and degradation in mutant and wild-type Chinese hamster ovary cells

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Abstract
The production of Chinese hamster ovary (CHO) cell mutants which are defective in endocytosis has led to a greater understanding of the process by which cells sort ligands and their receptors. Robbins and coworkers have obtained CHO mutants which are resistant to diphtheria toxin, defective in the delivery of endocytosed lysosomal enzymes to lysosomes, and have a decreased uptake of iron from transferrin (Robbins et al.: 1. Cell Biol. 96:1064–1071, 1983). We have previously shown that these CHO mutants are markedly deficient in the acidification of early endocytic compartments (Yamashiro and Maxfield: J. Cell Biol. 105:2713–2721, 1987). In this study we examined the endocytosis of α2-macroglobulin (α2M) to determine whether the defects in early endosome acidification would alter the processing of this ligand. We found that the CHO mutants DTG 1–5–4 and DTF 1–5–1 bind, internalize, and degrade 125l-α2M in a manner similar to the wild-type cells. We also found that the CHO mutants retain the ability to recycle the receptors for α2M. Since the binding of α2M is greatly reduced at mildly acidic pH (≈︁6.8), only slight acidification of the endosomal compartment should be sufficient to achieve sorting of a2M from its receptor. In contrast, lysosomal enzymes require more acidic conditions (pH < 6.0) for dissociation. The different behavior of the two ligands provides biochemical evidence for a partial (but not complete) defect in early endosome acidification in the mutants. The data also indicate that pH regulation in a relatively narrow range can achieve differential sorting of various ligands.