Abstract
Members of the Myc family of proteins share a number of protein motifs that are found in regulators of gene transcription. Conserved stretches of amino acids found in the N-terminal transcriptional activation domain of c-Myc are required for cotransforming activity. Most of the Myc proteins contain the basic helix-loop-helix zipper (bHLH-Zip) DNA-binding motif which is also required for the cotransforming activity of c-Myc. L-Myc, the product of a myc family gene that is highly amplified in many human lung carcinomas, was found to cotransform primary rat embryo cells with an activated ras gene. However, L-Myc cotransforming activity was only 1 to 10% of that of c-Myc (M. J. Birrer, S. Segal, J. S. DeGreve, F. Kaye, E. A. Sausville, and J. D. Minna, Mol. Cell. Biol. 8:2668-2673, 1988). We sought to determine whether functional differences between c-Myc and L-Myc in either the N-terminal or the C-terminal domain could account for the relatively diminished L-Myc cotransforming activity. Although the N-terminal domain of L-Myc could activate transcription when fused to the yeast GAL4 DNA-binding domain, the activity was only 5% of that of a comparable c-Myc domain. We next determined that the interaction of the C-terminal bHLH-Zip region of L-Myc or c-Myc with that of a Myc partner protein, Max, was equivalent in transfected cells. A Max expression vector was found to augment the cotransforming activity of L-Myc as well as that of c-Myc. In addition, a bacterially synthesized DNA-binding domain of L-Myc, like that o c-Myc, heterodimerizes with purified Max protein to bind the core DNA sequence CACGTG. To determine the region of L-Myc responsible for its relatively diminished cotransforming activity, we constructed chimeras containing exons 2 (constituting activation domains) and 3 (constituting DNA-binding domains) of c-Myc fused to those of L-Myc. The cotransforming potencies of these chimeras were compared with those of full-length L-Myc of c-Myc in rat embryo cells. The relative cotransforming activities suggest that the potencies of the activation domains determine the cotransforming efficiencies for c-Myc and L-Myc. This correlation supports the hypothesis that the Myc proteins function in neoplastic cotransformation as transcription factors.
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Selected References
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- Abate C., Luk D., Gentz R., Rauscher F. J., 3rd, Curran T. Expression and purification of the leucine zipper and DNA-binding domains of Fos and Jun: both Fos and Jun contact DNA directly. Proc Natl Acad Sci U S A. 1990 Feb;87(3):1032–1036. doi: 10.1073/pnas.87.3.1032. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Alitalo K., Koskinen P., Mäkelä T. P., Saksela K., Sistonen L., Winqvist R. myc oncogenes: activation and amplification. Biochim Biophys Acta. 1987 Apr 20;907(1):1–32. doi: 10.1016/0304-419x(87)90016-3. [DOI] [PubMed] [Google Scholar]
- Asker C., Steinitz M., Andersson K., Sümegi J., Klein G., Ingvarsson S. Nucleotide sequence of the rat Bmyc gene. Oncogene. 1989 Dec;4(12):1523–1527. [PubMed] [Google Scholar]
- Beckmann H., Kadesch T. The leucine zipper of TFE3 dictates helix-loop-helix dimerization specificity. Genes Dev. 1991 Jun;5(6):1057–1066. doi: 10.1101/gad.5.6.1057. [DOI] [PubMed] [Google Scholar]
