Cloning and functional characterization of a new phosphatidyl-inositol anchored molecule of a metastasizing rat pancreatic tumor

Abstract

We have described recently a panel of metastasisassociated antigens expressed on a rat pancreatic tumor. One of these molecules, recognized by the monoclonal antibody C4.4 and named accordingly C4.4A, was under physiological conditions expressed only in the gravid uterus and on epithelial of the upper gastrointestinal tract. The cDNA of the antigen has been isolated and cloned. The 1,637 b cDNA codes for a 352 amino acid long glycosylphosphatidyl-inositol (GP) anchored molecule, whose molecular weight varies in di erent cells between 94 ± 98 kD according to the degree of N-and Oglycosylation. Data base searches have revealed a low degree of homology to the receptor for the plasminogen activator (uPAR). After intrafootpad and intravenous application of C4.4A transfected and mocktransfected tumor cells, an increased number of lung nodules was detected with the former, whereby the individual metastatic nodules amalgamated without any encapsulation of the tumor tissue. Furthermore, C4.4A is involved in adhesion to laminin and, although transfection of a non-metastasizing tumor line with the molecule was not su cient, constitutively C4.4A-positive tumor cells penetrated through matrigel. This process could be completely prevented by C4.4. Finally, we could demonstrate that uPA, albeit weakly, bound to the C4.4A molecule. In view of the observed in¯uence of C4.4A on metastasis formation and matrix penetration it is tempting to speculate that this newly described metastasis-associated molecule may exert functional activity similar to the uPAR, i.e. via activation of matrix degrading enzymes. By the very restricted expression of the molecule in the adult organism, modulation of C4.4A could well be of therapeutic interest.

Ly6 family member C4.4A binds laminins 1 and 5, associates with galectin‐3 and supports cell migration

“…18 C4.4A shows 46.9% homology to uPAR. 18 Both proteins belong to the Ly-6 family, characterized by repetitive cysteine residues which allow typical ''3-finger'' folding via disulfide bridges. 19 While uPAR has 3 of these cysteines containing domains, C4.4A has only 2 and the third domain is devoid of cysteines.…”

“…17 The GPI-anchored, glycosylated C4.4A protein was first identified in a highly metastasizing rat pancreatic adenocarcinoma line. 18 C4.4A shows 46.9% homology to uPAR. 18 Both proteins belong to the Ly-6 family, characterized by repetitive cysteine residues which allow typical ''3-finger'' folding via disulfide bridges.…”

“…19 While uPAR has 3 of these cysteines containing domains, C4.4A has only 2 and the third domain is devoid of cysteines. 18 C4.4A expression in nontransformed tissue is restricted mostly to the basal epithelial layers. 20 C4.4A expression is upregulated in different types of cancer, including malignant melanoma, 21 breast, 22 lung 21 and urothelial 23 cancer as well as renal cell carcinoma.…”

“…24 While the functions of uPAR in cell migration, as a proteinase receptor and as a signaling molecule, are well characterized, 25 less is known about C4.4A; and no ligand has been identified so far, though C4.4A has been suggested to be involved in LN adhesion. 18 Stable transfection of C4.4A in the low metastasizing pancreatic adenocarcinoma line BSp73AS (BSp73AS-1B1) influenced tumor growth inasmuch as BSp73AS-1B1 tumors showed no encapsulation, whereas BSp73AS tumors were wellencapsulated. 18 We interpreted this finding as a hint that C4.4A might be involved in the organization and/or degradation of the extracellular matrix.…”

“…18 Stable transfection of C4.4A in the low metastasizing pancreatic adenocarcinoma line BSp73AS (BSp73AS-1B1) influenced tumor growth inasmuch as BSp73AS-1B1 tumors showed no encapsulation, whereas BSp73AS tumors were wellencapsulated. 18 We interpreted this finding as a hint that C4.4A might be involved in the organization and/or degradation of the extracellular matrix. Further evidence for C4.4A-matrix interactions is the observation that C4.4A is upregulated in human urothelial cells when grown on Matrigel.…”