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. 1987 Dec 20;6(13):3995–4002. doi: 10.1002/j.1460-2075.1987.tb02742.x

Molecular cloning and expression of cDNA encoding a murine myeloid leukaemia inhibitory factor (LIF).

D P Gearing 1, N M Gough 1, J A King 1, D J Hilton 1, N A Nicola 1, R J Simpson 1, E C Nice 1, A Kelso 1, D Metcalf 1
PMCID: PMC553879  PMID: 3127201

Abstract

Leukaemia inhibitory factor (LIF) can induce macrophage differentiation in M1 murine myeloid leukaemic cells and suppress their proliferation in vitro. It does not stimulate the proliferation of normal progenitor cells and is apparently distinct from known colony-stimulating factors. We have used oligo-nucleotides complementary to partial amino acid sequence of LIF to isolate a LIF clone from a T lymphocyte cDNA library. When this cDNA was coupled to a yeast expression vector (YEpsec1) and introduced into yeast cells, a molecule with the biological properties characteristic of native LIF was secreted into the growth medium. The amino acid sequence of LIF established it to be a unique molecular entity, distinct from the other known haemopoietic growth factors. Since LIF is encoded by a unique gene, two biochemically separable forms of LIF probably represent post-transcriptional or posttranslational variants of the same gene product. In contrast to several other haemopoietic regulators, the 0.8- to 1-kb LIF mRNA was expressed constitutively in two murine T lymphocyte cell lines examined, and its abundance was not enhanced by stimulation with concanavalin A. Cloning, sequencing and expressing LIF has resolved several discrepancies in the literature concerning the identity of factors capable of inducing differentiation of murine myeloid leukaemic cells in vitro.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Abraham J. A., Mergia A., Whang J. L., Tumolo A., Friedman J., Hjerrild K. A., Gospodarowicz D., Fiddes J. C. Nucleotide sequence of a bovine clone encoding the angiogenic protein, basic fibroblast growth factor. Science. 1986 Aug 1;233(4763):545–548. doi: 10.1126/science.2425435. [DOI] [PubMed] [Google Scholar]
  2. Auron P. E., Webb A. C., Rosenwasser L. J., Mucci S. F., Rich A., Wolff S. M., Dinarello C. A. Nucleotide sequence of human monocyte interleukin 1 precursor cDNA. Proc Natl Acad Sci U S A. 1984 Dec;81(24):7907–7911. doi: 10.1073/pnas.81.24.7907. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Baldari C., Murray J. A., Ghiara P., Cesareni G., Galeotti C. L. A novel leader peptide which allows efficient secretion of a fragment of human interleukin 1 beta in Saccharomyces cerevisiae. EMBO J. 1987 Jan;6(1):229–234. doi: 10.1002/j.1460-2075.1987.tb04743.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Burgess A. W., Metcalf D., Sparrow L. G., Simpson R. J., Nice E. C. Granulocyte/macrophage colony-stimulating factor from mouse lung conditioned medium. Purification of multiple forms and radioiodination. Biochem J. 1986 May 1;235(3):805–814. doi: 10.1042/bj2350805. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Capaldi R. A., Vanderkooi G. The low polarity of many membrane proteins. Proc Natl Acad Sci U S A. 1972 Apr;69(4):930–932. doi: 10.1073/pnas.69.4.930. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chen E. Y., Seeburg P. H. Supercoil sequencing: a fast and simple method for sequencing plasmid DNA. DNA. 1985 Apr;4(2):165–170. doi: 10.1089/dna.1985.4.165. [DOI] [PubMed] [Google Scholar]
