Skip to main content
NCBI home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1994 Sep;14(9):6232–6243. doi: 10.1128/mcb.14.9.6232

Dimerization through the helix-loop-helix motif enhances phosphorylation of the transcription activation domains of myogenin.

J Zhou 1, E N Olson 1
PMCID: PMC359150  PMID: 8065355

Abstract

The muscle-specific basic helix-loop-helix (bHLH) protein myogenin activates muscle transcription by binding to target sequences in muscle-specific promoters and enhancers as a heterodimer with ubiquitous bHLH proteins, such as the E2A gene products E12 and E47. We show that dimerization with E2A products potentiates phosphorylation of myogenin at sites within its amino- and carboxyl-terminal transcription activation domains. Phosphorylation of myogenin at these sites was mediated by the bHLH region of E2A products and was dependent on dimerization but not on DNA binding. Mutations of the dimerization-dependent phosphorylation sites resulted in enhanced transcriptional activity of myogenin, suggesting that their phosphorylation diminishes myogenin's transcriptional activity. The ability of E2A products to potentiate myogenin phosphorylation suggests that dimerization induces a conformational change in myogenin that unmasks otherwise cryptic phosphorylation sites or that E2A proteins recruit a kinase for which myogenin is a substrate. That phosphorylation of these dimerization-dependent sites diminished myogenin's transcriptional activity suggests that these sites are targets for a kinase that interferes with muscle-specific gene expression.

Full text

PDF
6232

Images in this article

6234Image
on p.6234
6234Image
on p.6234
6235Image
on p.6235
6236Image
on p.6236
6237Image
on p.6237
6238Image
on p.6238
6239Image
on p.6239

Selected References

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

  1. Benezra R., Davis R. L., Lockshon D., Turner D. L., Weintraub H. The protein Id: a negative regulator of helix-loop-helix DNA binding proteins. Cell. 1990 Apr 6;61(1):49–59. doi: 10.1016/0092-8674(90)90214-y. [DOI] [PubMed] [Google Scholar]
  2. Blackwell T. K., Weintraub H. Differences and similarities in DNA-binding preferences of MyoD and E2A protein complexes revealed by binding site selection. Science. 1990 Nov 23;250(4984):1104–1110. doi: 10.1126/science.2174572. [DOI] [PubMed] [Google Scholar]
  3. Blenis J. Signal transduction via the MAP kinases: proceed at your own RSK. Proc Natl Acad Sci U S A. 1993 Jul 1;90(13):5889–5892. doi: 10.1073/pnas.90.13.5889. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Braun T., Bober E., Buschhausen-Denker G., Kohtz S., Grzeschik K. H., Arnold H. H., Kotz S. Differential expression of myogenic determination genes in muscle cells: possible autoactivation by the Myf gene products. EMBO J. 1989 Dec 1;8(12):3617–3625. doi: 10.1002/j.1460-2075.1989.tb08535.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Braun T., Bober E., Winter B., Rosenthal N., Arnold H. H. Myf-6, a new member of the human gene family of myogenic determination factors: evidence for a gene cluster on chromosome 12. EMBO J. 1990 Mar;9(3):821–831. doi: 10.1002/j.1460-2075.1990.tb08179.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Braun T., Buschhausen-Denker G., Bober E., Tannich E., Arnold H. H. A novel human muscle factor related to but distinct from MyoD1 induces myogenic conversion in 10T1/2 fibroblasts. EMBO J. 1989 Mar;8(3):701–709. doi: 10.1002/j.1460-2075.1989.tb03429.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Braun T., Winter B., Bober E., Arnold H. H. Transcriptional activation domain of the muscle-specific gene-regulatory protein myf5. Nature. 1990 Aug 16;346(6285):663–665. doi: 10.1038/346663a0. [DOI] [PubMed] [Google Scholar]
  8. Brennan T. J., Chakraborty T., Olson E. N. Mutagenesis of the myogenin basic region identifies an ancient protein motif critical for activation of myogenesis. Proc Natl Acad Sci U S A. 1991 Jul 1;88(13):5675–5679. doi: 10.1073/pnas.88.13.5675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Brennan T. J., Olson E. N. Myogenin resides in the nucleus and acquires high affinity for a conserved enhancer element on heterodimerization. Genes Dev. 1990 Apr;4(4):582–595. doi: 10.1101/gad.4.4.582. [DOI] [PubMed] [Google Scholar]
  10. Chakraborty T., Brennan T. J., Li L., Edmondson D., Olson E. N. Inefficient homooligomerization contributes to the dependence of myogenin on E2A products for efficient DNA binding. Mol Cell Biol. 1991 Jul;11(7):3633–3641. doi: 10.1128/mcb.11.7.3633. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cherry J. R., Johnson T. R., Dollard C., Shuster J. R., Denis C. L. Cyclic AMP-dependent protein kinase phosphorylates and inactivates the yeast transcriptional activator ADR1. Cell. 1989 Feb 10;56(3):409–419. doi: 10.1016/0092-8674(89)90244-4. [DOI] [PubMed] [Google Scholar]
  12. Davis R. L., Weintraub H., Lassar A. B. Expression of a single transfected cDNA converts fibroblasts to myoblasts. Cell. 1987 Dec 24;51(6):987–1000. doi: 10.1016/0092-8674(87)90585-x. [DOI] [PubMed] [Google Scholar]
  13. Edmondson D. G., Brennan T. J., Olson E. N. Mitogenic repression of myogenin autoregulation. J Biol Chem. 1991 Nov 15;266(32):21343–21346. [PubMed] [Google Scholar]
  14. Edmondson D. G., Olson E. N. A gene with homology to the myc similarity region of MyoD1 is expressed during myogenesis and is sufficient to activate the muscle differentiation program. Genes Dev. 1989 May;3(5):628–640. doi: 10.1101/gad.3.5.628. [DOI] [PubMed] [Google Scholar]
  15. Emerson C. P. Myogenesis and developmental control genes. Curr Opin Cell Biol. 1990 Dec;2(6):1065–1075. doi: 10.1016/0955-0674(90)90157-a. [DOI] [PubMed] [Google Scholar]
  16. Fujisawa-Sehara A., Nabeshima Y., Hosoda Y., Obinata T., Nabeshima Y. Myogenin contains two domains conserved among myogenic factors. J Biol Chem. 1990 Sep 5;265(25):15219–15223. [PubMed] [Google Scholar]
