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. 1995 Feb;15(2):925–931. doi: 10.1128/mcb.15.2.925

ets-1 in astrocytes: expression and transmitter-evoked phosphorylation.

L F Fleischman 1, L Holtzclaw 1, J T Russell 1, G Mavrothalassitis 1, R J Fisher 1
PMCID: PMC231978  PMID: 7823957

Abstract

The ets-1 protein has been primarily studied as a sequence-specific transcriptional regulator that is predominately expressed in lymphoid cells. In this report, we show that ets-1 is also expressed in astrocytes and astrocytoma cells and is regulated during both signal transduction and differentiation. Both isoforms of ets-1, p51 and p42, were found in astrocytes and astrocytoma cells, but whereas expression of p51 was strong, p42, the alternate splice product previously shown to lack the phosphorylation domain, was difficult to detect and was present at a level 10- to 40-fold lower than that of p51. This differed by roughly an order of magnitude from the ratio generally observable in T cells and thymocytes. In two astrocytoma lines of human origin, CCF and 1321N1, ets-1 phosphorylation was stimulated by bradykinin and carbachol, respectively. Glutamate, norepinephrine, and bradykinin elicited phosphorylation of p51 in cultures of primary rat type 1 astrocytes. ets-1 phosphorylation was dramatically blocked by KT5926, an inhibitor of myosin light-chain kinase, suggesting that this kinase may be involved in phosphorylation of ets-1 in vivo. Investigations of retinoic acid-induced differentiation in P19 cells provided further support for a strong correlation of ets-1 with the pathway for astrocyte differentiation.

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Selected References

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  1. Amouyel P., Gégonne A., Delacourte A., Défossez A., Stéhelin D. Expression of ETS proto-oncogenes in astrocytes in human cortex. Brain Res. 1988 Apr 26;447(1):149–153. doi: 10.1016/0006-8993(88)90976-6. [DOI] [PubMed] [Google Scholar]
  2. Ashkenazi A., Ramachandran J., Capon D. J. Acetylcholine analogue stimulates DNA synthesis in brain-derived cells via specific muscarinic receptor subtypes. Nature. 1989 Jul 13;340(6229):146–150. doi: 10.1038/340146a0. [DOI] [PubMed] [Google Scholar]
  3. Baorto D. M., Mellado W., Shelanski M. L. Astrocyte process growth induction by actin breakdown. J Cell Biol. 1992 Apr;117(2):357–367. doi: 10.1083/jcb.117.2.357. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bhat N. K., Fisher R. J., Fujiwara S., Ascione R., Papas T. S. Temporal and tissue-specific expression of mouse ets genes. Proc Natl Acad Sci U S A. 1987 May;84(10):3161–3165. doi: 10.1073/pnas.84.10.3161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bhat N. K., Komschlies K. L., Fujiwara S., Fisher R. J., Mathieson B. J., Gregorio T. A., Young H. A., Kasik J. W., Ozato K., Papas T. S. Expression of ets genes in mouse thymocyte subsets and T cells. J Immunol. 1989 Jan 15;142(2):672–678. [PubMed] [Google Scholar]
  6. Bhat N. K., Thompson C. B., Lindsten T., June C. H., Fujiwara S., Koizumi S., Fisher R. J., Papas T. S. Reciprocal expression of human ETS1 and ETS2 genes during T-cell activation: regulatory role for the protooncogene ETS1. Proc Natl Acad Sci U S A. 1990 May;87(10):3723–3727. doi: 10.1073/pnas.87.10.3723. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. 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]
  8. 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]
  9. Chen J. H. The proto-oncogene c-ets is preferentially expressed in lymphoid cells. Mol Cell Biol. 1985 Nov;5(11):2993–3000. doi: 10.1128/mcb.5.11.2993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cleveland D. W., Fischer S. G., Kirschner M. W., Laemmli U. K. Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. J Biol Chem. 1977 Feb 10;252(3):1102–1106. [PubMed] [Google Scholar]
  11. Cornell-Bell A. H., Finkbeiner S. M., Cooper M. S., Smith S. J. Glutamate induces calcium waves in cultured astrocytes: long-range glial signaling. Science. 1990 Jan 26;247(4941):470–473. doi: 10.1126/science.1967852. [DOI] [PubMed] [Google Scholar]
  12. Dani J. W., Chernjavsky A., Smith S. J. Neuronal activity triggers calcium waves in hippocampal astrocyte networks. Neuron. 1992 Mar;8(3):429–440. doi: 10.1016/0896-6273(92)90271-e. [DOI] [PubMed] [Google Scholar]
  13. Fisher R. J., Fivash M., Casas-Finet J., Erickson J. W., Kondoh A., Bladen S. V., Fisher C., Watson D. K., Papas T. Real-time DNA binding measurements of the ETS1 recombinant oncoproteins reveal significant kinetic differences between the p42 and p51 isoforms. Protein Sci. 1994 Feb;3(2):257–266. doi: 10.1002/pro.5560030210. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fisher R. J., Mavrothalassitis G., Kondoh A., Papas T. S. High-affinity DNA-protein interactions of the cellular ETS1 protein: the determination of the ETS binding motif. Oncogene. 1991 Dec;6(12):2249–2254. [PubMed] [Google Scholar]
  15. Fleischman L. F., Pilaro A. M., Murakami K., Kondoh A., Fisher R. J., Papas T. S. c-Ets-1 protein is hyperphosphorylated during mitosis. Oncogene. 1993 Mar;8(3):771–780. [PubMed] [Google Scholar]
  16. Fujiwara S., Fisher R. J., Bhat N. K., Diaz de la Espina S. M., Papas T. S. A short-lived nuclear phosphoprotein encoded by the human ets-2 proto-oncogene is stabilized by activation of protein kinase C. Mol Cell Biol. 1988 Nov;8(11):4700–4706. doi: 10.1128/mcb.8.11.4700. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Fujiwara S., Fisher R. J., Seth A., Bhat N. K., Showalter S. D., Zweig M., Papas T. S. Characterization and localization of the products of the human homologs of the v-ets oncogene. Oncogene. 1988 Feb;2(2):99–103. [PubMed] [Google Scholar]
  18. Fujiwara S., Koizumi S., Fisher R. J., Bhat N. K., Papas T. S. Phosphorylation of the ETS-2 protein: regulation by the T-cell antigen receptor-CD3 complex. Mol Cell Biol. 1990 Mar;10(3):1249–1253. doi: 10.1128/mcb.10.3.1249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Ghysdael J., Gegonne A., Pognonec P., Dernis D., Leprince D., Stehelin D. Identification and preferential expression in thymic and bursal lymphocytes of a c-ets oncogene-encoded Mr 54,000 cytoplasmic protein. Proc Natl Acad Sci U S A. 1986 Mar;83(6):1714–1718. doi: 10.1073/pnas.83.6.1714. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Gunther C. V., Nye J. A., Bryner R. S., Graves B. J. Sequence-specific DNA binding of the proto-oncoprotein ets-1 defines a transcriptional activator sequence within the long terminal repeat of the Moloney murine sarcoma virus. Genes Dev. 1990 Apr;4(4):667–679. doi: 10.1101/gad.4.4.667. [DOI] [PubMed] [Google Scholar]
  21. 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]
  22. Ho I. C., Bhat N. K., Gottschalk L. R., Lindsten T., Thompson C. B., Papas T. S., Leiden J. M. Sequence-specific binding of human Ets-1 to the T cell receptor alpha gene enhancer. Science. 1990 Nov 9;250(4982):814–818. doi: 10.1126/science.2237431. [DOI] [PubMed] [Google Scholar]
  23. Jones-Villeneuve E. M., McBurney M. W., Rogers K. A., Kalnins V. I. Retinoic acid induces embryonal carcinoma cells to differentiate into neurons and glial cells. J Cell Biol. 1982 Aug;94(2):253–262. doi: 10.1083/jcb.94.2.253. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Jorcyk C. L., Watson D. K., Mavrothalassitis G. J., Papas T. S. The human ETS1 gene: genomic structure, promoter characterization and alternative splicing. Oncogene. 1991 Apr;6(4):523–532. [PubMed] [Google Scholar]
  25. 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]
  26. Klämbt C. The Drosophila gene pointed encodes two ETS-like proteins which are involved in the development of the midline glial cells. Development. 1993 Jan;117(1):163–176. doi: 10.1242/dev.117.1.163. [DOI] [PubMed] [Google Scholar]
  27. Koizumi S., Fisher R. J., Fujiwara S., Jorcyk C., Bhat N. K., Seth A., Papas T. S. Isoforms of the human ets-1 protein: generation by alternative splicing and differential phosphorylation. Oncogene. 1990 May;5(5):675–681. [PubMed] [Google Scholar]
  28. Macleod K., Leprince D., Stehelin D. The ets gene family. Trends Biochem Sci. 1992 Jul;17(7):251–256. doi: 10.1016/0968-0004(92)90404-w. [DOI] [PubMed] [Google Scholar]
  29. Maroulakou I. G., Papas T. S., Green J. E. Differential expression of ets-1 and ets-2 proto-oncogenes during murine embryogenesis. Oncogene. 1994 Jun;9(6):1551–1565. [PubMed] [Google Scholar]
  30. McCarthy K. D., de Vellis J. Preparation of separate astroglial and oligodendroglial cell cultures from rat cerebral tissue. J Cell Biol. 1980 Jun;85(3):890–902. doi: 10.1083/jcb.85.3.890. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. McDonough P. M., Eubanks J. H., Brown J. H. Desensitization and recovery of muscarinic and histaminergic Ca2+ mobilization in 1321N1 astrocytoma cells. Biochem J. 1988 Jan 1;249(1):135–141. doi: 10.1042/bj2490135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Nakahata N., Harden T. K. Regulation of inositol trisphosphate accumulation by muscarinic cholinergic and H1-histamine receptors on human astrocytoma cells. Differential induction of desensitization by agonists. Biochem J. 1987 Jan 15;241(2):337–344. doi: 10.1042/bj2410337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Nakanishi S., Catt K. J., Balla T. Inhibition of agonist-stimulated inositol 1,4,5-trisphosphate production and calcium signaling by the myosin light chain kinase inhibitor, wortmannin. J Biol Chem. 1994 Mar 4;269(9):6528–6535. [PubMed] [Google Scholar]
  34. Nakanishi S., Yamada K., Iwahashi K., Kuroda K., Kase H. KT5926, a potent and selective inhibitor of myosin light chain kinase. Mol Pharmacol. 1990 Apr;37(4):482–488. [PubMed] [Google Scholar]
  35. Noronha-Blob L., Richard C., U'Prichard D. C. Calcium mobilization by muscarinic receptors in human astrocytoma cells: measurements with quin 2. Biochem Biophys Res Commun. 1987 Aug 31;147(1):182–188. doi: 10.1016/s0006-291x(87)80104-3. [DOI] [PubMed] [Google Scholar]
  36. Parpura V., Basarsky T. A., Liu F., Jeftinija K., Jeftinija S., Haydon P. G. Glutamate-mediated astrocyte-neuron signalling. Nature. 1994 Jun 30;369(6483):744–747. doi: 10.1038/369744a0. [DOI] [PubMed] [Google Scholar]
  37. Pognonec P., Boulukos K. E., Gesquière J. C., Stéhelin D., Ghysdael J. Mitogenic stimulation of thymocytes results in the calcium-dependent phosphorylation of c-ets-1 proteins. EMBO J. 1988 Apr;7(4):977–983. doi: 10.1002/j.1460-2075.1988.tb02904.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pongubala J. M., Van Beveren C., Nagulapalli S., Klemsz M. J., McKercher S. R., Maki R. A., Atchison M. L. Effect of PU.1 phosphorylation on interaction with NF-EM5 and transcriptional activation. Science. 1993 Mar 12;259(5101):1622–1625. doi: 10.1126/science.8456286. [DOI] [PubMed] [Google Scholar]
  39. 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]
  40. 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]
  41. Sacchi N., de Klein A., Showalter S. D., Bigi G., Papas T. S. High expression of ets-1 gene in human thymocytes and immature T leukemic cells. Leukemia. 1988 Jan;2(1):12–18. [PubMed] [Google Scholar]
  42. Saitoh M., Naka M., Hidaka H. The modulatory role of myosin light chain phosphorylation in human platelet activation. Biochem Biophys Res Commun. 1986 Oct 15;140(1):280–287. doi: 10.1016/0006-291x(86)91087-9. [DOI] [PubMed] [Google Scholar]
  43. Scholz W. K., Baitinger C., Schulman H., Kelly P. T. Developmental changes in Ca2+/calmodulin-dependent protein kinase II in cultures of hippocampal pyramidal neurons and astrocytes. J Neurosci. 1988 Mar;8(3):1039–1051. doi: 10.1523/JNEUROSCI.08-03-01039.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Thiele C. J., Deutsch L. A., Israel M. A. The expression of multiple proto-oncogenes is differentially regulated during retinoic acid induced maturation of human neuroblastoma cell lines. Oncogene. 1988 Sep;3(3):281–288. [PubMed] [Google Scholar]
  45. 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]
  46. Wasylyk C., Flores P., Gutman A., Wasylyk B. PEA3 is a nuclear target for transcription activation by non-nuclear oncogenes. EMBO J. 1989 Nov;8(11):3371–3378. doi: 10.1002/j.1460-2075.1989.tb08500.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Wasylyk C., Gutman A., Nicholson R., Wasylyk B. The c-Ets oncoprotein activates the stromelysin promoter through the same elements as several non-nuclear oncoproteins. EMBO J. 1991 May;10(5):1127–1134. doi: 10.1002/j.1460-2075.1991.tb08053.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Watson D. K., McWilliams-Smith M. J., Nunn M. F., Duesberg P. H., O'Brien S. J., Papas T. S. The ets sequence from the transforming gene of avian erythroblastosis virus, E26, has unique domains on human chromosomes 11 and 21: both loci are transcriptionally active. Proc Natl Acad Sci U S A. 1985 Nov;82(21):7294–7298. doi: 10.1073/pnas.82.21.7294. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Wernert N., Raes M. B., Lassalle P., Dehouck M. P., Gosselin B., Vandenbunder B., Stehelin D. c-ets1 proto-oncogene is a transcription factor expressed in endothelial cells during tumor vascularization and other forms of angiogenesis in humans. Am J Pathol. 1992 Jan;140(1):119–127. [PMC free article] [PubMed] [Google Scholar]
  50. Xin J. H., Cowie A., Lachance P., Hassell J. A. Molecular cloning and characterization of PEA3, a new member of the Ets oncogene family that is differentially expressed in mouse embryonic cells. Genes Dev. 1992 Mar;6(3):481–496. doi: 10.1101/gad.6.3.481. [DOI] [PubMed] [Google Scholar]
  51. Yagodin S. V., Holtzclaw L., Sheppard C. A., Russell J. T. Nonlinear propagation of agonist-induced cytoplasmic calcium waves in single astrocytes. J Neurobiol. 1994 Mar;25(3):265–280. doi: 10.1002/neu.480250307. [DOI] [PubMed] [Google Scholar]
  52. Yano S., Fukunaga K., Ushio Y., Miyamoto E. Activation of Ca2+/calmodulin-dependent protein kinase II and phosphorylation of intermediate filament proteins by stimulation of glutamate receptors in cultured rat cortical astrocytes. J Biol Chem. 1994 Feb 18;269(7):5428–5439. [PubMed] [Google Scholar]
  53. Yoshikawa K., Aizawa T., Hayashi Y. Degeneration in vitro of post-mitotic neurons overexpressing the Alzheimer amyloid protein precursor. Nature. 1992 Sep 3;359(6390):64–67. doi: 10.1038/359064a0. [DOI] [PubMed] [Google Scholar]

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