Identification and biological characterization of an epidermal growth factor-related protein: Cripto-1

Ralf Brandt, Nicola Normanno, William J. Gullick, Jiing Huey Lin, Richard Harkins, Douglas Schneider, Brenda Wallace Jones, Fortunato Ciardiello, Maria G. Persico, Franca Armenante, Nancy Kim, David S. Salomon

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Abstract

The human and mouse cripto-1 (CR-1) genes can code for proteins related in structure to epidermal growth factor (EGF). A specific 36-kDa immunoreactive protein was detected by Western blot analysis in human cell lines that express CR-1 mRNA but not in cell lines that fail to express this transcript. Immunoprecipitation of GEO colon carcinoma or mouse embryonal carcinoma cells detected 27-29-kDa and 24-kDa proteins, respectively. Cell lysates and conditioned medium that were prepared from several CHO clones and were expressing either a recombinant human or mouse CR-1 cDNA contained immunospecific 27-29-kDa and 24-kDa proteins, respectively. Monensin or tunicamycin treatment resulted in a shift of the 27-29-kDa human CR-1 protein to 24 kDa and 20 kDa, respectively. 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 expressing CHO cells, growth stimulatory activity could be detected on non-transformed human mammary epithelial cells and on two human breast cancer cell lines. EGF receptor-blocking antibody did not inhibit the growth stimulatory action of the CR-1 protein. Likewise, the CR-1 refolded peptides or conditioned medium from the human CR-1-expressing CHO cells failed to inhibit the binding of 125I-EGF in an EGF-radioreceptor assay. These data demonstrate that the CR-1 is a glycoprotein that can function as a growth factor through an EGF receptor-independent pathway.

Original languageEnglish
Pages (from-to)17320-17328
Number of pages9
JournalJournal of Biological Chemistry
Volume269
Issue number25
Publication statusPublished - Jun 24 1994

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ASJC Scopus subject areas

  • Biochemistry

Cite this

Brandt, R., Normanno, N., Gullick, W. J., Lin, J. H., Harkins, R., Schneider, D., Jones, B. W., Ciardiello, F., Persico, M. G., Armenante, F., Kim, N., & Salomon, D. S. (1994). Identification and biological characterization of an epidermal growth factor-related protein: Cripto-1. Journal of Biological Chemistry, 269(25), 17320-17328.