Protein processing by the placental protease, cathepsin P.
Cathepsin P is a member of a family of placentally expressed cathepsins (PECs). The closest human homolog of cathepsin P is cathepsin L, a broad specificity enzyme that has functions in many tissues in addition to placenta. The gene duplications that gave rise to the PECs provide a rare opportunity to define proteolytic functions in placenta, a transient organ unique to mammals. Peptidyl substrate and inhibitor libraries have shown that cathepsin P has evolved an unusually restricted preference for substrates containing hydrophobic amino acids. Proteomic techniques were used to probe for substrates of this enzyme. Recombinant cathepsin P was incubated with rat choriocarcinoma (Rcho-1) cell proteins to identify substrates using two-dimensional difference gel electrophoresis. Substrate proteins were excised from gels and characterized by trypsin digestion and MALDI MS/MS. Two endoplasmic reticulum (ER) proteins, gp96 and calreticulin, emerged as potential substrates, and western blotting showed that these proteins are processed by cathepsin P from their C-terminus, removing the KDEL ER retention signal. Immunohistochemistry showed that a portion of cathepsin P co-localizes with calreticulin in Rcho-1 cells. Extracellular calreticulin induces differentiation of Rcho-1 cells, indicating a potential role of cathepsin P in processing and secretion of calreticulin during differentiation of trophoblast giant cells.
Department of Biomedical Research, Alfred I duPont Hospital for Children, Wilmington, DE 19803, USA.
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Hassanein
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Glazewski
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Mason
D000818:Animals; D015153:Blotting, Western; D037282:Calreticulin / Q000378:metabolism / Q000494:pharmacology; D002403:Cathepsins / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects; D045744:Cell Line, Tumor; D015180:Electrophoresis, Gel, Two-Dimensional; D005260:Female; D007150:Immunohistochemistry; D010920:Placenta / Q000201:enzymology*; D051381:Rats; D014327:Trophoblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism
D037282:Calreticulin; D002403:Cathepsins