IDENTIFICATION AND BIOLOGICAL CHARACTERIZATION OF AN EPIDERMAL GROWTHFACTOR-RELATED PROTEIN - CRIPTO-1

The human and mouse cripto-1 (CR-1) genes can code for proteins relate d in structure to epidermal growth factor (EGF). A specific 36-kDa imm unoreactive protein was detected by Western blot analysis in human cel l lines that express CR-1 mRNA but not in cell lines that fail to expr ess this transcript. Immunoprecipitation of GEO colon carcinoma or mou se embryonal carcinoma cells detected 27-29-kDa and 24-kDa proteins, r espectively. Cell lysates and conditioned medium that were prepared fr om several CHO clones and were expressing either a recombinant human o r mouse CR-1 cDNA contained immunospecific 27-29-kDa and 24 kDa protei ns, respectively. Monensin or tunicamycin treatment resulted in a shif t of the 27-29-kDa human CR-1 protein to 24 kDa and 20 kDa, respective ly. The 20-kDa protein was also observed after digestion of the 27-29- kDa human CR-1 protein with N-glycosidase F. Using two CR-1 synthetic refolded peptides that correspond to the EGF-like domain of the human CR-1 sequence or conditioned medium obtained from human CR-1 expressin g CHO cells, growth stimulatory activity could be detected on nontrans formed human mammary epithelial cells and on two human breast cancer c ell lines. EGF receptor-blocking antibody did not inhibit the growth s timulatory action of the CR-1 protein. Likewise, the CR-1 refolded pep tides or conditioned medium from the human CR-1-expressing CHO cells f ailed to inhibit the binding of I-125-EGF in an EGF-radioreceptor assa y. These data demonstrate that the CR-1 is a glycoprotein that can fun ction as a growth factor through an EGF receptor-independent pathway.