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Mutational analysis of cysteine-string protein function in insulin exocytosis

Contributors

Zhang, H.; Kelley, William; Chamberlain, L. H.; Burgoyne, R. D.; Lang, Jochen

Published in

Journal of cell science, vol. 112, no. 9, p. 1345-1351

Publication date

1999

Abstract

Cysteine-string proteins (Csps) are vesicle proteins involved in neurotransmission. They contain at least four domains: an N-terminal J-domain which can interact with the chaperone Hsc70, an adjacent linker region, the defining cysteine rich domain and a variable C terminus. As the relevance of these domains for the function of Csps in exocytosis is unknown, we have performed a mutational analysis of Csp domains using insulin release by large dense core vesicles (LDCVs) as a model of regulated exocytosis. All mutants were apparently palmitoylated and their subcellular distribution was similar to endogenous Csp. Point mutations within the highly conserved HPD motif of the J-domain abolished activation of Hsc70. However, these mutations altered the effect of Csp on exocytosis only after additional truncation of the extreme C terminus as found in the Csp splice variant Csp2. Furthermore, the strikingly conserved linker region adjacent to the J-domain was important for Csp function in exocytosis, but not for the activation of Hsc70 ATPase. The effects of Csp wild-type or mutants were preserved in permeabilized cells excluding an effect on transmembrane ion fluxes. These observations demonstrate a functional difference between the two isoforms and suggest a role for the J-domain co-chaperone function as well as for the newly defined linker region in LDCV exocytosis.

Keywords

  • Cells, Cultured
  • Exocytosis/ physiology
  • HSP40 Heat-Shock Proteins
  • Humans
  • Insulin/ secretion
  • Membrane Proteins/ genetics
  • Mutagenesis, Site-Directed
  • Nerve Tissue Proteins/ genetics
  • Point Mutation
  • Subcellular Fractions/chemistry

UNIGE affiliation entities

Citation (ISO format)

ZHANG, H. et al. Mutational analysis of cysteine-string protein function in insulin exocytosis. In: Journal of cell science, 1999, vol. 112, n° 9, p. 1345–1351.

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Article

Identifiers

Additional URL for this publication

http://jcs.biologists.org/cgi/content/abstract/112/9/1345

Journal ISSN

0021-9533
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332downloads

Technical informations

Creation

21/06/2010 10:27:17

First validation

21/06/2010 10:27:17

Update

14/03/2023 15:46:21

Status update

14/03/2023 15:46:21

Last indexation

29/10/2024 15:22:35
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