Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1992 Jul 11;20(13):3317–3324. doi: 10.1093/nar/20.13.3317

Elk-1 protein domains required for direct and SRF-assisted DNA-binding.

R Janknecht 1, A Nordheim 1
PMCID: PMC312483  PMID: 1630903

Abstract

The Ets-related Elk-1 protein can bind to purine-rich DNA target sites in a sequence specific fashion and, in addition, can form a ternary complex with the c-fos serum response element (SRE) and the serum response factor (SRF). We demonstrate that Elk-1 can readily interchange between its different interaction partners. The amino terminal ETS-domain of Elk-1 was shown to be necessary and sufficient for direct DNA-binding activity. For ternary complex formation with the SRE and SRF, both the Elk-1 ETS-domain as well as flanking sequences up to amino acid 169 were required. Removal of sequences between the ETS-domain and amino acids 137-169 did not abolish ternary complex formation. This suggests the Elk-1 region spanning amino acids 137-169 to contain a protein-protein interaction domain. Furthermore, we have shown that a single amino acid exchange introduced into the ETS-domain can drastically alter the direct DNA-binding affinity of Elk-1 without severely affecting SRF-assisted binding to the SRE. Thus, Elk-1 requires different propensities of the ETS-domain to exert its different modes of DNA sequence recognition.

