Structural and functional changes of lysosomal acid alpha-glucosidase during intracellular transport and maturation.
The synthesis and posttranslational modification of lysosomal acid alpha-glucosidase were studied in a cell-free translation system and in mammalian cells transfected with acid alpha-glucosidase cDNA constructs. The newly synthesized precursor, sequestered in the endoplasmic reticulum, was demonstrated to be membrane-bound by lack of signal peptide cleavage, and to be catalytically inactive. Sugar chain modification was shown to occur in the Golgi complex and to be dependent on the rate of transport. From the trans-Golgi network different routes were found to be followed by acid alpha-glucosidase. A fraction of precursor molecules, proteolytically released from the membrane anchor, appeared to enter the secretory pathway and was recovered from the cell culture medium in a catalytically active form. A second fraction was transported to the lysosomes and was trimmed in a stepwise process at both the amino- and carboxyl-terminal ends. The intramolecular cleavage sites were determined. Involvement of thiol proteinases was demonstrated. Specificity for the natural substrate glycogen was gained during the maturation process. The phosphomannosyl receptor is assumed to be instrumental in the lysosomal targeting of acid alpha-glucosidase, but a phosphomannosyl receptor-independent transport of membrane-bound precursor molecules to the lysosomes, either directly or via the plasma membrane, cannot be excluded.