Association of carboxyl esterase with facilitative glucose transporter isoform 4 (GLUT4) intracellular compartments in rat adipocytes and its possible role in insulin-induced GLUT4 recruitment

Facilitative glucose transporter isoform 4 (GLUT4) in rat adipocytes is lar gely sequestered in intracellular sites, and insulin recruits GLUT4 from th ese sites to the cell surface. The process is known to involve multiple int racellular compartments and associated proteins, many of which are yet to b e identified. Recently, we purified three distinct insulin-sensitive intrac ellular GLUT4 compartments (G4T(L), G4H, and G4L) in rat adipocytes and unr aveled several new resident proteins in these compartments. Here, we descri be one of them, a 62-kDa protein, purified and identified as rat adipose ti ssue carboxyl esterase (p62/CE) by matrix-assisted laser desorption/ionizat ion time of flight mass spectroscopy, reverse transcription-polymerase chai n reaction, gene cloning, and immunological and enzymatic activity measurem ents. p62/CE in rat adipocytes was 80% cytosolic and 20% microsome-associat ed. It was found in all of the three insulin-sensitive intracellular GLUT4 compartments, and particularly enriched in G4T(L), a compartment thought to represent GLUT4 endocytic vesicles. Significantly, an antibody against p62 /CE introduced into rat adipocytes completely abolished the insulin-induced GLUT4 recruitment to the plasma membrane in host cells without affecting t he basal GLUT4 distribution. Together, these findings suggest that p62/CE p lays a key role in insulin-induced GLUT4 recruitment in rat adipocytes, pro bably by hydrolyzing acylglycerols or acyl-CoA esters to the respective fre e acids that are required for GLUT4 transport vesicle budding and/or fusion .