Full text

PDF
3322

Images in this article

Selected References

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

  1. Ben-David Y., Giddens E. B., Letwin K., Bernstein A. Erythroleukemia induction by Friend murine leukemia virus: insertional activation of a new member of the ets gene family, Fli-1, closely linked to c-ets-1. Genes Dev. 1991 Jun;5(6):908–918. doi: 10.1101/gad.5.6.908. [DOI] [PubMed] [Google Scholar]
  2. Bosselut R., Duvall J. F., Gégonne A., Bailly M., Hémar A., Brady J., Ghysdael J. The product of the c-ets-1 proto-oncogene and the related Ets2 protein act as transcriptional activators of the long terminal repeat of human T cell leukemia virus HTLV-1. EMBO J. 1990 Oct;9(10):3137–3144. doi: 10.1002/j.1460-2075.1990.tb07511.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Boulukos K. E., Pognonec P., Begue A., Galibert F., Gesquière J. C., Stéhelin D., Ghysdael J. Identification in chickens of an evolutionarily conserved cellular ets-2 gene (c-ets-2) encoding nuclear proteins related to the products of the c-ets proto-oncogene. EMBO J. 1988 Mar;7(3):697–705. doi: 10.1002/j.1460-2075.1988.tb02865.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Burtis K. C., Thummel C. S., Jones C. W., Karim F. D., Hogness D. S. The Drosophila 74EF early puff contains E74, a complex ecdysone-inducible gene that encodes two ets-related proteins. Cell. 1990 Apr 6;61(1):85–99. doi: 10.1016/0092-8674(90)90217-3. [DOI] [PubMed] [Google Scholar]
  5. Dalton S., Treisman R. Characterization of SAP-1, a protein recruited by serum response factor to the c-fos serum response element. Cell. 1992 Feb 7;68(3):597–612. doi: 10.1016/0092-8674(92)90194-h. [DOI] [PubMed] [Google Scholar]
  6. Feigl G., Gram M., Pongs O. A member of the steroid hormone receptor gene family is expressed in the 20-OH-ecdysone inducible puff 75B in Drosophila melanogaster. Nucleic Acids Res. 1989 Sep 25;17(18):7167–7178. doi: 10.1093/nar/17.18.7167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Golay J., Introna M., Graf T. A single point mutation in the v-ets oncogene affects both erythroid and myelomonocytic cell differentiation. Cell. 1988 Dec 23;55(6):1147–1158. doi: 10.1016/0092-8674(88)90259-0. [DOI] [PubMed] [Google Scholar]
  8. Graham R., Gilman M. Distinct protein targets for signals acting at the c-fos serum response element. Science. 1991 Jan 11;251(4990):189–192. doi: 10.1126/science.1898992. [DOI] [PubMed] [Google Scholar]
  9. Herrera R. E., Shaw P. E., Nordheim A. Occupation of the c-fos serum response element in vivo by a multi-protein complex is unaltered by growth factor induction. Nature. 1989 Jul 6;340(6228):68–70. doi: 10.1038/340068a0. [DOI] [PubMed] [Google Scholar]
  10. Hipskind R. A., Rao V. N., Mueller C. G., Reddy E. S., Nordheim A. Ets-related protein Elk-1 is homologous to the c-fos regulatory factor p62TCF. Nature. 1991 Dec 19;354(6354):531–534. doi: 10.1038/354531a0. [DOI] [PubMed] [Google Scholar]
  11. Janknecht R., Sander C., Pongs O. (HX)n repeats: a pH-controlled protein-protein interaction motif of eukaryotic transcription factors? FEBS Lett. 1991 Dec 16;295(1-3):1–2. doi: 10.1016/0014-5793(91)81369-j. [DOI] [PubMed] [Google Scholar]
  12. Janknecht R., Taube W., Lüdecke H. J., Pongs O. Characterization of a putative transcription factor gene expressed in the 20-OH-ecdysone inducible puff 74EF in Drosophila melanogaster. Nucleic Acids Res. 1989 Jun 26;17(12):4455–4464. doi: 10.1093/nar/17.12.4455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Janknecht R., de Martynoff G., Lou J., Hipskind R. A., Nordheim A., Stunnenberg H. G. Rapid and efficient purification of native histidine-tagged protein expressed by recombinant vaccinia virus. Proc Natl Acad Sci U S A. 1991 Oct 15;88(20):8972–8976. doi: 10.1073/pnas.88.20.8972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Karim F. D., Urness L. D., Thummel C. S., Klemsz M. J., McKercher S. R., Celada A., Van Beveren C., Maki R. A., Gunther C. V., Nye J. A. The ETS-domain: a new DNA-binding motif that recognizes a purine-rich core DNA sequence. Genes Dev. 1990 Sep;4(9):1451–1453. doi: 10.1101/gad.4.9.1451. [DOI] [PubMed] [Google Scholar]
  15. Klemsz M. J., McKercher S. R., Celada A., Van Beveren C., Maki R. A. The macrophage and B cell-specific transcription factor PU.1 is related to the ets oncogene. Cell. 1990 Apr 6;61(1):113–124. doi: 10.1016/0092-8674(90)90219-5. [DOI] [PubMed] [Google Scholar]
  16. Koelle M. R., Talbot W. S., Segraves W. A., Bender M. T., Cherbas P., Hogness D. S. The Drosophila EcR gene encodes an ecdysone receptor, a new member of the steroid receptor superfamily. Cell. 1991 Oct 4;67(1):59–77. doi: 10.1016/0092-8674(91)90572-g. [DOI] [PubMed] [Google Scholar]
  17. LaMarco K., Thompson C. C., Byers B. P., Walton E. M., McKnight S. L. Identification of Ets- and notch-related subunits in GA binding protein. Science. 1991 Aug 16;253(5021):789–792. doi: 10.1126/science.1876836. [DOI] [PubMed] [Google Scholar]
  18. Leprince D., Gegonne A., Coll J., de Taisne C., Schneeberger A., Lagrou C., Stehelin D. A putative second cell-derived oncogene of the avian leukaemia retrovirus E26. Nature. 1983 Nov 24;306(5941):395–397. doi: 10.1038/306395a0. [DOI] [PubMed] [Google Scholar]
