Huntingtin-Interacting Protein HIP14 Is a Palmitoyl Transferase Involved in Palmitoylation and Trafficking of Multiple Neuronal Proteins

In neurons, posttranslational modification by palmitate regulates the trafficking and function of signaling molecules, neurotransmitter receptors, and associated synaptic scaffolding proteins. However, the enzymatic machinery involved in protein palmitoylation has remained elusive. Here, using biochemical assays, we show that huntingtin (htt) interacting protein, HIP14, is a neuronal palmitoyl transferase (PAT). HIP14 shows remarkable substrate specificity for neuronal proteins, including SNAP-25, PSD-95, GAD65, synaptotagmin I, and htt. Conversely, HIP14 is catalytically invariant toward paralemmin and synaptotagmin VII. Exogenous HIP14 enhances palmitoylation-dependent vesicular trafficking of several acylated proteins in both heterologous cells and neurons. Moreover, interference with endogenous expression of HIP14 reduces clustering of PSD-95 and GAD65 in neurons. These findings define HIP14 as a mammalian palmitoyl transferase involved in the palmitoylation and trafficking of multiple neuronal proteins.

[1]  M. Sheng,et al.  Development of neuron–neuron synapses , 2000, Current Opinion in Neurobiology.

[2]  T. Murphy,et al.  Regulation of dendritic branching and filopodia formation in hippocampal neurons by specific acylated protein motifs. , 2004, Molecular biology of the cell.

[3]  Asher Mullard,et al.  Presynaptic Trafficking of Synaptotagmin I Is Regulated by Protein Palmitoylation* , 2004, Journal of Biological Chemistry.

[4]  M. Linder,et al.  SNAP-25 palmitoylation and plasma membrane targeting require a functional secretory pathway. , 1998, Molecular biology of the cell.

[5]  D. Bredt,et al.  Synaptic Targeting of the Postsynaptic Density Protein PSD-95 Mediated by Lipid and Protein Motifs , 1999, Neuron.

[6]  P. Beachy,et al.  Skinny Hedgehog, an Acyltransferase Required for Palmitoylation and Activity of the Hedgehog Signal , 2001, Science.

[7]  P. Washbourne,et al.  Cysteine residues of SNAP-25 are required for SNARE disassembly and exocytosis, but not for membrane targeting. , 2001, The Biochemical journal.

[8]  T. Südhof,et al.  Fatty acylation of synaptotagmin in PC12 cells and synaptosomes. , 1996, Biochemical and biophysical research communications.

[9]  M. Schmidt.  Fatty acylation of proteins , 1989, Biochimica et Biophysica Acta (BBA) - Reviews on Biomembranes.

[10]  Linyi Chen,et al.  The yeast DHHC cysteine-rich domain protein Akr1p is a palmitoyl transferase , 2002, The Journal of cell biology.

[11]  Maria Karayiorgou,et al.  Evidence that the gene encoding ZDHHC8 contributes to the risk of schizophrenia , 2004, Nature Genetics.

[12]  E. Petrasch‐Parwez,et al.  Paralemmin, a Prenyl-Palmitoyl–anchored Phosphoprotein Abundant in Neurons and Implicated in Plasma Membrane Dynamics and Cell Process Formation , 1998, The Journal of cell biology.

[13]  R. Deschenes,et al.  Model organisms lead the way to protein palmitoyltransferases , 2004, Journal of Cell Science.

[14]  Carlos G. Dotti,et al.  Breaking the neuronal sphere: regulation of the actin cytoskeleton in neuritogenesis , 2002, Nature Reviews Neuroscience.

[15]  Dane M. Chetkovich,et al.  Dual Palmitoylation of Psd-95 Mediates Its Vesiculotubular Sorting, Postsynaptic Targeting, and Ion Channel Clustering , 2000, The Journal of cell biology.

[16]  M. Linder,et al.  G-protein Palmitoyltransferase Activity Is Enriched in Plasma Membranes (*) , 1996, The Journal of Biological Chemistry.

[17]  S. Hersch,et al.  HIP14, a novel ankyrin domain-containing protein, links huntingtin to intracellular trafficking and endocytosis. , 2002, Human molecular genetics.

[18]  R. Carraway,et al.  Huntingtin is a cytoplasmic protein associated with vesicles in human and rat brain neurons , 1995, Neuron.

[19]  D. Bredt,et al.  Protein palmitoylation: a regulator of neuronal development and function , 2002, Nature Reviews Neuroscience.

[20]  M. Resh.  Fatty acylation of proteins: new insights into membrane targeting of myristoylated and palmitoylated proteins. , 1999, Biochimica et biophysica acta.

[21]  C. Keller,et al.  The gamma2 subunit of GABA(A) receptors is a substrate for palmitoylation by GODZ. , 2004, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[22]  Neal Sweeney,et al.  Synaptic Strength Regulated by Palmitate Cycling on PSD-95 , 2002, Cell.

[23]  J. Skene,et al.  The 25 kDa synaptosomal-associated protein SNAP-25 is the major methionine-rich polypeptide in rapid axonal transport and a major substrate for palmitoylation in adult CNS , 1992, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[24]  M. Veit,et al.  Palmitoylation sites and processing of synaptotagmin I, the putative calcium sensor for neurosecretion , 2003, FEBS letters.

[25]  S. Moss,et al.  Palmitoylation regulates the clustering and cell surface stability of GABAA receptors , 2004, Molecular and Cellular Neuroscience.

[26]  M. Fishman,et al.  Neuronal pathfinding is abnormal in mice lacking the neuronal growth cone protein GAP-43 , 1995, Cell.

[27]  C. Schwarz,et al.  Wild-Type and Mutant Huntingtins Function in Vesicle Trafficking in the Secretory and Endocytic Pathways , 1998, Experimental Neurology.

[28]  D. Bredt,et al.  A combination of three distinct trafficking signals mediates axonal targeting and presynaptic clustering of GAD65 , 2002, The Journal of cell biology.

[29]  K. Mikoshiba,et al.  Mechanism of the SDS-resistant Synaptotagmin Clustering Mediated by the Cysteine Cluster at the Interface between the Transmembrane and Spacer Domains* , 2001, The Journal of Biological Chemistry.

[30]  T. Südhof.  Synaptotagmins: Why So Many?* , 2002, The Journal of Biological Chemistry.

[31]  J. Smotrys,et al.  Palmitoylation of intracellular signaling proteins: regulation and function. , 2004, Annual review of biochemistry.

[32]  K. Ueno.  Involvement of fatty acid synthase in axonal development in mouse embryos , 2000, Genes to cells : devoted to molecular & cellular mechanisms.