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pubmed:abstractText |
Cathepsin L, a lysosomal cysteine protease, is overexpressed and secreted by malignantly transformed cells. However, the reason for secretion of this man 6-phosphate-containing lysosomal protease into the extracellular medium is not clear. We wished to determine whether there is a region within the primary sequence of the proenzyme form of cathepsin L which affects its subcellular and extracellular localization. High-level transient expression of human procathepsin L in mouse NIH 3T3 cells results in the secretion of most of this protein into the extracellular medium. At the same time, the endogenous mouse procathepsin L in these nontransformed cells is found in its usual location in lysosomes. Mutants of human procathepsin L with carboxy-terminus deletions involving the last 11 amino acids are not secreted into the medium. Deletion of as little as two amino acids, Thr and Val, from the carboxy terminus, blocked the secretion of the protein but did not affect its enzyme activity, posttranslational processing, or subcellular distribution. Replacement of Thr-Val by two bulky amino acids Tyr-Asn allowed secretion of the procathepsin L, but the replacement of these two amino acids by nonbulky alanines prevented its secretion. Single alanine substitutions of the last six amino acids (ASYPTV) indicated that substitution by alanine of Y or T does not affect the secretion of hproCAT L, but alanine substitutions of S, P, or V completely blocked its secretion into the culture medium. We therefore conclude that the carboxy terminus of procathepsin L contains a sequence essential for its secretion.
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