- Birrer M. J., Segal S., DeGreve J. S., Kaye F., Sausville E. A., Minna J. D. L-myc cooperates with ras to transform primary rat embryo fibroblasts. Mol Cell Biol. 1988 Jun;8(6):2668–2673. doi: 10.1128/mcb.8.6.2668. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bishop J. M. Molecular themes in oncogenesis. Cell. 1991 Jan 25;64(2):235–248. doi: 10.1016/0092-8674(91)90636-d. [DOI] [PubMed] [Google Scholar]
- Blackwell T. K., Kretzner L., Blackwood E. M., Eisenman R. N., Weintraub H. Sequence-specific DNA binding by the c-Myc protein. Science. 1990 Nov 23;250(4984):1149–1151. doi: 10.1126/science.2251503. [DOI] [PubMed] [Google Scholar]
- Blackwood E. M., Eisenman R. N. Max: a helix-loop-helix zipper protein that forms a sequence-specific DNA-binding complex with Myc. Science. 1991 Mar 8;251(4998):1211–1217. doi: 10.1126/science.2006410. [DOI] [PubMed] [Google Scholar]
- Blackwood E. M., Lüscher B., Eisenman R. N. Myc and Max associate in vivo. Genes Dev. 1992 Jan;6(1):71–80. doi: 10.1101/gad.6.1.71. [DOI] [PubMed] [Google Scholar]
- Cai M., Davis R. W. Yeast centromere binding protein CBF1, of the helix-loop-helix protein family, is required for chromosome stability and methionine prototrophy. Cell. 1990 May 4;61(3):437–446. doi: 10.1016/0092-8674(90)90525-j. [DOI] [PubMed] [Google Scholar]
- Cole M. D. Myc meets its Max. Cell. 1991 May 31;65(5):715–716. doi: 10.1016/0092-8674(91)90377-b. [DOI] [PubMed] [Google Scholar]
- Cole M. D. The myc oncogene: its role in transformation and differentiation. Annu Rev Genet. 1986;20:361–384. doi: 10.1146/annurev.ge.20.120186.002045. [DOI] [PubMed] [Google Scholar]
- Dang C. V., Dolde C., Gillison M. L., Kato G. J. Discrimination between related DNA sites by a single amino acid residue of Myc-related basic-helix-loop-helix proteins. Proc Natl Acad Sci U S A. 1992 Jan 15;89(2):599–602. doi: 10.1073/pnas.89.2.599. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dang C. V., Lee W. M. Identification of the human c-myc protein nuclear translocation signal. Mol Cell Biol. 1988 Oct;8(10):4048–4054. doi: 10.1128/mcb.8.10.4048. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dang C. V., McGuire M., Buckmire M., Lee W. M. Involvement of the 'leucine zipper' region in the oligomerization and transforming activity of human c-myc protein. Nature. 1989 Feb 16;337(6208):664–666. doi: 10.1038/337664a0. [DOI] [PubMed] [Google Scholar]
- Dang C. V. c-myc oncoprotein function. Biochim Biophys Acta. 1991 Dec 10;1072(2-3):103–113. doi: 10.1016/0304-419x(91)90009-a. [DOI] [PubMed] [Google Scholar]
- De Greve J., Battey J., Fedorko J., Birrer M., Evan G., Kaye F., Sausville E., Minna J. The human L-myc gene encodes multiple nuclear phosphoproteins from alternatively processed mRNAs. Mol Cell Biol. 1988 Oct;8(10):4381–4388. doi: 10.1128/mcb.8.10.4381. [DOI] [PMC free article] [PubMed] [Google Scholar]
- DePinho R. A., Schreiber-Agus N., Alt F. W. myc family oncogenes in the development of normal and neoplastic cells. Adv Cancer Res. 1991;57:1–46. doi: 10.1016/s0065-230x(08)60994-x. [DOI] [PubMed] [Google Scholar]
- Dosaka-Akita H., Rosenberg R. K., Minna J. D., Birrer M. J. A complex pattern of translational initiation and phosphorylation in L-myc proteins. Oncogene. 1991 Mar;6(3):371–378. [PubMed] [Google Scholar]
- Halazonetis T. D., Kandil A. N. Determination of the c-MYC DNA-binding site. Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6162–6166. doi: 10.1073/pnas.88.14.6162. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Halazonetis T. D., Kandil A. N. Predicted structural similarities of the DNA binding domains of c-Myc and endonuclease Eco RI. Science. 1992 Jan 24;255(5043):464–466. doi: 10.1126/science.1734524. [DOI] [PubMed] [Google Scholar]
