AJP - Cell Physiology, Vol 248, Issue 1 135-C144, Copyright © 1985 by American Physiological Society
Transport and proteolytic processing of rabbit cardiac cathepsin D
A. M. Samarel, A. G. Ferguson, S. W. Worobec and M. Lesch
Rabbit cardiac cathepsin D is synthesized as a 53,000-mol wt precursor that
undergoes limited proteolysis at an unknown intracellular site to a
48,000-mol wt active form. To examine the site of proteolytic processing,
isolated perfused rabbit hearts were fractionated by differential
centrifugation 150 or 300 min after pulse labeling with [35S]methionine.
Newly synthesized precursor and processed cathepsin D were quantitatively
isolated from each fraction by extraction, immunoadsorption, and sodium
dodecyl sulfate-polyacrylamide gel electrophoresis. After 30-min pulse
perfusions, all of the 35S-labeled cathepsin D was present as precursor,
with the greatest amounts found in low-density subcellular fractions.
Proteolytic processing of cathepsin D precursor occurred after chase
perfusions that were coincident with the subcellular redistribution of
newly synthesized enzyme from sites of synthesis to heavier subcellular
structures. Pulse-chase perfusions with chloroquine (10 microM) inhibited
precursor proteolytic processing and the time-dependent subcellular
redistribution of newly synthesized cathepsin D. The data are consistent
with a model for cardiac lysosomal enzyme maturation in which limited
proteolytic processing occurs coincident with or soon after the transport
of precursors to an acidic intracellular compartment. The results thus
suggest that cathepsin D proteolytic processing occurs within cardiac
lysosomes.