  7. Dente L., Cesareni G., Cortese R. pEMBL: a new family of single stranded plasmids. Nucleic Acids Res. 1983 Mar 25;11(6):1645–1655. doi: 10.1093/nar/11.6.1645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fung M. C., Hapel A. J., Ymer S., Cohen D. R., Johnson R. M., Campbell H. D., Young I. G. Molecular cloning of cDNA for murine interleukin-3. Nature. 1984 Jan 19;307(5948):233–237. doi: 10.1038/307233a0. [DOI] [PubMed] [Google Scholar]
  9. Gough N. M. Core and E antigen synthesis in rodent cells transformed with hepatitis B virus DNA is associated with greater than genome length viral messenger RNAs. J Mol Biol. 1983 Apr 25;165(4):683–699. doi: 10.1016/s0022-2836(83)80274-5. [DOI] [PubMed] [Google Scholar]
  10. Gough N. M., Gough J., Metcalf D., Kelso A., Grail D., Nicola N. A., Burgess A. W., Dunn A. R. Molecular cloning of cDNA encoding a murine haematopoietic growth regulator, granulocyte-macrophage colony stimulating factor. 1984 Jun 28-Jul 4Nature. 309(5971):763–767. doi: 10.1038/309763a0. [DOI] [PubMed] [Google Scholar]
  11. Gough N. M., Metcalf D., Gough J., Grail D., Dunn A. R. Structure and expression of the mRNA for murine granulocyte-macrophage colony stimulating factor. EMBO J. 1985 Mar;4(3):645–653. doi: 10.1002/j.1460-2075.1985.tb03678.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Grantham R., Gautier C., Gouy M., Jacobzone M., Mercier R. Codon catalog usage is a genome strategy modulated for gene expressivity. Nucleic Acids Res. 1981 Jan 10;9(1):r43–r74. doi: 10.1093/nar/9.1.213-b. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jaye M., Howk R., Burgess W., Ricca G. A., Chiu I. M., Ravera M. W., O'Brien S. J., Modi W. S., Maciag T., Drohan W. N. Human endothelial cell growth factor: cloning, nucleotide sequence, and chromosome localization. Science. 1986 Aug 1;233(4763):541–545. doi: 10.1126/science.3523756. [DOI] [PubMed] [Google Scholar]
  14. Jeffery W. R., Brawerman G. Characterization of the steady-state population of messenger RNA and its poly(adenylic acid) segment in mammalian cells. Biochemistry. 1974 Oct 22;13(22):4633–4637. doi: 10.1021/bi00719a026. [DOI] [PubMed] [Google Scholar]
  15. Kelso A., Metcalf D., Gough N. M. Independent regulation of granulocyte-macrophage colony-stimulating factor and multi-lineage colony-stimulating factor production in T lymphocyte clones. J Immunol. 1986 Mar 1;136(5):1718–1725. [PubMed] [Google Scholar]
  16. Klebe R. J., Harriss J. V., Sharp Z. D., Douglas M. G. A general method for polyethylene-glycol-induced genetic transformation of bacteria and yeast. Gene. 1983 Nov;25(2-3):333–341. doi: 10.1016/0378-1119(83)90238-x. [DOI] [PubMed] [Google Scholar]
  17. Laird C. D. Chromatid structure: relationship between DNA content and nucleotide sequence diversity. Chromosoma. 1971 Mar 16;32(4):378–406. doi: 10.1007/BF00285251. [DOI] [PubMed] [Google Scholar]
  18. Lipton J. H., Sachs L. Characterization of macrophage- and granulocyte-inducing proteins for normal and leukemic myeloid cells produced by the Krebs ascites tumor. Biochim Biophys Acta. 1981 Apr 3;673(4):552–569. doi: 10.1016/0304-4165(81)90486-4. [DOI] [PubMed] [Google Scholar]
  19. Metcalf D. The Wellcome Foundation lecture, 1986. The molecular control of normal and leukaemic granulocytes and macrophages. Proc R Soc Lond B Biol Sci. 1987 May 22;230(1261):389–423. doi: 10.1098/rspb.1987.0026. [DOI] [PubMed] [Google Scholar]
