Regulated assembly of vacuolar ATPase is increased during cluster disruption-induced maturation of dendritic cells through a phosphatidylinositol 3-Kinase/mTOR-dependent pathway

Abstract

The vacuolar (H+)-ATPases (V-ATPases) are ATP-driven proton pumps composed of a peripheral V1 domain and a membrane-embedded V0 domain. Regulated assembly of V1 and V0 represents an important regulatory mechanism for controlling V-ATPase activity in vivo. Previous work has shown that VATPase assembly increases during maturation of bone marrowderived dendritic cells induced by activation of Toll-like receptors. This increased assembly is essential for antigen processing, which is dependent upon an acidic lysosomal pH. Cluster disruption of dendritic cells induces a semi-mature phenotype associated with immune tolerance. Thus, semi-mature dendritic cells are able to process and present self-peptides to suppress autoimmune responses. We have investigated V-ATPase assembly in bone marrow-derived, murine dendritic cells and observed an increase in assembly following cluster disruption. This increased assembly is not dependent upon new protein synthesis and is associated with an increase in concanamycin A-sensitive proton transport in FITC-loaded lysosomes. Inhibition of phosphatidylinositol 3-kinase with wortmannin or mTORC1 with rapamycin effectively inhibits the increased assembly observed upon cluster disruption. These results suggest that the phosphatidylinositol 3-kinase/mTOR pathway is involved in controlling V-ATPase assembly during dendritic cell maturation. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.