MOLECULAR-CLONING AND EXPRESSION OF COLLAGENASE-3, A NOVEL HUMAN MATRIX METALLOPROTEINASE PRODUCED BY BREAST CARCINOMAS

A cDNA coding for a new human matrix metalloproteinase (MMP) has been cloned from a cDNA library derived from a breast tumor. The isolated c DNA contains an open reading frame coding for a polypeptide of 471 ami no acids. The predicted protein sequence displays extensive similarity to the previously known MMPs and presents all the structural features characteristic of the members of this protein family, including the w ell conserved PRCGXPD motif, involved in the latency of the enzyme and the zinc-binding domain (HEXGHXXXXXHS). In addition, this novel human MMP contains in its amino acid sequence several residues specific to the collagenase subfamily (Tyr-214, Asp-235, and Gly-237) and lacks th e 9-residue insertion present in the stromelysins. According to these structural characteristics, the MMP described herein has been tentativ ely called collagenase-3, since it represents the third member of this subfamily, composed at present of fibroblast and neutrophil collagena ses. The collagenase-3 cDNA was expressed in a vaccinia virus system, and the recombinant protein was able to degrade fibrillar collagens, p roviding support to the hypothesis that the isolated cDNA codes for an authentic collagenase. Northern blot analysis of RNA from normal and pathological tissues demonstrated the existence in breast tumors of th ree different mRNA species, which seem to be the result of the utiliza tion of different polyadenylation sites present in the 3'-noncoding re gion of the gene. By contrast, no collagenase-3 mRNA was detected eith er by Northern blot or RNA polymerase chain reaction analysis with RNA from other human tissues, including normal breast, mammary fibroadeno mas, liver, placenta, ovary, uterus, prostate, and parotid gland. On t he basis of the increased expression of collagenase-3 in breast carcin omas and the absence of detectable expression in normal tissues, a pos sible role for this metalloproteinase in the tumoral process is propos ed.