The amino‐terminal fragment of human urokinase directs a recombinant chimeric toxin to target cells: internalization is toxin mediated

In contrast to two‐chain urokinase (uPA), a chemical conjugate between uPA and native saporin (a cytotoxic plant seed ribosome‐inactivating protein) did not require plasminogen activator inhibitors to be internalized. To dissect this pathway, we constructed a chimera consisting of the amino‐terminal fragment (ATF) of human urokinase fused to a saporin isoform (SAP‐3). The chimeric ATF‐SAP toxin was expressed in Escherichia coli, purified, and characterized for its ribosome‐inactivating activity. Besides being a potent inhibitor of protein synthesis in cell‐free assays, ATF‐SAP was specifically cytotoxic toward cells expressing human uPAR. Competition experiments indicated that both the human uPAR and the LDL‐related receptor protein are involved in mediating the cell killing ability of ATF‐SAP. We conclude that neither plasminogen activator inhibitors nor the catalytic moiety of urokinase are necessary to initiate these internalization pathways. Thus, saporin may play a role similar to plasminogen activator inhibitors in its ability to trigger internalization of uPAR‐bound ligands through endocytic receptors.—Fabbrini, M. S., Carpani, D., Bello‐Rivero, I., Soria, M. R. The amino‐terminal fragment of human urokinase directs a recombinant chimeric toxin to target cells: internalization is toxin‐mediated. FASEB J. 11, 1169–1176 (1997)