- Ingvarsson S., Asker C., Axelson H., Klein G., Sümegi J. Structure and expression of B-myc, a new member of the myc gene family. Mol Cell Biol. 1988 Aug;8(8):3168–3174. doi: 10.1128/mcb.8.8.3168. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Johnson P. F., McKnight S. L. Eukaryotic transcriptional regulatory proteins. Annu Rev Biochem. 1989;58:799–839. doi: 10.1146/annurev.bi.58.070189.004055. [DOI] [PubMed] [Google Scholar]
- Kato G. J., Barrett J., Villa-Garcia M., Dang C. V. An amino-terminal c-myc domain required for neoplastic transformation activates transcription. Mol Cell Biol. 1990 Nov;10(11):5914–5920. doi: 10.1128/mcb.10.11.5914. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kato G. J., Lee W. M., Chen L. L., Dang C. V. Max: functional domains and interaction with c-Myc. Genes Dev. 1992 Jan;6(1):81–92. doi: 10.1101/gad.6.1.81. [DOI] [PubMed] [Google Scholar]
- Kaye F., Battey J., Nau M., Brooks B., Seifter E., De Greve J., Birrer M., Sausville E., Minna J. Structure and expression of the human L-myc gene reveal a complex pattern of alternative mRNA processing. Mol Cell Biol. 1988 Jan;8(1):186–195. doi: 10.1128/mcb.8.1.186. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kerkhoff E., Bister K., Klempnauer K. H. Sequence-specific DNA binding by Myc proteins. Proc Natl Acad Sci U S A. 1991 May 15;88(10):4323–4327. doi: 10.1073/pnas.88.10.4323. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- Land H., Parada L. F., Weinberg R. A. Tumorigenic conversion of primary embryo fibroblasts requires at least two cooperating oncogenes. Nature. 1983 Aug 18;304(5927):596–602. doi: 10.1038/304596a0. [DOI] [PubMed] [Google Scholar]
- Landschulz W. H., Johnson P. F., McKnight S. L. The leucine zipper: a hypothetical structure common to a new class of DNA binding proteins. Science. 1988 Jun 24;240(4860):1759–1764. doi: 10.1126/science.3289117. [DOI] [PubMed] [Google Scholar]
- Lee W. M., Schwab M., Westaway D., Varmus H. E. Augmented expression of normal c-myc is sufficient for cotransformation of rat embryo cells with a mutant ras gene. Mol Cell Biol. 1985 Dec;5(12):3345–3356. doi: 10.1128/mcb.5.12.3345. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Legouy E., DePinho R., Zimmerman K., Collum R., Yancopoulos G., Mitsock L., Kriz R., Alt F. W. Structure and expression of the murine L-myc gene. EMBO J. 1987 Nov;6(11):3359–3366. doi: 10.1002/j.1460-2075.1987.tb02657.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lillie J. W., Green M. R. Transcription activation by the adenovirus E1a protein. Nature. 1989 Mar 2;338(6210):39–44. doi: 10.1038/338039a0. [DOI] [PubMed] [Google Scholar]
- Lüscher B., Eisenman R. N. New light on Myc and Myb. Part I. Myc. Genes Dev. 1990 Dec;4(12A):2025–2035. doi: 10.1101/gad.4.12a.2025. [DOI] [PubMed] [Google Scholar]
- Mitchell P. J., Tjian R. Transcriptional regulation in mammalian cells by sequence-specific DNA binding proteins. Science. 1989 Jul 28;245(4916):371–378. doi: 10.1126/science.2667136. [DOI] [PubMed] [Google Scholar]
- Murre C., McCaw P. S., Baltimore D. A new DNA binding and dimerization motif in immunoglobulin enhancer binding, daughterless, MyoD, and myc proteins. Cell. 1989 Mar 10;56(5):777–783. doi: 10.1016/0092-8674(89)90682-x. [DOI] [PubMed] [Google Scholar]
