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Endothelial Cell Palmitoylproteomic Identifies Novel Lipid-Modified Targets and Potential Substrates for Protein Acyl Transferases

2012; Lippincott Williams & Wilkins; Volume: 110; Issue: 10 Linguagem: Inglês

10.1161/circresaha.112.269514

1524-4571

Ethan P. Marin, Behrad Derakhshan, TuKiet T. Lam, Alberto Dávalos, William C. Sessa,

Ubiquitin and proteasome pathways

Rationale: Protein S-palmitoylation is the posttranslational attachment of a saturated 16-carbon palmitic acid to a cysteine side chain via a thioester bond. Palmitoylation can affect protein localization, trafficking, stability, and function. The extent and roles of palmitoylation in endothelial cell (EC) biology is not well-understood, partly because of technological limits on palmitoylprotein detection. Objective: To develop a method using acyl-biotinyl exchange technology coupled with mass spectrometry to globally isolate and identify palmitoylproteins in ECs. Methods and Results: More than 150 putative palmitoyl proteins were identified in ECs using acyl-biotinyl exchange and mass spectrometry. Among the novel palmitoylproteins identified is superoxide dismutase-1, an intensively studied enzyme that protects all cells from oxidative damage. Mutation of cysteine-6 prevents palmitoylation, leads to reduction in superoxide dismutase-1 activity in vivo and in vitro, and inhibits nuclear localization, thereby supporting a functional role for superoxide dismutase-1 palmitoylation. Moreover, we used acyl-biotinyl exchange to search for substrates of particular protein acyl transferases in ECs. We found that palmitoylation of the cell adhesion protein platelet endothelial cell adhesion molecule-1 is dependent on the protein acyl transferase ZDHHC21. We show that knockdown of ZDHHC21 leads to reduced levels of platelet endothelial cell adhesion molecule-1 at the cell surface. Conclusions: Our data demonstrate the utility of EC palmitoylproteomics to reveal new insights into the role of this important posttranslational lipid modification in EC biology.