Processing and secretion of lysosomal acid alpha-glucosidase in Tetrahymena wild type and secretion-deficient mutant cells.
The proteolytic processing and secretion of a lysosomal enzyme, acid alpha-glucosidase, was studied by pulse-chase labeling with [35S]methionine in Tetrahymena thermophila CU-399 cells treated with ammonium chloride. This cell secreted a large amount of acid alpha-glucosidase into the cultured medium during starvation. The secretion was found to be repressed by addition of ammonium chloride (NH4Cl). Acid alpha-glucosidase was produced as a precursor form (108 kDa) and then processed to a mature polypeptide (105 kDa) within 60 min. This mature enzyme was secreted into the media within 2-3 h after chase, whereas the precursor form was not secreted by either control cells or NH4Cl-treated cells. NH4Cl did not affect the processing of the precursor acid alpha-glucosidase. Processing profile of this enzyme was apparently indistinguishable from that of the mutant MS-1 defective in lysosomal enzyme secretion. Furthermore, the purified extracellular (CU-399) and intracellular (MS-1) acid alpha-glucosidases were the same in molecular mass (105 kDa) and enzymatic properties. They contained no mannose 6-phosphate residues in N-linked oligosaccharides. These results suggested that unlike mammalian cells, Tetrahymena acid alpha-glucosidase may be transferred to lysosomes by a mannose 6-phosphate receptor-independent mechanism, and also that low pH was not essential for the proteolytic processing of precursor polypeptide.
Banno
Y
Y
Department of Biochemistry, Gifu University School of Medicine, Japan.
Okano
Y
Y
Furukawa
K
K
Tiedtke
A
A
Kobata
A
A
Nozawa
Y
Y
D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000032:analysis; D005087:Glucan 1,4-alpha-Glucosidase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D013768:Tetrahymena / Q000187:drug effects / Q000378:metabolism*
D002241:Carbohydrates; D000643:Ammonium Chloride; D005087:Glucan 1,4-alpha-Glucosidase