- Nau M. M., Brooks B. J., Battey J., Sausville E., Gazdar A. F., Kirsch I. R., McBride O. W., Bertness V., Hollis G. F., Minna J. D. L-myc, a new myc-related gene amplified and expressed in human small cell lung cancer. Nature. 1985 Nov 7;318(6041):69–73. doi: 10.1038/318069a0. [DOI] [PubMed] [Google Scholar]
- Prendergast G. C., Lawe D., Ziff E. B. Association of Myn, the murine homolog of max, with c-Myc stimulates methylation-sensitive DNA binding and ras cotransformation. Cell. 1991 May 3;65(3):395–407. doi: 10.1016/0092-8674(91)90457-a. [DOI] [PubMed] [Google Scholar]
- Prendergast G. C., Ziff E. B. Methylation-sensitive sequence-specific DNA binding by the c-Myc basic region. Science. 1991 Jan 11;251(4990):186–189. doi: 10.1126/science.1987636. [DOI] [PubMed] [Google Scholar]
- Roy A. L., Meisterernst M., Pognonec P., Roeder R. G. Cooperative interaction of an initiator-binding transcription initiation factor and the helix-loop-helix activator USF. Nature. 1991 Nov 21;354(6350):245–248. doi: 10.1038/354245a0. [DOI] [PubMed] [Google Scholar]
- Rustgi A. K., Dyson N., Bernards R. Amino-terminal domains of c-myc and N-myc proteins mediate binding to the retinoblastoma gene product. Nature. 1991 Aug 8;352(6335):541–544. doi: 10.1038/352541a0. [DOI] [PubMed] [Google Scholar]
- Saksela K., Mäkelä T. P., Hughes K., Woodgett J. R., Alitalo K. Activation of protein kinase C increases phosphorylation of the L-myc trans-activator domain at a GSK-3 target site. Oncogene. 1992 Feb;7(2):347–353. [PubMed] [Google Scholar]
- Sarid J., Halazonetis T. D., Murphy W., Leder P. Evolutionarily conserved regions of the human c-myc protein can be uncoupled from transforming activity. Proc Natl Acad Sci U S A. 1987 Jan;84(1):170–173. doi: 10.1073/pnas.84.1.170. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Seed B., Sheen J. Y. A simple phase-extraction assay for chloramphenicol acyltransferase activity. Gene. 1988 Jul 30;67(2):271–277. doi: 10.1016/0378-1119(88)90403-9. [DOI] [PubMed] [Google Scholar]
- Seth A., Alvarez E., Gupta S., Davis R. J. A phosphorylation site located in the NH2-terminal domain of c-Myc increases transactivation of gene expression. J Biol Chem. 1991 Dec 15;266(35):23521–23524. [PubMed] [Google Scholar]
- Skeiky Y. A., Iatrou K. Synergistic interactions of silkmoth chorion promoter-binding factors. Mol Cell Biol. 1991 Apr;11(4):1954–1964. doi: 10.1128/mcb.11.4.1954. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Smith M. J., Charron-Prochownik D. C., Prochownik E. V. The leucine zipper of c-Myc is required for full inhibition of erythroleukemia differentiation. Mol Cell Biol. 1990 Oct;10(10):5333–5339. doi: 10.1128/mcb.10.10.5333. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Spencer C. A., Groudine M. Control of c-myc regulation in normal and neoplastic cells. Adv Cancer Res. 1991;56:1–48. doi: 10.1016/s0065-230x(08)60476-5. [DOI] [PubMed] [Google Scholar]
- Stone J., de Lange T., Ramsay G., Jakobovits E., Bishop J. M., Varmus H., Lee W. Definition of regions in human c-myc that are involved in transformation and nuclear localization. Mol Cell Biol. 1987 May;7(5):1697–1709. doi: 10.1128/mcb.7.5.1697. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wenzel A., Cziepluch C., Hamann U., Schürmann J., Schwab M. The N-Myc oncoprotein is associated in vivo with the phosphoprotein Max(p20/22) in human neuroblastoma cells. EMBO J. 1991 Dec;10(12):3703–3712. doi: 10.1002/j.1460-2075.1991.tb04938.x. [DOI] [PMC free article] [PubMed] [Google Scholar]