  20. Nagata S., Tsuchiya M., Asano S., Kaziro Y., Yamazaki T., Yamamoto O., Hirata Y., Kubota N., Oheda M., Nomura H. Molecular cloning and expression of cDNA for human granulocyte colony-stimulating factor. 1986 Jan 30-Feb 5Nature. 319(6052):415–418. doi: 10.1038/319415a0. [DOI] [PubMed] [Google Scholar]
  21. Nicola N. A., Metcalf D. Binding of the differentiation-inducer, granulocyte-colony-stimulating factor, to responsive but not unresponsive leukemic cell lines. Proc Natl Acad Sci U S A. 1984 Jun;81(12):3765–3769. doi: 10.1073/pnas.81.12.3765. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Nicola N. A., Metcalf D., Matsumoto M., Johnson G. R. Purification of a factor inducing differentiation in murine myelomonocytic leukemia cells. Identification as granulocyte colony-stimulating factor. J Biol Chem. 1983 Jul 25;258(14):9017–9023. [PubMed] [Google Scholar]
  23. Parnes J. R., Seidman J. G. Structure of wild-type and mutant mouse beta 2-microglobulin genes. Cell. 1982 Jun;29(2):661–669. doi: 10.1016/0092-8674(82)90182-9. [DOI] [PubMed] [Google Scholar]
  24. SWARTZ M. N., TRAUTNER T. A., KORNBERG A. Enzymatic synthesis of deoxyribonucleic acid. XI. Further studies on nearest neighbor base sequences in deoxyribonucleic acids. J Biol Chem. 1962 Jun;237:1961–1967. [PubMed] [Google Scholar]
  25. Sachs L. Control of growth and normal differentiation in leukemic cells: regulation of the developmental program and restoration of the normal phenotype in myeloid leukemia. J Cell Physiol Suppl. 1982;1:151–164. doi: 10.1002/jcp.1041130423. [DOI] [PubMed] [Google Scholar]
  26. 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]
  27. Stanley E., Metcalf D., Sobieszczuk P., Gough N. M., Dunn A. R. The structure and expression of the murine gene encoding granulocyte-macrophage colony stimulating factor: evidence for utilisation of alternative promoters. EMBO J. 1985 Oct;4(10):2569–2573. doi: 10.1002/j.1460-2075.1985.tb03972.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Takahashi N., Takahashi Y., Putnam F. W. Structure of human hemopexin: O-glycosyl and N-glycosyl sites and unusual clustering of tryptophan residues. Proc Natl Acad Sci U S A. 1984 Apr;81(7):2021–2025. doi: 10.1073/pnas.81.7.2021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Takeda K., Iwamoto S., Sugimoto H., Takuma T., Kawatani N., Noda M., Masaki A., Morise H., Arimura H., Konno K. Identity of differentiation inducing factor and tumour necrosis factor. 1986 Sep 25-Oct 1Nature. 323(6086):338–340. doi: 10.1038/323338a0. [DOI] [PubMed] [Google Scholar]
  30. Tomida M., Yamamoto-Yamaguchi Y., Hozumi M., Okabe T., Takaku F. Induction by recombinant human granulocyte colony-stimulating factor of differentiation of mouse myeloid leukemic M1 cells. FEBS Lett. 1986 Oct 27;207(2):271–275. doi: 10.1016/0014-5793(86)81503-4. [DOI] [PubMed] [Google Scholar]
  31. Tomida M., Yamamoto-Yamaguchi Y., Hozumi M. Purification of a factor inducing differentiation of mouse myeloid leukemic M1 cells from conditioned medium of mouse fibroblast L929 cells. J Biol Chem. 1984 Sep 10;259(17):10978–10982. [PubMed] [Google Scholar]
  32. Tsuchiya M., Asano S., Kaziro Y., Nagata S. Isolation and characterization of the cDNA for murine granulocyte colony-stimulating factor. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7633–7637. doi: 10.1073/pnas.83.20.7633. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. von Heijne G. Patterns of amino acids near signal-sequence cleavage sites. Eur J Biochem. 1983 Jun 1;133(1):17–21. doi: 10.1111/j.1432-1033.1983.tb07424.x. [DOI] [PubMed] [Google Scholar]

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