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. 1992 Sep 1;89(17):8107–8111. doi: 10.1073/pnas.89.17.8107

A mitogenic peptide amide encoded within the E peptide domain of the insulin-like growth factor IB prohormone.

J M Siegfried 1, P G Kasprzyk 1, A M Treston 1, J L Mulshine 1, K A Quinn 1, F Cuttitta 1
PMCID: PMC49865  PMID: 1325646

Abstract

We have identified an amino acid sequence within the E peptide of the insulin-like growth factor IB (IGF-IB) precursor that is biologically active and designated this peptide insulin-like growth factor IB-(103-124) E1 amide (IBE1). Its existence was predicted by a flanking Gly-Lys-Lys-Lys, a signal sequence for sequential proteolytic cleavage and peptidyl C-terminal amidation. A synthetic analog of the predicted IBE1 peptide, designated Y-23-R-NH2, was generated with tyrosine added at position 0. This peptide at 2-20 nM had growth-promoting effects on both normal and malignant human bronchial epithelial cells. Y-23-R-NH2 bound to specific high-affinity receptors (Kd = 2.8 +/- 1.4 x 10(-11) M) present at 1-2 x 10(4) binding sites per cell. Ligand binding was not inhibited by recombinant insulin or recombinant IGF-I. Furthermore, a monoclonal antibody antagonist to the IGF-I receptor (alpha IR3) did not suppress the proliferative response induced by Y-23-R-NH2. In addition, C-terminal amidation was shown to be important in receptor recognition since the free-acid analog of IBE1 (Y-23-R-OH) did not effectively compete for binding and was not a potent agonist of proliferation. Immunoblot analysis of human lung tumor cell line extracts using an antibody raised against Y-23-R-NH2 detected a low molecular mass band of approximately 5 kDa, implying that a protein product is produced that has immunological similarity to IBE1. Extracts of human, mammalian, and avian livers analyzed on an immunoblot with the anti-Y-23-R-NH2 antibody contained proteins of approximately 21 kDa that were specifically recognized by the antiserum and presumably represent an IGF-I precursor molecule. This implies that in species where an IGF-I mRNA with homology to the human IGF-IB E domain has not yet been described, an alternate mRNA must be produced that contains a sequence similar to that of the midportion of the human IGF-IB E domain. Our findings demonstrate that IBE1 is a growth factor that mediates its effect through a specific high-affinity receptor and is most likely conserved in many species.

