The cysteine string secretory vesicle protein activates Hsc70 ATPase.
Cysteine string protein (CSP) is a 34 kDa secretory vesicle protein bearing a "J-domain" as well as a palmitoylated cysteine-rich "string" region used for membrane attachment. Mutation of the CSP gene causes impaired presynaptic neuromuscular transmission in Drosophila melanogaster, implicating CSP as part of the exocytotic protein machinery. The J-domain of CSP shares homology with the universally conserved DnaJ family, a group of proteins that act as co-chaperones with Hsc70 and its homologs. Hsc70 is an abundant neural protein with coupled protein binding and ATPase activities. We have investigated the CSP modulation of Hsc70 ATPase activity. Here we demonstrated that CSP enhances Hsc70 ATPase activity in a dose-dependent manner. CSP activation of Hsc70 was maximal ( approximately 12 times) at 1:1 stoichiometry and above. We show that a J-domain-containing fragment (amino acids 1-82) of CSP is sufficient for the activation of Hsc70. Neither CSP nor the amino-terminal fragment stimulate the activity of the isolated Hsc70 ATPase domain (amino acids 1-386). CSP does not significantly increase the activity of N-ethylmaleimide-sensitive fusion protein, another ATPase required for transport vesicle function. Our results suggest that CSP, a DnaJ family member associated with the secretory vesicle cycle regulates Hsc70 functions. Hsc70 may function within the biochemical pathways of exo- and endocytosis to promote the formation or dissociation of multimeric complexes or to regulate conformational changes.
Braun
J E
JE
Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University Medical Center, Stanford, California 94305-5428, USA.
Wilbanks
S M
SM
Scheller
R H
RH
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D001923:Brain Chemistry; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D004789:Enzyme Activation; D050883:HSC70 Heat-Shock Proteins; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity
D002352:Carrier Proteins; D050883:HSC70 Heat-Shock Proteins; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011994:Recombinant Proteins; C088460:cysteine string protein; D000251:Adenosine Triphosphatases