Palmitoylation is a post-translational modification of Alix regulating the membrane organization of exosome-like small extracellular vesicles

• Published in: Biochimica et biophysica acta. General subjects 2018-12, Vol.1862 (12), p.2879-2887
• Main Authors: Romancino, Daniele P., Buffa, Valentina, Caruso, Stefano, Ferrara, Ines, Raccosta, Samuele, Notaro, Antonietta, Campos, Yvan, Noto, Rosina, Martorana, Vincenzo, Cupane, Antonio, Giallongo, Agata, d'Azzo, Alessandra, Manno, Mauro, Bongiovanni, Antonella
• Format: Article
• Language: English
• Subjects: Alix (also known as PDCD6IP); Calcium-Binding Proteins - metabolism; Cell Cycle Proteins - metabolism; Cell Line; Cell Membrane - metabolism; Cells, Cultured; Endosomal Sorting Complexes Required for Transport - metabolism; Exosome; Exosomes - metabolism; extracellular vesicles (EVs); Extracellular Vesicles - metabolism; Humans; Lipid Bilayers; Lipoylation; Muscle, Skeletal - cytology; Muscle, Skeletal - metabolism; Protein Binding; Protein Conformation; Protein Processing, Post-Translational; Protein Transport; S-palmitoylation; Signal Transduction; Skeletal muscle cells; Tetraspanin; Tetraspanin 29 - metabolism
• ISSN: 0304-4165; 1872-8006
• DOI: 10.1016/j.bbagen.2018.09.004

Virtually all cell types have the capacity to secrete nanometer-sized extracellular vesicles, which have emerged in recent years as potent signal transducers and cell-cell communicators. The multifunctional protein Alix is a bona fide exosomal regulator and skeletal muscle cells can release Alix-positive nano-sized extracellular vesicles, offering a new paradigm for understanding how myofibers communicate within skeletal muscle and with other organs. S-palmitoylation is a reversible lipid post-translational modification, involved in different biological processes, such as the trafficking of membrane proteins, achievement of stable protein conformations, and stabilization of protein interactions. Here, we have used an integrated biochemical-biophysical approach to determine whether S-palmitoylation contributes to the regulation of extracellular vesicle production in skeletal muscle cells. We ascertained that Alix is S-palmitoylated and that this post-translational modification influences its protein-protein interaction with CD9, a member of the tetraspanin protein family. Furthermore, we showed that the structural organization of the lipid bilayer of the small (nano-sized) extracellular vesicle membrane with altered palmitoylation is qualitatively different compared to mock control vesicles. We propose that S-palmitoylation regulates the function of Alix in facilitating the interactions among extracellular vesicle-specific regulators and maintains the proper structural organization of exosome-like extracellular vesicle membranes. Beyond its biological relevance, our study also provides the means for a comprehensive structural characterization of EVs. [Display omitted] •Role of a post-translational modification on exosome-like extracellular vesicles•Palmitoylation maintains the lipid bilayer organization of extracellular vesicle.•Palmitoylation is a post-translational modification of Alix.•Palmitoylation-dependent interaction between Alix and CD9