  19. Lim F., Kraut N., Framptom J., Graf T. DNA binding by c-Ets-1, but not v-Ets, is repressed by an intramolecular mechanism. EMBO J. 1992 Feb;11(2):643–652. doi: 10.1002/j.1460-2075.1992.tb05096.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Matthias P., Müller M. M., Schreiber E., Rusconi S., Schaffner W. Eukaryotic expression vectors for the analysis of mutant proteins. Nucleic Acids Res. 1989 Aug 11;17(15):6418–6418. doi: 10.1093/nar/17.15.6418. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Mueller C. G., Nordheim A. A protein domain conserved between yeast MCM1 and human SRF directs ternary complex formation. EMBO J. 1991 Dec;10(13):4219–4229. doi: 10.1002/j.1460-2075.1991.tb05000.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Norman C., Runswick M., Pollock R., Treisman R. Isolation and properties of cDNA clones encoding SRF, a transcription factor that binds to the c-fos serum response element. Cell. 1988 Dec 23;55(6):989–1003. doi: 10.1016/0092-8674(88)90244-9. [DOI] [PubMed] [Google Scholar]
  23. Nunn M. F., Seeburg P. H., Moscovici C., Duesberg P. H. Tripartite structure of the avian erythroblastosis virus E26 transforming gene. Nature. 1983 Nov 24;306(5941):391–395. doi: 10.1038/306391a0. [DOI] [PubMed] [Google Scholar]
  24. Pongs O. Ecdysteroid-regulated gene expression in Drosophila melanogaster. Eur J Biochem. 1988 Aug 1;175(2):199–204. doi: 10.1111/j.1432-1033.1988.tb14184.x. [DOI] [PubMed] [Google Scholar]
  25. Pribyl L. J., Watson D. K., McWilliams M. J., Ascione R., Papas T. S. The Drosophila ets-2 gene: molecular structure, chromosomal localization, and developmental expression. Dev Biol. 1988 May;127(1):45–53. doi: 10.1016/0012-1606(88)90187-x. [DOI] [PubMed] [Google Scholar]
  26. Pribyl L. J., Watson D. K., Schulz R. A., Papas T. S. D-elg, a member of the Drosophila ets gene family: sequence, expression and evolutionary comparison. Oncogene. 1991 Jul;6(7):1175–1183. [PubMed] [Google Scholar]
  27. Rao V. N., Huebner K., Isobe M., ar-Rushdi A., Croce C. M., Reddy E. S. elk, tissue-specific ets-related genes on chromosomes X and 14 near translocation breakpoints. Science. 1989 Apr 7;244(4900):66–70. doi: 10.1126/science.2539641. [DOI] [PubMed] [Google Scholar]
  28. Rao V. N., Papas T. S., Reddy E. S. erg, a human ets-related gene on chromosome 21: alternative splicing, polyadenylation, and translation. Science. 1987 Aug 7;237(4815):635–639. doi: 10.1126/science.3299708. [DOI] [PubMed] [Google Scholar]
  29. Rao V. N., Reddy E. S. A divergent ets-related protein, elk-1, recognizes similar c-ets-1 proto-oncogene target sequences and acts as a transcriptional activator. Oncogene. 1992 Jan;7(1):65–70. [PubMed] [Google Scholar]
  30. Reddy E. S., Rao V. N. erg, an ets-related gene, codes for sequence-specific transcriptional activators. Oncogene. 1991 Dec;6(12):2285–2289. [PubMed] [Google Scholar]
  31. Rivera V. M., Greenberg M. E. Growth factor-induced gene expression: the ups and downs of c-fos regulation. New Biol. 1990 Sep;2(9):751–758. [PubMed] [Google Scholar]
  32. Schröter H., Mueller C. G., Meese K., Nordheim A. Synergism in ternary complex formation between the dimeric glycoprotein p67SRF, polypeptide p62TCF and the c-fos serum response element. EMBO J. 1990 Apr;9(4):1123–1130. doi: 10.1002/j.1460-2075.1990.tb08218.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Segraves W. A., Hogness D. S. The E75 ecdysone-inducible gene responsible for the 75B early puff in Drosophila encodes two new members of the steroid receptor superfamily. Genes Dev. 1990 Feb;4(2):204–219. doi: 10.1101/gad.4.2.204. [DOI] [PubMed] [Google Scholar]
  34. Shaw P. E., Schröter H., Nordheim A. The ability of a ternary complex to form over the serum response element correlates with serum inducibility of the human c-fos promoter. Cell. 1989 Feb 24;56(4):563–572. doi: 10.1016/0092-8674(89)90579-5. [DOI] [PubMed] [Google Scholar]
  35. Thompson C. C., Brown T. A., McKnight S. L. Convergence of Ets- and notch-related structural motifs in a heteromeric DNA binding complex. Science. 1991 Aug 16;253(5021):762–768. doi: 10.1126/science.1876833. [DOI] [PubMed] [Google Scholar]
  36. Urness L. D., Thummel C. S. Molecular interactions within the ecdysone regulatory hierarchy: DNA binding properties of the Drosophila ecdysone-inducible E74A protein. Cell. 1990 Oct 5;63(1):47–61. doi: 10.1016/0092-8674(90)90287-o. [DOI] [PubMed] [Google Scholar]
  37. Wasylyk B., Wasylyk C., Flores P., Begue A., Leprince D., Stehelin D. The c-ets proto-oncogenes encode transcription factors that cooperate with c-Fos and c-Jun for transcriptional activation. Nature. 1990 Jul 12;346(6280):191–193. doi: 10.1038/346191a0. [DOI] [PubMed] [Google Scholar]
  38. Watson D. K., McWilliams M. J., Lapis P., Lautenberger J. A., Schweinfest C. W., Papas T. S. Mammalian ets-1 and ets-2 genes encode highly conserved proteins. Proc Natl Acad Sci U S A. 1988 Nov;85(21):7862–7866. doi: 10.1073/pnas.85.21.7862. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Woods D. B., Ghysdael J., Owen M. J. Identification of nucleotide preferences in DNA sequences recognised specifically by c-Ets-1 protein. Nucleic Acids Res. 1992 Feb 25;20(4):699–704. doi: 10.1093/nar/20.4.699. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES