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. 1997 Apr 15;16(8):1961–1969. doi: 10.1093/emboj/16.8.1961

The heat shock protein 83 (Hsp83) is required for Raf-mediated signalling in Drosophila.

A van der Straten 1, C Rommel 1, B Dickson 1, E Hafen 1
PMCID: PMC1169799  PMID: 9155022

Abstract

The heat shock protein Hsp90 has been shown to associate with various cellular signalling proteins such as steroid hormone receptors, src-like kinases and the serine/threonine kinase Raf. While the interaction between steroid hormone receptors and Hsp90 appears to be essential for ligand binding and activation of the receptors, the role of Hsp90 in Raf activation is less clear. We have identified mutations in the hsp83 gene, the Drosophila homologue of hsp90, in a search for dominant mutations that attenuate signalling from Raf in the developing eye. The mutations result in single amino acid substitutions in the Hsp83 protein and cause a dominant-negative effect on the function of the wild-type protein. We show that both wild-type and mutant forms of Hsp83 bind to the activated Drosophila Raf but the mutant Hsp83 protein causes a reduction in the kinase activity of Raf. Our results indicate that Hsp83 is essential for Raf function in vivo.

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Selected References

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  1. Aitken A. 14-3-3 proteins on the MAP. Trends Biochem Sci. 1995 Mar;20(3):95–97. doi: 10.1016/s0968-0004(00)88971-9. [DOI] [PubMed] [Google Scholar]
  2. Biggs W. H., 3rd, Zavitz K. H., Dickson B., van der Straten A., Brunner D., Hafen E., Zipursky S. L. The Drosophila rolled locus encodes a MAP kinase required in the sevenless signal transduction pathway. EMBO J. 1994 Apr 1;13(7):1628–1635. doi: 10.1002/j.1460-2075.1994.tb06426.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blackman R. K., Meselson M. Interspecific nucleotide sequence comparisons used to identify regulatory and structural features of the Drosophila hsp82 gene. J Mol Biol. 1986 Apr 20;188(4):499–515. doi: 10.1016/s0022-2836(86)80001-8. [DOI] [PubMed] [Google Scholar]
  4. Brunner D., Oellers N., Szabad J., Biggs W. H., 3rd, Zipursky S. L., Hafen E. A gain-of-function mutation in Drosophila MAP kinase activates multiple receptor tyrosine kinase signaling pathways. Cell. 1994 Mar 11;76(5):875–888. doi: 10.1016/0092-8674(94)90362-x. [DOI] [PubMed] [Google Scholar]
  5. Catelli M. G., Binart N., Jung-Testas I., Renoir J. M., Baulieu E. E., Feramisco J. R., Welch W. J. The common 90-kd protein component of non-transformed '8S' steroid receptors is a heat-shock protein. EMBO J. 1985 Dec 1;4(12):3131–3135. doi: 10.1002/j.1460-2075.1985.tb04055.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chang H. C., Solomon N. M., Wassarman D. A., Karim F. D., Therrien M., Rubin G. M., Wolff T. phyllopod functions in the fate determination of a subset of photoreceptors in Drosophila. Cell. 1995 Feb 10;80(3):463–472. doi: 10.1016/0092-8674(95)90497-2. [DOI] [PubMed] [Google Scholar]
  7. Cutforth T., Rubin G. M. Mutations in Hsp83 and cdc37 impair signaling by the sevenless receptor tyrosine kinase in Drosophila. Cell. 1994 Jul 1;77(7):1027–1036. doi: 10.1016/0092-8674(94)90442-1. [DOI] [PubMed] [Google Scholar]
  8. Dickson B. J., Domínguez M., van der Straten A., Hafen E. Control of Drosophila photoreceptor cell fates by phyllopod, a novel nuclear protein acting downstream of the Raf kinase. Cell. 1995 Feb 10;80(3):453–462. doi: 10.1016/0092-8674(95)90496-4. [DOI] [PubMed] [Google Scholar]
  9. Dickson B. J., van der Straten A., Dominguez M., Hafen E. Mutations Modulating Raf signaling in Drosophila eye development. Genetics. 1996 Jan;142(1):163–171. doi: 10.1093/genetics/142.1.163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dickson B. Nuclear factors in sevenless signalling. Trends Genet. 1995 Mar;11(3):106–111. doi: 10.1016/S0168-9525(00)89011-3. [DOI] [PubMed] [Google Scholar]
  11. Dickson B., Sprenger F., Morrison D., Hafen E. Raf functions downstream of Ras1 in the Sevenless signal transduction pathway. Nature. 1992 Dec 10;360(6404):600–603. doi: 10.1038/360600a0. [DOI] [PubMed] [Google Scholar]
  12. Evan G. I., Lewis G. K., Ramsay G., Bishop J. M. Isolation of monoclonal antibodies specific for human c-myc proto-oncogene product. Mol Cell Biol. 1985 Dec;5(12):3610–3616. doi: 10.1128/mcb.5.12.3610. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Fortini M. E., Simon M. A., Rubin G. M. Signalling by the sevenless protein tyrosine kinase is mimicked by Ras1 activation. Nature. 1992 Feb 6;355(6360):559–561. doi: 10.1038/355559a0. [DOI] [PubMed] [Google Scholar]
  14. Greenwald I., Rubin G. M. Making a difference: the role of cell-cell interactions in establishing separate identities for equivalent cells. Cell. 1992 Jan 24;68(2):271–281. doi: 10.1016/0092-8674(92)90470-w. [DOI] [PubMed] [Google Scholar]
  15. Hackett R. W., Lis J. T. Localization of the hsp83 transcript within a 3292 nucleotide sequence from the 63B heat shock locus of D. melanogaster. Nucleic Acids Res. 1983 Oct 25;11(20):7011–7030. doi: 10.1093/nar/11.20.7011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Herbst R., Carroll P. M., Allard J. D., Schilling J., Raabe T., Simon M. A. Daughter of sevenless is a substrate of the phosphotyrosine phosphatase Corkscrew and functions during sevenless signaling. Cell. 1996 Jun 14;85(6):899–909. doi: 10.1016/s0092-8674(00)81273-8. [DOI] [PubMed] [Google Scholar]
  17. Holgado-Madruga M., Emlet D. R., Moscatello D. K., Godwin A. K., Wong A. J. A Grb2-associated docking protein in EGF- and insulin-receptor signalling. Nature. 1996 Feb 8;379(6565):560–564. doi: 10.1038/379560a0. [DOI] [PubMed] [Google Scholar]
  18. Krasnow M. A., Saffman E. E., Kornfeld K., Hogness D. S. Transcriptional activation and repression by Ultrabithorax proteins in cultured Drosophila cells. Cell. 1989 Jun 16;57(6):1031–1043. doi: 10.1016/0092-8674(89)90341-3. [DOI] [PubMed] [Google Scholar]
  19. Leevers S. J., Paterson H. F., Marshall C. J. Requirement for Ras in Raf activation is overcome by targeting Raf to the plasma membrane. Nature. 1994 Jun 2;369(6479):411–414. doi: 10.1038/369411a0. [DOI] [PubMed] [Google Scholar]
  20. Lu X., Melnick M. B., Hsu J. C., Perrimon N. Genetic and molecular analyses of mutations involved in Drosophila raf signal transduction. EMBO J. 1994 Jun 1;13(11):2592–2599. doi: 10.1002/j.1460-2075.1994.tb06549.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Marais R., Light Y., Paterson H. F., Marshall C. J. Ras recruits Raf-1 to the plasma membrane for activation by tyrosine phosphorylation. EMBO J. 1995 Jul 3;14(13):3136–3145. doi: 10.1002/j.1460-2075.1995.tb07316.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Miyata Y., Yahara I. The 90-kDa heat shock protein, HSP90, binds and protects casein kinase II from self-aggregation and enhances its kinase activity. J Biol Chem. 1992 Apr 5;267(10):7042–7047. [PubMed] [Google Scholar]
  23. Mlodzik M., Hiromi Y., Weber U., Goodman C. S., Rubin G. M. The Drosophila seven-up gene, a member of the steroid receptor gene superfamily, controls photoreceptor cell fates. Cell. 1990 Jan 26;60(2):211–224. doi: 10.1016/0092-8674(90)90737-y. [DOI] [PubMed] [Google Scholar]
  24. O'Connor D., Lis J. T. Two closely linked transcription units within the 63B heat shock puff locus of D. melanogaster display strikingly different regulation. Nucleic Acids Res. 1981 Oct 10;9(19):5075–5092. doi: 10.1093/nar/9.19.5075. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Olivier J. P., Raabe T., Henkemeyer M., Dickson B., Mbamalu G., Margolis B., Schlessinger J., Hafen E., Pawson T. A Drosophila SH2-SH3 adaptor protein implicated in coupling the sevenless tyrosine kinase to an activator of Ras guanine nucleotide exchange, Sos. Cell. 1993 Apr 9;73(1):179–191. doi: 10.1016/0092-8674(93)90170-u. [DOI] [PubMed] [Google Scholar]
  26. Parsell D. A., Lindquist S. The function of heat-shock proteins in stress tolerance: degradation and reactivation of damaged proteins. Annu Rev Genet. 1993;27:437–496. doi: 10.1146/annurev.ge.27.120193.002253. [DOI] [PubMed] [Google Scholar]
  27. Pawson T. Protein modules and signalling networks. Nature. 1995 Feb 16;373(6515):573–580. doi: 10.1038/373573a0. [DOI] [PubMed] [Google Scholar]
  28. Raabe T., Riesgo-Escovar J., Liu X., Bausenwein B. S., Deak P., Maröy P., Hafen E. DOS, a novel pleckstrin homology domain-containing protein required for signal transduction between sevenless and Ras1 in Drosophila. Cell. 1996 Jun 14;85(6):911–920. doi: 10.1016/s0092-8674(00)81274-x. [DOI] [PubMed] [Google Scholar]
  29. Rogge R. D., Karlovich C. A., Banerjee U. Genetic dissection of a neurodevelopmental pathway: Son of sevenless functions downstream of the sevenless and EGF receptor tyrosine kinases. Cell. 1991 Jan 11;64(1):39–48. doi: 10.1016/0092-8674(91)90207-f. [DOI] [PubMed] [Google Scholar]
  30. Sanchez E. R., Faber L. E., Henzel W. J., Pratt W. B. The 56-59-kilodalton protein identified in untransformed steroid receptor complexes is a unique protein that exists in cytosol in a complex with both the 70- and 90-kilodalton heat shock proteins. Biochemistry. 1990 May 29;29(21):5145–5152. doi: 10.1021/bi00473a021. [DOI] [PubMed] [Google Scholar]
  31. Simon M. A., Bowtell D. D., Dodson G. S., Laverty T. R., Rubin G. M. Ras1 and a putative guanine nucleotide exchange factor perform crucial steps in signaling by the sevenless protein tyrosine kinase. Cell. 1991 Nov 15;67(4):701–716. doi: 10.1016/0092-8674(91)90065-7. [DOI] [PubMed] [Google Scholar]
  32. Simon M. A., Dodson G. S., Rubin G. M. An SH3-SH2-SH3 protein is required for p21Ras1 activation and binds to sevenless and Sos proteins in vitro. Cell. 1993 Apr 9;73(1):169–177. doi: 10.1016/0092-8674(93)90169-q. [DOI] [PubMed] [Google Scholar]
  33. Stancato L. F., Chow Y. H., Hutchison K. A., Perdew G. H., Jove R., Pratt W. B. Raf exists in a native heterocomplex with hsp90 and p50 that can be reconstituted in a cell-free system. J Biol Chem. 1993 Oct 15;268(29):21711–21716. [PubMed] [Google Scholar]
  34. Stokoe D., Macdonald S. G., Cadwallader K., Symons M., Hancock J. F. Activation of Raf as a result of recruitment to the plasma membrane. Science. 1994 Jun 3;264(5164):1463–1467. doi: 10.1126/science.7811320. [DOI] [PubMed] [Google Scholar]
  35. Therrien M., Chang H. C., Solomon N. M., Karim F. D., Wassarman D. A., Rubin G. M. KSR, a novel protein kinase required for RAS signal transduction. Cell. 1995 Dec 15;83(6):879–888. doi: 10.1016/0092-8674(95)90204-x. [DOI] [PubMed] [Google Scholar]
  36. Tsuda L., Inoue Y. H., Yoo M. A., Mizuno M., Hata M., Lim Y. M., Adachi-Yamada T., Ryo H., Masamune Y., Nishida Y. A protein kinase similar to MAP kinase activator acts downstream of the raf kinase in Drosophila. Cell. 1993 Feb 12;72(3):407–414. doi: 10.1016/0092-8674(93)90117-9. [DOI] [PubMed] [Google Scholar]
  37. Van Vactor D. L., Jr, Cagan R. L., Krämer H., Zipursky S. L. Induction in the developing compound eye of Drosophila: multiple mechanisms restrict R7 induction to a single retinal precursor cell. Cell. 1991 Dec 20;67(6):1145–1155. doi: 10.1016/0092-8674(91)90291-6. [DOI] [PubMed] [Google Scholar]
  38. Wartmann M., Davis R. J. The native structure of the activated Raf protein kinase is a membrane-bound multi-subunit complex. J Biol Chem. 1994 Mar 4;269(9):6695–6701. [PubMed] [Google Scholar]
  39. Wohlwill A. D., Bonner J. J. Genetic analysis of chromosome region 63 of Drosophila melanogaster. Genetics. 1991 Aug;128(4):763–775. doi: 10.1093/genetics/128.4.763. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Xu Y., Lindquist S. Heat-shock protein hsp90 governs the activity of pp60v-src kinase. Proc Natl Acad Sci U S A. 1993 Aug 1;90(15):7074–7078. doi: 10.1073/pnas.90.15.7074. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Zipursky S. L., Rubin G. M. Determination of neuronal cell fate: lessons from the R7 neuron of Drosophila. Annu Rev Neurosci. 1994;17:373–397. doi: 10.1146/annurev.ne.17.030194.002105. [DOI] [PubMed] [Google Scholar]

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