  17. Hardy S., Kong Y., Konieczny S. F. Fibroblast growth factor inhibits MRF4 activity independently of the phosphorylation status of a conserved threonine residue within the DNA-binding domain. Mol Cell Biol. 1993 Oct;13(10):5943–5956. doi: 10.1128/mcb.13.10.5943. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hasty P., Bradley A., Morris J. H., Edmondson D. G., Venuti J. M., Olson E. N., Klein W. H. Muscle deficiency and neonatal death in mice with a targeted mutation in the myogenin gene. Nature. 1993 Aug 5;364(6437):501–506. doi: 10.1038/364501a0. [DOI] [PubMed] [Google Scholar]
  19. Henthorn P., Kiledjian M., Kadesch T. Two distinct transcription factors that bind the immunoglobulin enhancer microE5/kappa 2 motif. Science. 1990 Jan 26;247(4941):467–470. doi: 10.1126/science.2105528. [DOI] [PubMed] [Google Scholar]
  20. Hibi M., Lin A., Smeal T., Minden A., Karin M. Identification of an oncoprotein- and UV-responsive protein kinase that binds and potentiates the c-Jun activation domain. Genes Dev. 1993 Nov;7(11):2135–2148. doi: 10.1101/gad.7.11.2135. [DOI] [PubMed] [Google Scholar]
  21. Hu J. S., Olson E. N., Kingston R. E. HEB, a helix-loop-helix protein related to E2A and ITF2 that can modulate the DNA-binding ability of myogenic regulatory factors. Mol Cell Biol. 1992 Mar;12(3):1031–1042. doi: 10.1128/mcb.12.3.1031. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. James G., Olson E. Deletion of the regulatory domain of protein kinase C alpha exposes regions in the hinge and catalytic domains that mediate nuclear targeting. J Cell Biol. 1992 Feb;116(4):863–874. doi: 10.1083/jcb.116.4.863. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kadesch T. Helix-loop-helix proteins in the regulation of immunoglobulin gene transcription. Immunol Today. 1992 Jan;13(1):31–36. doi: 10.1016/0167-5699(92)90201-h. [DOI] [PubMed] [Google Scholar]
  24. Kemp B. E., Pearson R. B. Protein kinase recognition sequence motifs. Trends Biochem Sci. 1990 Sep;15(9):342–346. doi: 10.1016/0968-0004(90)90073-k. [DOI] [PubMed] [Google Scholar]
  25. Lassar A. B., Davis R. L., Wright W. E., Kadesch T., Murre C., Voronova A., Baltimore D., Weintraub H. Functional activity of myogenic HLH proteins requires hetero-oligomerization with E12/E47-like proteins in vivo. Cell. 1991 Jul 26;66(2):305–315. doi: 10.1016/0092-8674(91)90620-e. [DOI] [PubMed] [Google Scholar]
  26. Li L., Heller-Harrison R., Czech M., Olson E. N. Cyclic AMP-dependent protein kinase inhibits the activity of myogenic helix-loop-helix proteins. Mol Cell Biol. 1992 Oct;12(10):4478–4485. doi: 10.1128/mcb.12.10.4478. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Li L., Zhou J., James G., Heller-Harrison R., Czech M. P., Olson E. N. FGF inactivates myogenic helix-loop-helix proteins through phosphorylation of a conserved protein kinase C site in their DNA-binding domains. Cell. 1992 Dec 24;71(7):1181–1194. doi: 10.1016/s0092-8674(05)80066-2. [DOI] [PubMed] [Google Scholar]
  28. 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]
  29. Martin J. F., Li L., Olson E. N. Repression of myogenin function by TGF-beta 1 is targeted at the basic helix-loop-helix motif and is independent of E2A products. J Biol Chem. 1992 Jun 5;267(16):10956–10960. [PubMed] [Google Scholar]
  30. Miner J. H., Wold B. Herculin, a fourth member of the MyoD family of myogenic regulatory genes. Proc Natl Acad Sci U S A. 1990 Feb;87(3):1089–1093. doi: 10.1073/pnas.87.3.1089. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Mitsui K., Shirakata M., Paterson B. M. Phosphorylation inhibits the DNA-binding activity of MyoD homodimers but not MyoD-E12 heterodimers. J Biol Chem. 1993 Nov 15;268(32):24415–24420. [PubMed] [Google Scholar]
  32. 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]
  33. Murre C., McCaw P. S., Vaessin H., Caudy M., Jan L. Y., Jan Y. N., Cabrera C. V., Buskin J. N., Hauschka S. D., Lassar A. B. Interactions between heterologous helix-loop-helix proteins generate complexes that bind specifically to a common DNA sequence. Cell. 1989 Aug 11;58(3):537–544. doi: 10.1016/0092-8674(89)90434-0. [DOI] [PubMed] [Google Scholar]
  34. Ofir R., Dwarki V. J., Rashid D., Verma I. M. Phosphorylation of the C terminus of Fos protein is required for transcriptional transrepression of the c-fos promoter. Nature. 1990 Nov 1;348(6296):80–82. doi: 10.1038/348080a0. [DOI] [PubMed] [Google Scholar]
  35. Olson E. N. Interplay between proliferation and differentiation within the myogenic lineage. Dev Biol. 1992 Dec;154(2):261–272. doi: 10.1016/0012-1606(92)90066-p. [DOI] [PubMed] [Google Scholar]
  36. Olson E. N. MyoD family: a paradigm for development? Genes Dev. 1990 Sep;4(9):1454–1461. doi: 10.1101/gad.4.9.1454. [DOI] [PubMed] [Google Scholar]