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Selected References

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  1. Barr P. J. Mammalian subtilisins: the long-sought dibasic processing endoproteases. Cell. 1991 Jul 12;66(1):1–3. doi: 10.1016/0092-8674(91)90129-m. [DOI] [PubMed] [Google Scholar]
  2. Bell G. I., Sanchez-Pescador R., Laybourn P. J., Najarian R. C. Exon duplication and divergence in the human preproglucagon gene. 1983 Jul 28-Aug 3Nature. 304(5924):368–371. doi: 10.1038/304368a0. [DOI] [PubMed] [Google Scholar]
  3. Bradbury A. F., Finnie M. D., Smyth D. G. Mechanism of C-terminal amide formation by pituitary enzymes. Nature. 1982 Aug 12;298(5875):686–688. doi: 10.1038/298686a0. [DOI] [PubMed] [Google Scholar]
  4. Clemmons D. R., Van Wyk J. J. Somatomedin-C and platelet-derived growth factor stimulate human fibroblast replication. J Cell Physiol. 1981 Mar;106(3):361–367. doi: 10.1002/jcp.1041060305. [DOI] [PubMed] [Google Scholar]
  5. Cuttitta F., Carney D. N., Mulshine J., Moody T. W., Fedorko J., Fischler A., Minna J. D. Bombesin-like peptides can function as autocrine growth factors in human small-cell lung cancer. 1985 Aug 29-Sep 4Nature. 316(6031):823–826. doi: 10.1038/316823a0. [DOI] [PubMed] [Google Scholar]
  6. Cuttitta F., Fedorko J., Gu J. A., Lebacq-Verheyden A. M., Linnoila R. I., Battey J. F. Gastrin-releasing peptide gene-associated peptides are expressed in normal human fetal lung and small cell lung cancer: a novel peptide family found in man. J Clin Endocrinol Metab. 1988 Sep;67(3):576–583. doi: 10.1210/jcem-67-3-576. [DOI] [PubMed] [Google Scholar]
  7. Daughaday W. H. Hormonal regulation of growth by somatomedin and other tissue growth factors. Clin Endocrinol Metab. 1977 Mar;6(1):117–135. doi: 10.1016/s0300-595x(77)80059-5. [DOI] [PubMed] [Google Scholar]
  8. Eipper B. A., Perkins S. N., Husten E. J., Johnson R. C., Keutmann H. T., Mains R. E. Peptidyl-alpha-hydroxyglycine alpha-amidating lyase. Purification, characterization, and expression. J Biol Chem. 1991 Apr 25;266(12):7827–7833. [PubMed] [Google Scholar]
  9. Frolik C. A., De Larco J. E. Radioreceptor assays for transforming growth factors. Methods Enzymol. 1987;146:95–102. doi: 10.1016/s0076-6879(87)46012-6. [DOI] [PubMed] [Google Scholar]
  10. Gainer H., Russell J. T., Loh Y. P. The enzymology and intracellular organization of peptide precursor processing: the secretory vesicle hypothesis. Neuroendocrinology. 1985 Feb;40(2):171–184. doi: 10.1159/000124070. [DOI] [PubMed] [Google Scholar]
  11. Han V. K., D'Ercole A. J., Lund P. K. Cellular localization of somatomedin (insulin-like growth factor) messenger RNA in the human fetus. Science. 1987 Apr 10;236(4798):193–197. doi: 10.1126/science.3563497. [DOI] [PubMed] [Google Scholar]
  12. Huff K. K., Kaufman D., Gabbay K. H., Spencer E. M., Lippman M. E., Dickson R. B. Secretion of an insulin-like growth factor-I-related protein by human breast cancer cells. Cancer Res. 1986 Sep;46(9):4613–4619. [PubMed] [Google Scholar]
  13. Jacobs S., Cook S., Svoboda M. E., Van Wyk J. J. Interaction of the monoclonal antibodies alpha IR-1 and alpha IR-3 with insulin and somatomedin-C receptors. Endocrinology. 1986 Jan;118(1):223–226. doi: 10.1210/endo-118-1-223. [DOI] [PubMed] [Google Scholar]
  14. Jansen M., van Schaik F. M., Ricker A. T., Bullock B., Woods D. E., Gabbay K. H., Nussbaum A. L., Sussenbach J. S., Van den Brande J. L. Sequence of cDNA encoding human insulin-like growth factor I precursor. Nature. 1983 Dec 8;306(5943):609–611. doi: 10.1038/306609a0. [DOI] [PubMed] [Google Scholar]
  15. Jensen R. T., Moody T., Pert C., Rivier J. E., Gardner J. D. Interaction of bombesin and litorin with specific membrane receptors on pancreatic acinar cells. Proc Natl Acad Sci U S A. 1978 Dec;75(12):6139–6143. doi: 10.1073/pnas.75.12.6139. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lopez L. C., Frazier M. L., Su C. J., Kumar A., Saunders G. F. Mammalian pancreatic preproglucagon contains three glucagon-related peptides. Proc Natl Acad Sci U S A. 1983 Sep;80(18):5485–5489. doi: 10.1073/pnas.80.18.5485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Macauly V. M., Teale J. D., Everard M. J., Joshi G. P., Smith I. E., Millar J. L. Somatomedin-C/insulin-like growth factor-I is a mitogen for human small cell lung cancer. Br J Cancer. 1988 Jan;57(1):91–93. doi: 10.1038/bjc.1988.16. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Nagaoka I., Someya A., Iwabuchi K., Yamashita T. Expression of insulin-like growth factor-IA and factor-IB mRNA in human liver, hepatoma cells, macrophage-like cells and fibroblasts. FEBS Lett. 1991 Mar 11;280(1):79–83. doi: 10.1016/0014-5793(91)80208-k. [DOI] [PubMed] [Google Scholar]
