Curation of the Mammalian Palmitoylome Indicates a Pivotal Role for Palmitoylation in Diseases and Disorders of the Nervous System and Cancers

Abstract

Palmitoylation involves the reversible posttranslational addition of palmitate to cysteines and promotes membrane binding and subcellular localization. Recent advancements in the detection and identification of palmitoylated proteins have led to multiple palmitoylation proteomics studies but these datasets are contained within large supplemental tables, making downstream analysis and data mining time-consuming and difficult. Consequently, we curated the data from 15 palmitoylation proteomics studies into one compendium containing 1,838 genes encoding palmitoylated proteins; representing approximately 10% of the genome. Enrichment analysis revealed highly significant enrichments for Gene Ontology biological processes, pathway maps, and process networks related to the nervous system. Strikingly, 41% of synaptic genes encode a palmitoylated protein in the compendium. The top disease associations included cancers and diseases and disorders of the nervous system, with Schizophrenia, HD, and pancreatic ductal carcinoma among the top five, suggesting that aberrant palmitoylation may play a pivotal role in the balance of cell death and survival. This compendium provides a much-needed resource for cell biologists and the palmitoylation field, providing new perspectives for cancer and neurodegeneration.

Fig 1. Palmitoylation and detection methods.

(a) Palmitoylation involves the reversible addition of long-chain fatty acids (FA) to cysteine residues via thioester bonds. (b) The ABE and Acyl-RAC assays use N-ethylmaleimide (NEM) to block free cysteines and hydroxylamine (HAM) to remove palmitate. The Acyl-RAC assay uses thiopropyl-sepharose beads that covalently react with the free cysteines, allowing enrichment and elution, using β-mercaptoethanol (β-ME), of palmitoyl-proteins and detection by MS. Following HAM treatment in the ABE assay free cysteines are labeled using Biotin-HPDP, allowing streptavidin-sepharose enrichment for MS. (c) The bioorthogonal-labeling assay uses alkyne-FA analogues followed by click chemistry to covalently link alkynyl-palmitate with biotin, allowing enrichment of palmitoyl-proteins for MS.

Fig 2. Hierarchical clustering of 15 published palmitoylomes.

The gene list of each palmitoyl proteome were subjected to hierarchical clustering using R. Studies that used ABE, bioorthogonal labeling (CLICK) or Acyl-RAC (ARAC) assays are in black, blue, and green font, respectively. Studies that used neuronal sample sources are outlined with black boxes. Bootstrap values are only shown for significant clusters.

Fig 3. Gene Ontology biological process enrichments among the curated palmitoylome.

The top 15 GO biological process enrichments that obtained a FDR<0.001 and a FE≥2 plotted by-log (FDR). The FE is displayed to the right of each bar.

Fig 4. Pathway map enrichments of the curated palmitoylome and enrichment of synaptic proteins for palmitoylated proteins.

(a) The 14 pathway map enrichments that obtained a FDR < 0.001 and FE ≥ 2 plotted by-log (FDR). The FE is displayed to the right of each bar. Venn diagram of genes in the compendium and the synaptic gene list from SynSysNet is shown in (b). The overlap between proteins from neuronal sources versus non-neuronal sources is shown in a Venn diagram in (c).

Fig 5. Biomarker-based disease-association enrichments of the curated palmitoylome.

(a) The top 15 biomarker-based disease-association enrichments that obtained a FDR < 0.001 and a FE ≥ 2 plotted by–log (FDR). The FE is displayed to the right of each bar. All of the 40 statistically significant biomarker-based disease-association enrichments of the compendium were broadly classified and are displayed in (b). GI = Gastrointestinal. Other = those diseases that do not fit into any other category.