Abstract
We previously identified, on normal or tumour cells, two membrane proteinases, p57 and p65, that cleave human C3, the third component of complement, thus regulating C3's biological properties. Whereas p57 was purified from human erythrocytes, p65 was identified using polyclonal anti-p57 antibodies on a human melanoma cell line resistant to complement lysis. Analysis of cell distribution of C3-cleaving proteinases established that DSm, a murine melanoma cell line, expressed a C3-cleaving proteinase distinct from p57 and p65 proteinases. Thus we purified the C3-cleaving proteinase solubilized from membranes of DSm cells. The purified proteinase, termed 'p39' on the basis of its molecular mass of 39 kDa, was identified, using specific proteinase inhibitors, as a cysteine proteinase. Anti-p39 antibodies, prepared against highly purified p39, localized the p39 C3-cleaving proteinase mainly at the cell surface and demonstrated that p39 is also secreted. Anti-p39 antibodies inhibited solubilized C3-cleaving activity. Preincubation of DSm cells with anti-p39 F(ab')2 fragments increased up to 60% complement cell susceptibility. Amino acid analysis of N-terminal and three other regions of p39 demonstrated that this C3-cleaving proteinase carries 100% identity within four regions of procathepsin L. This is the first demonstration that a melanoma cell line expresses on its surface and secretes a p39 C3-cleaving cysteine proteinase that shares sequence identities with procathepsin L. Thus the p39 cysteine proteinase represents a new member of the C3-cleaving proteinase family associated with, and/or expressed on, the cell surface.