  19. Nakanishi Y., Mulshine J. L., Kasprzyk P. G., Natale R. B., Maneckjee R., Avis I., Treston A. M., Gazdar A. F., Minna J. D., Cuttitta F. Insulin-like growth factor-I can mediate autocrine proliferation of human small cell lung cancer cell lines in vitro. J Clin Invest. 1988 Jul;82(1):354–359. doi: 10.1172/JCI113594. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Orskov C., Bersani M., Johnsen A. H., Højrup P., Holst J. J. Complete sequences of glucagon-like peptide-1 from human and pig small intestine. J Biol Chem. 1989 Aug 5;264(22):12826–12829. [PubMed] [Google Scholar]
  21. Roberts C. T., Jr, Lasky S. R., Lowe W. L., Jr, Seaman W. T., LeRoith D. Molecular cloning of rat insulin-like growth factor I complementary deoxyribonucleic acids: differential messenger ribonucleic acid processing and regulation by growth hormone in extrahepatic tissues. Mol Endocrinol. 1987 Mar;1(3):243–248. doi: 10.1210/mend-1-3-243. [DOI] [PubMed] [Google Scholar]
  22. Rosenfeld M. G., Mermod J. J., Amara S. G., Swanson L. W., Sawchenko P. E., Rivier J., Vale W. W., Evans R. M. Production of a novel neuropeptide encoded by the calcitonin gene via tissue-specific RNA processing. Nature. 1983 Jul 14;304(5922):129–135. doi: 10.1038/304129a0. [DOI] [PubMed] [Google Scholar]
  23. Rotwein P., Folz R. J., Gordon J. I. Biosynthesis of human insulin-like growth factor I (IGF-I). The primary translation product of IGF-I mRNA contains an unusual 48-amino acid signal peptide. J Biol Chem. 1987 Aug 25;262(24):11807–11812. [PubMed] [Google Scholar]
  24. Rotwein P., Pollock K. M., Didier D. K., Krivi G. G. Organization and sequence of the human insulin-like growth factor I gene. Alternative RNA processing produces two insulin-like growth factor I precursor peptides. J Biol Chem. 1986 Apr 15;261(11):4828–4832. [PubMed] [Google Scholar]
  25. Rotwein P. Two insulin-like growth factor I messenger RNAs are expressed in human liver. Proc Natl Acad Sci U S A. 1986 Jan;83(1):77–81. doi: 10.1073/pnas.83.1.77. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Schoenle E., Zapf J., Humbel R. E., Froesch E. R. Insulin-like growth factor I stimulates growth in hypophysectomized rats. Nature. 1982 Mar 18;296(5854):252–253. doi: 10.1038/296252a0. [DOI] [PubMed] [Google Scholar]
  27. Siegfried J. M. Detection of human lung epithelial cell growth factors produced by a lung carcinoma cell line: use in culture of primary solid lung tumors. Cancer Res. 1987 Jun 1;47(11):2903–2910. [PubMed] [Google Scholar]
  28. Siegfried J. M., Nesnow S. Cytotoxicity of chemical carcinogens towards human bronchial epithelial cells evaluated in a clonal assay. Carcinogenesis. 1984 Oct;5(10):1317–1322. doi: 10.1093/carcin/5.10.1317. [DOI] [PubMed] [Google Scholar]
  29. Steiner D. F., Quinn P. S., Chan S. J., Marsh J., Tager H. S. Processing mechanisms in the biosynthesis of proteins. Ann N Y Acad Sci. 1980;343:1–16. doi: 10.1111/j.1749-6632.1980.tb47238.x. [DOI] [PubMed] [Google Scholar]
  30. Sussenbach J. S. The gene structure of the insulin-like growth factor family. Prog Growth Factor Res. 1989;1(1):33–48. doi: 10.1016/0955-2235(89)90040-9. [DOI] [PubMed] [Google Scholar]
  31. Tatemoto K., Mutt V. Isolation of two novel candidate hormones using a chemical method for finding naturally occurring polypeptides. Nature. 1980 Jun 5;285(5764):417–418. doi: 10.1038/285417a0. [DOI] [PubMed] [Google Scholar]
  32. Tricoli J. V., Rall L. B., Karakousis C. P., Herrera L., Petrelli N. J., Bell G. I., Shows T. B. Enhanced levels of insulin-like growth factor messenger RNA in human colon carcinomas and liposarcomas. Cancer Res. 1986 Dec;46(12 Pt 1):6169–6173. [PubMed] [Google Scholar]
  33. Vassilopoulou-Sellin R., Phillips L. S. Extraction of somatomedin activity from rat liver. Endocrinology. 1982 Feb;110(2):582–589. doi: 10.1210/endo-110-2-582. [DOI] [PubMed] [Google Scholar]
  34. Vigna S. R., Giraud A. S., Soll A. H., Walsh J. H., Mantyh P. W. Bombesin receptors on gastrin cells. Ann N Y Acad Sci. 1988;547:131–137. doi: 10.1111/j.1749-6632.1988.tb23881.x. [DOI] [PubMed] [Google Scholar]

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