Secretion of intact proteins and peptide fragments by lysosomal pathways of protein degradation.
We report that degradation of proteins microinjected into human fibroblasts is accompanied by release into the culture medium of peptide fragments and intact proteins as well as single amino acids. For the nine proteins and polypeptides microinjected, acid-precipitable radioactivity, i.e. peptide fragments and/or intact proteins, ranged from 10 to 67% of the total released radioactivity. Peptide fragments and/or intact protein accounted for 60% of the radioactivity released into the medium by cells microinjected with ribonuclease A. Two major radiolabeled peptide fragments were found, and one was of an appropriate size to function as an antigen in antigen-presenting cells. The peptides released from microinjected ribonuclease A were derived from lysosomal pathways of proteolysis based on several lines of evidence. Previous studies have shown that microinjected ribonuclease A is degraded to single amino acids entirely within lysosomes (McElligott, M. A., Miao, P., and Dice, J. F. (1985) J. Biol. Chem. 260, 11986-11993). We show that release of free amino acids and peptide fragments and/or intact protein was equivalently stimulated by serum deprivation and equivalently inhibited by NH4Cl. We also show that lysosomal degradation of endocytosed [3H]ribonuclease A was accompanied by the release of two peptide fragments similar in size and charge to those from microinjected [3H]ribonuclease A. These findings demonstrate that degradation within lysosomes occurs in a manner that spares specific peptides; they also suggest a previously unsuspected pathway by which cells can secrete cytosol-derived polypeptides.
Isenman
L D
LD
Department of Physiology, Tufts University School of Medicine, Boston, Massachusetts 02111.
Dice
J F
JF
D002460:Cell Line; D004705:Endocytosis; D006801:Humans; D007457:Iodine Radioisotopes; D007525:Isoelectric Focusing; D007700:Kinetics; D008247:Lysosomes / Q000201:enzymology*; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000378:metabolism*; D010455:Peptides / Q000302:isolation & purification / Q000378:metabolism*; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D011865:Radioisotope Dilution Technique; D014316:Tritium
D007457:Iodine Radioisotopes; D010455:Peptides; D011506:Proteins; D014316:Tritium; D010447:Peptide Hydrolases