Palmitoylation and membrane interactions of the neuroprotective chaperone cysteine-string protein

Jennifer Greaves, Christine Salaun, Yuko Fukata, Masaki Fukata, Luke H Chamberlain

Research output: Contribution to journalArticle

73 Citations (Scopus)

Abstract

Cysteine-string protein (CSP) is an extensively palmitoylated DnaJ-family chaperone, which exerts an important neuroprotective function. Palmitoylation is required for the intracellular sorting and function of CSP, and thus it is important to understand how this essential modification of CSP is regulated. Recent work identified 23 putative palmitoyl transferases containing a conserved DHHC domain in mammalian cells, and here we show that palmitoylation of CSP is enhanced specifically by co-expression of the Golgi-localized palmitoyl transferases DHHC3, DHHC7, DHHC15, or DHHC17. Indeed, these DHHC proteins promote stable membrane attachment of CSP, which is otherwise cytosolic. An inverse correlation was identified between membrane affinity of unpalmitoylated CSP mutants and subsequent palmitoylation: mutants with an increased membrane affinity localize to the endoplasmic reticulum (ER) and are physically separated from the Golgi-localized DHHC proteins. Palmitoylation of an ER-localized mutant could be rescued by brefeldin A treatment, which promotes the mixing of ER and Golgi membranes. Interestingly though, the palmitoylated mutant remained at the ER following brefeldin A washout and did not traffic to more distal membrane compartments. We propose that CSP has a weak membrane affinity that allows the protein to locate its partner Golgi-localized DHHC proteins directly by membrane "sampling." Mutations that enhance membrane association prevent sampling and lead to accumulation of CSP on cellular membranes such as the ER. The coupling of CSP palmitoylation to Golgi membranes may thus be an important requirement for subsequent sorting.

Original languageEnglish
Pages (from-to)25014-24026
Number of pages13
JournalJournal of Biological Chemistry
Volume283
Issue number36
Early online date2 Jul 2008
DOIs
Publication statusPublished - 5 Sep 2008
Externally publishedYes

Fingerprint

Lipoylation
Membranes
Endoplasmic Reticulum
Brefeldin A
Transferases
Sorting
Proteins
cysteine string protein
Sampling
Membrane Proteins
Cells

Keywords

  • Acyltransferases
  • Animals
  • Brefeldin A
  • Cell Membrane
  • Endoplasmic Reticulum
  • Golgi Apparatus
  • HSP40 Heat-Shock Proteins
  • Humans
  • Lipoylation
  • Membrane Proteins
  • Mice
  • Mutation
  • Neuroprotective Agents
  • PC12 Cells
  • Protein Structure, Tertiary
  • Protein Synthesis Inhibitors
  • Protein Transport
  • Rats
  • Journal Article
  • Research Support, Non-U.S. Gov't

Cite this

Palmitoylation and membrane interactions of the neuroprotective chaperone cysteine-string protein. / Greaves, Jennifer; Salaun, Christine; Fukata, Yuko; Fukata, Masaki; Chamberlain, Luke H.

In: Journal of Biological Chemistry, Vol. 283, No. 36, 05.09.2008, p. 25014-24026.

Research output: Contribution to journalArticle

Greaves, Jennifer ; Salaun, Christine ; Fukata, Yuko ; Fukata, Masaki ; Chamberlain, Luke H. / Palmitoylation and membrane interactions of the neuroprotective chaperone cysteine-string protein. In: Journal of Biological Chemistry. 2008 ; Vol. 283, No. 36. pp. 25014-24026.
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T1 - Palmitoylation and membrane interactions of the neuroprotective chaperone cysteine-string protein

AU - Greaves, Jennifer

AU - Salaun, Christine

AU - Fukata, Yuko

AU - Fukata, Masaki

AU - Chamberlain, Luke H

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N2 - Cysteine-string protein (CSP) is an extensively palmitoylated DnaJ-family chaperone, which exerts an important neuroprotective function. Palmitoylation is required for the intracellular sorting and function of CSP, and thus it is important to understand how this essential modification of CSP is regulated. Recent work identified 23 putative palmitoyl transferases containing a conserved DHHC domain in mammalian cells, and here we show that palmitoylation of CSP is enhanced specifically by co-expression of the Golgi-localized palmitoyl transferases DHHC3, DHHC7, DHHC15, or DHHC17. Indeed, these DHHC proteins promote stable membrane attachment of CSP, which is otherwise cytosolic. An inverse correlation was identified between membrane affinity of unpalmitoylated CSP mutants and subsequent palmitoylation: mutants with an increased membrane affinity localize to the endoplasmic reticulum (ER) and are physically separated from the Golgi-localized DHHC proteins. Palmitoylation of an ER-localized mutant could be rescued by brefeldin A treatment, which promotes the mixing of ER and Golgi membranes. Interestingly though, the palmitoylated mutant remained at the ER following brefeldin A washout and did not traffic to more distal membrane compartments. We propose that CSP has a weak membrane affinity that allows the protein to locate its partner Golgi-localized DHHC proteins directly by membrane "sampling." Mutations that enhance membrane association prevent sampling and lead to accumulation of CSP on cellular membranes such as the ER. The coupling of CSP palmitoylation to Golgi membranes may thus be an important requirement for subsequent sorting.

AB - Cysteine-string protein (CSP) is an extensively palmitoylated DnaJ-family chaperone, which exerts an important neuroprotective function. Palmitoylation is required for the intracellular sorting and function of CSP, and thus it is important to understand how this essential modification of CSP is regulated. Recent work identified 23 putative palmitoyl transferases containing a conserved DHHC domain in mammalian cells, and here we show that palmitoylation of CSP is enhanced specifically by co-expression of the Golgi-localized palmitoyl transferases DHHC3, DHHC7, DHHC15, or DHHC17. Indeed, these DHHC proteins promote stable membrane attachment of CSP, which is otherwise cytosolic. An inverse correlation was identified between membrane affinity of unpalmitoylated CSP mutants and subsequent palmitoylation: mutants with an increased membrane affinity localize to the endoplasmic reticulum (ER) and are physically separated from the Golgi-localized DHHC proteins. Palmitoylation of an ER-localized mutant could be rescued by brefeldin A treatment, which promotes the mixing of ER and Golgi membranes. Interestingly though, the palmitoylated mutant remained at the ER following brefeldin A washout and did not traffic to more distal membrane compartments. We propose that CSP has a weak membrane affinity that allows the protein to locate its partner Golgi-localized DHHC proteins directly by membrane "sampling." Mutations that enhance membrane association prevent sampling and lead to accumulation of CSP on cellular membranes such as the ER. The coupling of CSP palmitoylation to Golgi membranes may thus be an important requirement for subsequent sorting.

KW - Acyltransferases

KW - Animals

KW - Brefeldin A

KW - Cell Membrane

KW - Endoplasmic Reticulum

KW - Golgi Apparatus

KW - HSP40 Heat-Shock Proteins

KW - Humans

KW - Lipoylation

KW - Membrane Proteins

KW - Mice

KW - Mutation

KW - Neuroprotective Agents

KW - PC12 Cells

KW - Protein Structure, Tertiary

KW - Protein Synthesis Inhibitors

KW - Protein Transport

KW - Rats

KW - Journal Article

KW - Research Support, Non-U.S. Gov't

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DO - 10.1074/jbc.M802140200

M3 - Article

VL - 283

SP - 25014

EP - 24026

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 36

ER -