  37. Pelech S. L., Sanghera J. S. Mitogen-activated protein kinases: versatile transducers for cell signaling. Trends Biochem Sci. 1992 Jun;17(6):233–238. doi: 10.1016/s0968-0004(00)80005-5. [DOI] [PubMed] [Google Scholar]
  38. Pownall M. E., Emerson C. P., Jr Sequential activation of three myogenic regulatory genes during somite morphogenesis in quail embryos. Dev Biol. 1992 May;151(1):67–79. doi: 10.1016/0012-1606(92)90214-2. [DOI] [PubMed] [Google Scholar]
  39. Rhodes S. J., Konieczny S. F. Identification of MRF4: a new member of the muscle regulatory factor gene family. Genes Dev. 1989 Dec;3(12B):2050–2061. doi: 10.1101/gad.3.12b.2050. [DOI] [PubMed] [Google Scholar]
  40. Sadowski I., Niedbala D., Wood K., Ptashne M. GAL4 is phosphorylated as a consequence of transcriptional activation. Proc Natl Acad Sci U S A. 1991 Dec 1;88(23):10510–10514. doi: 10.1073/pnas.88.23.10510. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Scales J. B., Olson E. N., Perry M. Two distinct Xenopus genes with homology to MyoD1 are expressed before somite formation in early embryogenesis. Mol Cell Biol. 1990 Apr;10(4):1516–1524. doi: 10.1128/mcb.10.4.1516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Schwarz J. J., Chakraborty T., Martin J., Zhou J. M., Olson E. N. The basic region of myogenin cooperates with two transcription activation domains to induce muscle-specific transcription. Mol Cell Biol. 1992 Jan;12(1):266–275. doi: 10.1128/mcb.12.1.266. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Smeal T., Binetruy B., Mercola D., Grover-Bardwick A., Heidecker G., Rapp U. R., Karin M. Oncoprotein-mediated signalling cascade stimulates c-Jun activity by phosphorylation of serines 63 and 73. Mol Cell Biol. 1992 Aug;12(8):3507–3513. doi: 10.1128/mcb.12.8.3507. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Sternberg E. A., Spizz G., Perry W. M., Vizard D., Weil T., Olson E. N. Identification of upstream and intragenic regulatory elements that confer cell-type-restricted and differentiation-specific expression on the muscle creatine kinase gene. Mol Cell Biol. 1988 Jul;8(7):2896–2909. doi: 10.1128/mcb.8.7.2896. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Tapscott S. J., Davis R. L., Thayer M. J., Cheng P. F., Weintraub H., Lassar A. B. MyoD1: a nuclear phosphoprotein requiring a Myc homology region to convert fibroblasts to myoblasts. Science. 1988 Oct 21;242(4877):405–411. doi: 10.1126/science.3175662. [DOI] [PubMed] [Google Scholar]
  46. Thayer M. J., Tapscott S. J., Davis R. L., Wright W. E., Lassar A. B., Weintraub H. Positive autoregulation of the myogenic determination gene MyoD1. Cell. 1989 Jul 28;58(2):241–248. doi: 10.1016/0092-8674(89)90838-6. [DOI] [PubMed] [Google Scholar]
  47. Weintraub H., Davis R., Lockshon D., Lassar A. MyoD binds cooperatively to two sites in a target enhancer sequence: occupancy of two sites is required for activation. Proc Natl Acad Sci U S A. 1990 Aug;87(15):5623–5627. doi: 10.1073/pnas.87.15.5623. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Weintraub H., Davis R., Tapscott S., Thayer M., Krause M., Benezra R., Blackwell T. K., Turner D., Rupp R., Hollenberg S. The myoD gene family: nodal point during specification of the muscle cell lineage. Science. 1991 Feb 15;251(4995):761–766. doi: 10.1126/science.1846704. [DOI] [PubMed] [Google Scholar]
  49. Weintraub H., Dwarki V. J., Verma I., Davis R., Hollenberg S., Snider L., Lassar A., Tapscott S. J. Muscle-specific transcriptional activation by MyoD. Genes Dev. 1991 Aug;5(8):1377–1386. doi: 10.1101/gad.5.8.1377. [DOI] [PubMed] [Google Scholar]
  50. Winter B., Braun T., Arnold H. H. Co-operativity of functional domains in the muscle-specific transcription factor Myf-5. EMBO J. 1992 May;11(5):1843–1855. doi: 10.1002/j.1460-2075.1992.tb05236.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Winter B., Braun T., Arnold H. H. cAMP-dependent protein kinase represses myogenic differentiation and the activity of the muscle-specific helix-loop-helix transcription factors Myf-5 and MyoD. J Biol Chem. 1993 May 5;268(13):9869–9878. [PubMed] [Google Scholar]
  52. Wright W. E., Sassoon D. A., Lin V. K. Myogenin, a factor regulating myogenesis, has a domain homologous to MyoD. Cell. 1989 Feb 24;56(4):607–617. doi: 10.1016/0092-8674(89)90583-7. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES