Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1997 Mar 15;322(Pt 3):853–858. doi: 10.1042/bj3220853

Activation of the ATPase activity of heat-shock proteins Hsc70/Hsp70 by cysteine-string protein.

L H Chamberlain 1, R D Burgoyne 1
PMCID: PMC1218266  PMID: 9148760

Abstract

DnaJ proteins are characterized by a 'J' domain which is homologous to a region of the Escherichia coli protein DnaJ. DnaJ has been shown to interact with the chaperone protein DnaK, and a number of eukaryotic DnaJ-like proteins have been found to interact with the 70 kDa heat-shock protein/70 kDa heat-shock cognate protein (Hsp70/Hsc70), the eukaryotic homologues of DnaK. Cysteine-string proteins (Csps) are believed to function in calcium-stimulated exocytosis and in this paper we describe a specific ATP-dependent interaction between a Csp (Csp1) and Hsc70/Hsp70. We also show that Csp1 can stimulate the ATPase activity of both Hsc70 and Hsp70 several-fold. Furthermore, we demonstrate that Csp2, a Csp variant found in adrenal chromaffin cells, can enhance the ATPase activity of Hsc70 to a similar extent as Csp1, whereas Csp(137-198), a truncated protein lacking the 'J' domain of Csp1 is unable to stimulate the ATPase activity of Hsc70. This suggests that the functions of Csp1 and Csp2 must differ in some aspect other than interaction with Hsc70. This study is also important from a general view of DnaJ/Hsc70 interactions, as Csps lack a G/F-rich region which has been suggested to be essential for activation of the ATPase activity of DnaK by DnaJ. Thus, this work would imply that a G/F-rich region is not an essential feature of DnaJ proteins for stimulation of the ATPase activity of Hsp70 proteins.

Full Text

The Full Text of this article is available as a PDF (253.3 KB).

Selected References

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

  1. Barnard R. J., Morgan A., Burgoyne R. D. Domains of alpha-SNAP required for the stimulation of exocytosis and for N-ethylmalemide-sensitive fusion protein (NSF) binding and activation. Mol Biol Cell. 1996 May;7(5):693–701. doi: 10.1091/mbc.7.5.693. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Braun J. E., Scheller R. H. Cysteine string protein, a DnaJ family member, is present on diverse secretory vesicles. Neuropharmacology. 1995 Nov;34(11):1361–1369. doi: 10.1016/0028-3908(95)00114-l. [DOI] [PubMed] [Google Scholar]
  3. Braun J. E., Wilbanks S. M., Scheller R. H. The cysteine string secretory vesicle protein activates Hsc70 ATPase. J Biol Chem. 1996 Oct 18;271(42):25989–25993. doi: 10.1074/jbc.271.42.25989. [DOI] [PubMed] [Google Scholar]
  4. Chamberlain L. H., Burgoyne R. D. Identification of a novel cysteine string protein variant and expression of cysteine string proteins in non-neuronal cells. J Biol Chem. 1996 Mar 29;271(13):7320–7323. doi: 10.1074/jbc.271.13.7320. [DOI] [PubMed] [Google Scholar]
  5. Chamberlain L. H., Henry J., Burgoyne R. D. Cysteine string proteins are associated with chromaffin granules. J Biol Chem. 1996 Aug 9;271(32):19514–19517. doi: 10.1074/jbc.271.32.19514. [DOI] [PubMed] [Google Scholar]
  6. Cheetham M. E., Jackson A. P., Anderton B. H. Regulation of 70-kDa heat-shock-protein ATPase activity and substrate binding by human DnaJ-like proteins, HSJ1a and HSJ1b. Eur J Biochem. 1994 Nov 15;226(1):99–107. doi: 10.1111/j.1432-1033.1994.tb20030.x. [DOI] [PubMed] [Google Scholar]
  7. Cyr D. M., Langer T., Douglas M. G. DnaJ-like proteins: molecular chaperones and specific regulators of Hsp70. Trends Biochem Sci. 1994 Apr;19(4):176–181. doi: 10.1016/0968-0004(94)90281-x. [DOI] [PubMed] [Google Scholar]
  8. Gething M. J., Sambrook J. Protein folding in the cell. Nature. 1992 Jan 2;355(6355):33–45. doi: 10.1038/355033a0. [DOI] [PubMed] [Google Scholar]
  9. Gundersen C. B., Umbach J. A. Suppression cloning of the cDNA for a candidate subunit of a presynaptic calcium channel. Neuron. 1992 Sep;9(3):527–537. doi: 10.1016/0896-6273(92)90190-o. [DOI] [PubMed] [Google Scholar]
  10. Karzai A. W., McMacken R. A bipartite signaling mechanism involved in DnaJ-mediated activation of the Escherichia coli DnaK protein. J Biol Chem. 1996 May 10;271(19):11236–11246. doi: 10.1074/jbc.271.19.11236. [DOI] [PubMed] [Google Scholar]
  11. Lanzetta P. A., Alvarez L. J., Reinach P. S., Candia O. A. An improved assay for nanomole amounts of inorganic phosphate. Anal Biochem. 1979 Nov 15;100(1):95–97. doi: 10.1016/0003-2697(79)90115-5. [DOI] [PubMed] [Google Scholar]
  12. Lill R., Dowhan W., Wickner W. The ATPase activity of SecA is regulated by acidic phospholipids, SecY, and the leader and mature domains of precursor proteins. Cell. 1990 Jan 26;60(2):271–280. doi: 10.1016/0092-8674(90)90742-w. [DOI] [PubMed] [Google Scholar]
  13. Mastrogiacomo A., Gundersen C. B. The nucleotide and deduced amino acid sequence of a rat cysteine string protein. Brain Res Mol Brain Res. 1995 Jan;28(1):12–18. doi: 10.1016/0169-328x(94)00172-b. [DOI] [PubMed] [Google Scholar]
  14. Mastrogiacomo A., Parsons S. M., Zampighi G. A., Jenden D. J., Umbach J. A., Gundersen C. B. Cysteine string proteins: a potential link between synaptic vesicles and presynaptic Ca2+ channels. Science. 1994 Feb 18;263(5149):981–982. doi: 10.1126/science.7906056. [DOI] [PubMed] [Google Scholar]
  15. Morgan A., Dimaline R., Burgoyne R. D. The ATPase activity of N-ethylmaleimide-sensitive fusion protein (NSF) is regulated by soluble NSF attachment proteins. J Biol Chem. 1994 Nov 25;269(47):29347–29350. [PubMed] [Google Scholar]
  16. Palleros D. R., Reid K. L., Shi L., Welch W. J., Fink A. L. ATP-induced protein-Hsp70 complex dissociation requires K+ but not ATP hydrolysis. Nature. 1993 Oct 14;365(6447):664–666. doi: 10.1038/365664a0. [DOI] [PubMed] [Google Scholar]
  17. Rassow J., Voos W., Pfanner N. Partner proteins determine multiple functions of Hsp70. Trends Cell Biol. 1995 May;5(5):207–212. doi: 10.1016/s0962-8924(00)89001-7. [DOI] [PubMed] [Google Scholar]
  18. Schlossman D. M., Schmid S. L., Braell W. A., Rothman J. E. An enzyme that removes clathrin coats: purification of an uncoating ATPase. J Cell Biol. 1984 Aug;99(2):723–733. doi: 10.1083/jcb.99.2.723. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Sugito K., Yamane M., Hattori H., Hayashi Y., Tohnai I., Ueda M., Tsuchida N., Ohtsuka K. Interaction between hsp70 and hsp40, eukaryotic homologues of DnaK and DnaJ, in human cells expressing mutant-type p53. FEBS Lett. 1995 Jan 23;358(2):161–164. doi: 10.1016/0014-5793(94)01417-y. [DOI] [PubMed] [Google Scholar]
  20. Szabo A., Korszun R., Hartl F. U., Flanagan J. A zinc finger-like domain of the molecular chaperone DnaJ is involved in binding to denatured protein substrates. EMBO J. 1996 Jan 15;15(2):408–417. [PMC free article] [PubMed] [Google Scholar]
  21. Szyperski T., Pellecchia M., Wall D., Georgopoulos C., Wüthrich K. NMR structure determination of the Escherichia coli DnaJ molecular chaperone: secondary structure and backbone fold of the N-terminal region (residues 2-108) containing the highly conserved J domain. Proc Natl Acad Sci U S A. 1994 Nov 22;91(24):11343–11347. doi: 10.1073/pnas.91.24.11343. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Südhof T. C. The synaptic vesicle cycle: a cascade of protein-protein interactions. Nature. 1995 Jun 22;375(6533):645–653. doi: 10.1038/375645a0. [DOI] [PubMed] [Google Scholar]
  23. Umbach J. A., Zinsmaier K. E., Eberle K. K., Buchner E., Benzer S., Gundersen C. B. Presynaptic dysfunction in Drosophila csp mutants. Neuron. 1994 Oct;13(4):899–907. doi: 10.1016/0896-6273(94)90255-0. [DOI] [PubMed] [Google Scholar]
  24. Ungewickell E., Ungewickell H., Holstein S. E., Lindner R., Prasad K., Barouch W., Martin B., Greene L. E., Eisenberg E. Role of auxilin in uncoating clathrin-coated vesicles. Nature. 1995 Dec 7;378(6557):632–635. doi: 10.1038/378632a0. [DOI] [PubMed] [Google Scholar]
  25. Wall D., Zylicz M., Georgopoulos C. The NH2-terminal 108 amino acids of the Escherichia coli DnaJ protein stimulate the ATPase activity of DnaK and are sufficient for lambda replication. J Biol Chem. 1994 Feb 18;269(7):5446–5451. [PubMed] [Google Scholar]
  26. Wall D., Zylicz M., Georgopoulos C. The conserved G/F motif of the DnaJ chaperone is necessary for the activation of the substrate binding properties of the DnaK chaperone. J Biol Chem. 1995 Feb 3;270(5):2139–2144. doi: 10.1074/jbc.270.5.2139. [DOI] [PubMed] [Google Scholar]
  27. Zinsmaier K. E., Eberle K. K., Buchner E., Walter N., Benzer S. Paralysis and early death in cysteine string protein mutants of Drosophila. Science. 1994 Feb 18;263(5149):977–980. doi: 10.1126/science.8310297. [DOI] [PubMed] [Google Scholar]
  28. Zinsmaier K. E., Hofbauer A., Heimbeck G., Pflugfelder G. O., Buchner S., Buchner E. A cysteine-string protein is expressed in retina and brain of Drosophila. J Neurogenet. 1990 Nov;7(1):15–29. doi: 10.3109/01677069009084150. [DOI] [PubMed] [Google Scholar]
  29. van de Goor J., Ramaswami M., Kelly R. Redistribution of synaptic vesicles and their proteins in temperature-sensitive shibire(ts1) mutant Drosophila. Proc Natl Acad Sci U S A. 1995 Jun 6;92(12):5739–5743. doi: 10.1073/pnas.92.12.5739. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES