Abstract
Two independent cDNA clones corresponding to a 2.7-kilobase (kb) epidermal growth factor receptor (EGF-R) mRNA were isolated from a rat liver cDNA library. Sequence analysis revealed 100% homology in the external domain when compared with the full-length rat EGF-R nucleotide sequence and 80 to 90% similarity relative to the human EGF-R. However, the 3'-terminal sequence of these clones did not match EGF-R or any other known sequence(s) and was distinct from the 3' end of the 2.8-kb mRNA, which encodes a truncated EGF-R in A431 cells. The deduced amino acid sequence revealed an open reading frame which is homologous to the external domain of the EGF-R but which terminates prior to the transmembrane region. Southern blot analysis of rat genomic DNA indicated that the 3'-terminal sequence of this transcript is derived from the EGF-R gene. Analysis of a genomic clone containing the 3' end of the 2.7-kb transcript revealed that this sequence is present as a discrete exon in the mid-region of the receptor gene in proximity to the exon encoding the transmembrane domain. Introduction of an expression vector containing the truncated EGF-R cDNA into Chinese hamster ovary (CHO) cells led to the expression of a 95-kilodalton protein which was detected in conditioned media, by using antisera directed against the EGF-R. A similarly sized protein was also detected in the media of WB cells, a continuous, nontransformed line of rat hepatic epithelial cells. Northern (RNA blot) analysis established that the truncated receptor is encoded by a 2.7-kb transcript found in normal rat liver. Furthermore, Northern analysis of rat poly(A)+ RNA showed that the 2.7-kb EGF-R transcript is expressed at differing levels in various fetal and adult tissues. These data indicate that alternative splicing of the EGF-R primary transcript yields a 2.7-kb mRNA which codes for a truncated form of the receptor. This receptor is secreted by rat hepatic epithelial cells in culture, which suggests that it may be secreted by normal rat cells or tissues and perhaps serve an as yet unknown physiological function.
Full text
PDFImages in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Basu A., Raghunath M., Bishayee S., Das M. Inhibition of tyrosine kinase activity of the epidermal growth factor (EGF) receptor by a truncated receptor form that binds to EGF: role for interreceptor interaction in kinase regulation. Mol Cell Biol. 1989 Feb;9(2):671–677. doi: 10.1128/mcb.9.2.671. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Carpenter G., Cohen S. Epidermal growth factor. Annu Rev Biochem. 1979;48:193–216. doi: 10.1146/annurev.bi.48.070179.001205. [DOI] [PubMed] [Google Scholar]
- Carpenter G. Receptors for epidermal growth factor and other polypeptide mitogens. Annu Rev Biochem. 1987;56:881–914. doi: 10.1146/annurev.bi.56.070187.004313. [DOI] [PubMed] [Google Scholar]
- Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
- Davis J. A., Linzer D. I. Expression of multiple forms of the prolactin receptor in mouse liver. Mol Endocrinol. 1989 Apr;3(4):674–680. doi: 10.1210/mend-3-4-674. [DOI] [PubMed] [Google Scholar]
- Downward J., Yarden Y., Mayes E., Scrace G., Totty N., Stockwell P., Ullrich A., Schlessinger J., Waterfield M. D. Close similarity of epidermal growth factor receptor and v-erb-B oncogene protein sequences. Nature. 1984 Feb 9;307(5951):521–527. doi: 10.1038/307521a0. [DOI] [PubMed] [Google Scholar]
