A Novel Aspartyl Proteinase from Apocrine Epithelia and Breast Tumors

Emilia Caputo,Giuseppe Manco,Luigi Mandrich,John Guardiola

doi:10.1074/jbc.275.11.7935

Abstract

GCDFP-15 (gross cystic disease fluid protein, 15 kDa) is a secretory marker of apocrine differentiation in breast carcinoma. In human breast cancer cell lines, gene expression is regulated by hormones, including androgens and prolactin. The protein is also known under different names in different body fluids such as gp17 in seminal plasma. GCDFP-15/gp17 is a ligand of CD4 and is a potent inhibitor of T-cell apoptosis induced by sequential CD4/T-cell receptor triggering. We now report that GCDFP-15/gp17 is a protease exhibiting structural properties relating it to the aspartyl proteinase superfamily. Unexpectedly, GCDFP-15/gp17 appears to be related to the retroviral members rather than to the known cellular members of this class. Site-specific mutagenesis of Asp(22) (predicted to be catalytically important for the active site) and pepstatin A inhibition confirmed that the protein is an aspartic-type protease. We also show that, among the substrates tested, GCDFP-15/gp17 is specific for fibronectin. The study of GCDFP-15/gp17-mediated proteolysis may provide a handle to understand phenomena as diverse as mammary tumor progression and fertilization.

Highlights

GCDFP-15, known as prolactin-inducible protein [1], gp17 [2], secretory actin-binding protein [3], and extraparotid glycoprotein [4], is a protein secreted by various exocrine glands, including the seminal vesicle, salivary gland, and sweat glands
Studies from our laboratory on the gp17 protein purified from human seminal plasma have shown that it is a ligand for CD4 [11], in turn a T-cell co-receptor playing a key role in antigen recognition and T-cell activation
Structural Homology between GCDFP-15/gp17 and the Aspartyl Proteinases—To gain insight into the possible function(s) of GCDFP-15/gp17, we decided to made full use of the TOPITS “threading” method [21], a potent means used to search for remote homologous protein structures with sequence identity around the so-called “twilight zone” [22], for comparison of the predicted secondary structure of GCDFP-15/gp17 with those derived from proteins with known three-dimensional structure present in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University

Summary

A Novel Aspartyl Proteinase from Apocrine Epithelia and Breast Tumors*

(Received for publication, September 13, 1999, and in revised form, December 22, 1999). GCDFP-15 (gross cystic disease fluid protein, 15 kDa), known as prolactin-inducible protein [1], gp17 [2], secretory actin-binding protein [3], and extraparotid glycoprotein [4], is a protein secreted by various exocrine glands, including the seminal vesicle, salivary gland, and sweat glands. This protein is, in addition, expressed by cancer cells derived from a limited number of tissues, among which are prominent primary and secondary breast carcinomas exhibiting an apocrine differentiation [5]. The finding that a significant percentage of breast carcinomas have the ability to synthesize and secrete GCDFP15/gp17 [14], together with its absence in normal resting mammary gland, raises the possibility that this proteinase might play a role in the lytic processes associated with invasive breast cancer lesions, as already found for other proteinases, including matrix metalloproteinases [15], plasminogen activators [16], and secreted lysosomal enzymes [17]

EXPERIMENTAL PROCEDURES

A Novel Aspartic Protease from Breast Tumors

RESULTS

DISCUSSION