DHHC5 protein palmitoylates flotillin-2 and is rapidly degraded on induction of neuronal differentiation in cultured cells

J Biol Chem. 2012 Jan 2;287(1):523-530. doi: 10.1074/jbc.M111.306183. Epub 2011 Nov 11.


Post-translational palmitoylation of intracellular proteins is mediated by protein palmitoyltransferases belonging to the DHHC family, which share a common catalytic Asp-His-His-Cys (DHHC) motif. Several members have been implicated in neuronal development, neurotransmission, and synaptic plasticity. We previously observed that mice homozygous for a hypomorphic allele of the ZDHHC5 gene are impaired in context-dependent learning and memory. To identify potentially relevant protein substrates of DHHC5, we performed a quantitative proteomic analysis of stable isotope-labeled neuronal stem cell cultures from forebrains of normal and DHHC5-GT (gene-trapped) mice using the bioorthogonal palmitate analog 17-octadecynoic acid. We identified ∼300 17-octadecynoic acid-modified and hydroxylamine-sensitive proteins, of which a subset was decreased in abundance in DHHC5-GT cells. Palmitoylation and oligomerization of one of these proteins (flotillin-2) was abolished in DHHC5-GT neuronal stem cells. In COS-1 cells, overexpression of DHHC5 markedly stimulated the palmitoylation of flotillin-2, strongly suggesting a direct enzyme-substrate relationship. Serendipitously, we found that down-regulation of DHHC5 was triggered within minutes following growth factor withdrawal from normal neural stem cells, a maneuver that is used to induce neural differentiation in culture. The effect was reversible for up to 4 h, and degradation was partially prevented by inhibitors of ubiquitin-mediated proteolysis. These findings suggest that protein palmitoylation can be regulated through changes in DHHC PAT levels in response to differentiation signals.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acyltransferases
  • Animals
  • Cell Differentiation*
  • Cells, Cultured
  • Humans
  • Lipoylation*
  • Membrane Proteins / chemistry
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism*
  • Mice
  • Mutagenesis, Site-Directed
  • Mutation
  • Neural Stem Cells / cytology
  • Neural Stem Cells / metabolism
  • Neurons / cytology*
  • Neurons / metabolism*
  • Protein Multimerization
  • Protein Structure, Quaternary
  • Proteolysis*


  • Membrane Proteins
  • flotillins
  • Acyltransferases
  • DHHC5 protein, mouse