- Earp H. S., Austin K. S., Blaisdell J., Rubin R. A., Nelson K. G., Lee L. W., Grisham J. W. Epidermal growth factor (EGF) stimulates EGF receptor synthesis. J Biol Chem. 1986 Apr 15;261(11):4777–4780. [PubMed] [Google Scholar]
- Fabricant R. N., De Larco J. E., Todaro G. J. Nerve growth factor receptors on human melanoma cells in culture. Proc Natl Acad Sci U S A. 1977 Feb;74(2):565–569. doi: 10.1073/pnas.74.2.565. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Filmus J., Pollak M. N., Cairncross J. G., Buick R. N. Amplified, overexpressed and rearranged epidermal growth factor receptor gene in a human astrocytoma cell line. Biochem Biophys Res Commun. 1985 Aug 30;131(1):207–215. doi: 10.1016/0006-291x(85)91790-5. [DOI] [PubMed] [Google Scholar]
- Fung Y. K., Lewis W. G., Crittenden L. B., Kung H. J. Activation of the cellular oncogene c-erbB by LTR insertion: molecular basis for induction of erythroblastosis by avian leukosis virus. Cell. 1983 Jun;33(2):357–368. doi: 10.1016/0092-8674(83)90417-8. [DOI] [PubMed] [Google Scholar]
- Glisin V., Crkvenjakov R., Byus C. Ribonucleic acid isolated by cesium chloride centrifugation. Biochemistry. 1974 Jun 4;13(12):2633–2637. doi: 10.1021/bi00709a025. [DOI] [PubMed] [Google Scholar]
- Graf T., Beug H. Role of the v-erbA and v-erbB oncogenes of avian erythroblastosis virus in erythroid cell transformation. Cell. 1983 Aug;34(1):7–9. doi: 10.1016/0092-8674(83)90130-7. [DOI] [PubMed] [Google Scholar]
- Graham F. L., van der Eb A. J. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology. 1973 Apr;52(2):456–467. doi: 10.1016/0042-6822(73)90341-3. [DOI] [PubMed] [Google Scholar]
- Gullick W. J., Marsden J. J., Whittle N., Ward B., Bobrow L., Waterfield M. D. Expression of epidermal growth factor receptors on human cervical, ovarian, and vulval carcinomas. Cancer Res. 1986 Jan;46(1):285–292. [PubMed] [Google Scholar]
- Hayman M. J., Ramsay G. M., Savin K., Kitchener G., Graf T., Beug H. Identification and characterization of the avian erythroblastosis virus erbB gene product as a membrane glycoprotein. Cell. 1983 Feb;32(2):579–588. doi: 10.1016/0092-8674(83)90477-4. [DOI] [PubMed] [Google Scholar]
- Hendler F. J., Ozanne B. W. Human squamous cell lung cancers express increased epidermal growth factor receptors. J Clin Invest. 1984 Aug;74(2):647–651. doi: 10.1172/JCI111463. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hunts J., Ueda M., Ozawa S., Abe O., Pastan I., Shimizu N. Hyperproduction and gene amplification of the epidermal growth factor receptor in squamous cell carcinomas. Jpn J Cancer Res. 1985 Aug;76(8):663–666. [PubMed] [Google Scholar]
- Izumo S., Mahdavi V. Thyroid hormone receptor alpha isoforms generated by alternative splicing differentially activate myosin HC gene transcription. Nature. 1988 Aug 11;334(6182):539–542. doi: 10.1038/334539a0. [DOI] [PubMed] [Google Scholar]
- Johnson A. C., Garfield S. H., Merlino G. T., Pastan I. Expression of epidermal growth factor receptor proto-oncogene mRNA in regenerating rat liver. Biochem Biophys Res Commun. 1988 Jan 15;150(1):412–418. doi: 10.1016/0006-291x(88)90536-0. [DOI] [PubMed] [Google Scholar]
- Koenig R. J., Lazar M. A., Hodin R. A., Brent G. A., Larsen P. R., Chin W. W., Moore D. D. Inhibition of thyroid hormone action by a non-hormone binding c-erbA protein generated by alternative mRNA splicing. Nature. 1989 Feb 16;337(6208):659–661. doi: 10.1038/337659a0. [DOI] [PubMed] [Google Scholar]
- Kris R. M., Lax I., Gullick W., Waterfield M. D., Ullrich A., Fridkin M., Schlessinger J. Antibodies against a synthetic peptide as a probe for the kinase activity of the avian EGF receptor and v-erbB protein. Cell. 1985 Mar;40(3):619–625. doi: 10.1016/0092-8674(85)90210-7. [DOI] [PubMed] [Google Scholar]
- Lax I., Kris R., Sasson I., Ullrich A., Hayman M. J., Beug H., Schlessinger J. Activation of c-erbB in avian leukosis virus-induced erythroblastosis leads to the expression of a truncated EGF receptor kinase. EMBO J. 1985 Dec 1;4(12):3179–3182. doi: 10.1002/j.1460-2075.1985.tb04062.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lee D. C., Rose T. M., Webb N. R., Todaro G. J. Cloning and sequence analysis of a cDNA for rat transforming growth factor-alpha. Nature. 1985 Feb 7;313(6002):489–491. doi: 10.1038/313489a0. [DOI] [PubMed] [Google Scholar]
- Leung D. W., Spencer S. A., Cachianes G., Hammonds R. G., Collins C., Henzel W. J., Barnard R., Waters M. J., Wood W. I. Growth hormone receptor and serum binding protein: purification, cloning and expression. Nature. 1987 Dec 10;330(6148):537–543. doi: 10.1038/330537a0. [DOI] [PubMed] [Google Scholar]
- Libermann T. A., Nusbaum H. R., Razon N., Kris R., Lax I., Soreq H., Whittle N., Waterfield M. D., Ullrich A., Schlessinger J. Amplification, enhanced expression and possible rearrangement of EGF receptor gene in primary human brain tumours of glial origin. Nature. 1985 Jan 10;313(5998):144–147. doi: 10.1038/313144a0. [DOI] [PubMed] [Google Scholar]
- Lin C. R., Chen W. S., Kruiger W., Stolarsky L. S., Weber W., Evans R. M., Verma I. M., Gill G. N., Rosenfeld M. G. Expression cloning of human EGF receptor complementary DNA: gene amplification and three related messenger RNA products in A431 cells. Science. 1984 May 25;224(4651):843–848. doi: 10.1126/science.6326261. [DOI] [PubMed] [Google Scholar]
- Maihle N. J., Kung H. J. C-erbB and the epidermal growth-factor receptor: a molecule with dual identity. Biochim Biophys Acta. 1989 Feb;948(3):287–304. doi: 10.1016/0304-419x(89)90003-6. [DOI] [PubMed] [Google Scholar]
- Matsui T., Heidaran M., Miki T., Popescu N., La Rochelle W., Kraus M., Pierce J., Aaronson S. Isolation of a novel receptor cDNA establishes the existence of two PDGF receptor genes. Science. 1989 Feb 10;243(4892):800–804. doi: 10.1126/science.2536956. [DOI] [PubMed] [Google Scholar]
- Merlino G. T., Ishii S., Whang-Peng J., Knutsen T., Xu Y. H., Clark A. J., Stratton R. H., Wilson R. K., Ma D. P., Roe B. A. Structure and localization of genes encoding aberrant and normal epidermal growth factor receptor RNAs from A431 human carcinoma cells. Mol Cell Biol. 1985 Jul;5(7):1722–1734. doi: 10.1128/MCB.5.7.1722. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Merlino G. T., Xu Y. H., Ishii S., Clark A. J., Semba K., Toyoshima K., Yamamoto T., Pastan I. Amplification and enhanced expression of the epidermal growth factor receptor gene in A431 human carcinoma cells. Science. 1984 Apr 27;224(4647):417–419. doi: 10.1126/science.6200934. [DOI] [PubMed] [Google Scholar]
- Miles B. D., Robinson H. L. High-frequency transduction of c-erbB in avian leukosis virus-induced erythroblastosis. J Virol. 1985 May;54(2):295–303. doi: 10.1128/jvi.54.2.295-303.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nieto-Sampedro M. Astrocyte mitogen inhibitor related to epidermal growth factor receptor. Science. 1988 Jun 24;240(4860):1784–1786. doi: 10.1126/science.3289118. [DOI] [PubMed] [Google Scholar]
- Ozanne B., Richards C. S., Hendler F., Burns D., Gusterson B. Over-expression of the EGF receptor is a hallmark of squamous cell carcinomas. J Pathol. 1986 May;149(1):9–14. doi: 10.1002/path.1711490104. [DOI] [PubMed] [Google Scholar]
- Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Seino S., Bell G. I. Alternative splicing of human insulin receptor messenger RNA. Biochem Biophys Res Commun. 1989 Feb 28;159(1):312–316. doi: 10.1016/0006-291x(89)92439-x. [DOI] [PubMed] [Google Scholar]
- Seino S., Seino M., Nishi S., Bell G. I. Structure of the human insulin receptor gene and characterization of its promoter. Proc Natl Acad Sci U S A. 1989 Jan;86(1):114–118. doi: 10.1073/pnas.86.1.114. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shimizu N., Kondo I., Gamou S., Behzadian M. A., Shimizu Y. Genetic analysis of hyperproduction of epidermal growth factor receptors in human epidermoid carcinoma A431 cells. Somat Cell Mol Genet. 1984 Jan;10(1):45–53. doi: 10.1007/BF01534472. [DOI] [PubMed] [Google Scholar]
- Smith W. C., Linzer D. I., Talamantes F. Detection of two growth hormone receptor mRNAs and primary translation products in the mouse. Proc Natl Acad Sci U S A. 1988 Dec;85(24):9576–9579. doi: 10.1073/pnas.85.24.9576. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
- Steffens D. L., Gross R. W. Sequencing of cloned DNA using bacteriophage lambda gt11 templates. Biotechniques. 1989 Jul-Aug;7(7):674–680. [PubMed] [Google Scholar]
- Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ullrich A., Coussens L., Hayflick J. S., Dull T. J., Gray A., Tam A. W., Lee J., Yarden Y., Libermann T. A., Schlessinger J. Human epidermal growth factor receptor cDNA sequence and aberrant expression of the amplified gene in A431 epidermoid carcinoma cells. 1984 May 31-Jun 6Nature. 309(5967):418–425. doi: 10.1038/309418a0. [DOI] [PubMed] [Google Scholar]
- Vu T. H., Martin G. R., Lee P., Mark D., Wang A., Williams L. T. Developmentally regulated use of alternative promoters creates a novel platelet-derived growth factor receptor transcript in mouse teratocarcinoma and embryonic stem cells. Mol Cell Biol. 1989 Oct;9(10):4563–4567. doi: 10.1128/mcb.9.10.4563. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weber W., Gill G. N., Spiess J. Production of an epidermal growth factor receptor-related protein. Science. 1984 Apr 20;224(4646):294–297. doi: 10.1126/science.6324343. [DOI] [PubMed] [Google Scholar]
- Wigler M., Silverstein S., Lee L. S., Pellicer A., Cheng Y. c., Axel R. Transfer of purified herpes virus thymidine kinase gene to cultured mouse cells. Cell. 1977 May;11(1):223–232. doi: 10.1016/0092-8674(77)90333-6. [DOI] [PubMed] [Google Scholar]
- Xu Y. H., Ishii S., Clark A. J., Sullivan M., Wilson R. K., Ma D. P., Roe B. A., Merlino G. T., Pastan I. Human epidermal growth factor receptor cDNA is homologous to a variety of RNAs overproduced in A431 carcinoma cells. 1984 Jun 28-Jul 4Nature. 309(5971):806–810. doi: 10.1038/309806a0. [DOI] [PubMed] [Google Scholar]
- Yamamoto T., Kamata N., Kawano H., Shimizu S., Kuroki T., Toyoshima K., Rikimaru K., Nomura N., Ishizaki R., Pastan I. High incidence of amplification of the epidermal growth factor receptor gene in human squamous carcinoma cell lines. Cancer Res. 1986 Jan;46(1):414–416. [PubMed] [Google Scholar]
- Yamamoto T., Nishida T., Miyajima N., Kawai S., Ooi T., Toyoshima K. The erbB gene of avian erythroblastosis virus is a member of the src gene family. Cell. 1983 Nov;35(1):71–78. doi: 10.1016/0092-8674(83)90209-x. [DOI] [PubMed] [Google Scholar]
- Yarden Y., Ullrich A. Growth factor receptor tyrosine kinases. Annu Rev Biochem. 1988;57:443–478. doi: 10.1146/annurev.bi.57.070188.002303. [DOI] [PubMed] [Google Scholar]