title: Constitutively active mitogen-activated protein kinase kinase increases GLUT1 expression and recruits both GLUT1 and GLUT4 at the cell surface in 3T3-L1 adipocytes.
abstract: To address a role of mitogen-activated protein kinase (MAPK) in the regulation of glucose transport, we made a constitutively active mutant of MAPK kinase (MAPKK) and introduced it into 3T3-L1 preadipocytes by using a retrovirus-mediated transfection procedure. The deletion of 20 amino acids (those between and including 32 and 51) in the amino terminal region of Xenopus MAPKK and the replacement of serine residues on the 218 and 222 positions by glutamic acid (dSESE-MAPKK) let Xenopus MAPKK constitutively active. The isolated cell clones differently expressing dSESE-MAPKK (clone 219 higher expression, clone 233 lower expression) efficiently differentiated to adipocytes by a standard differentiation cocktail. Accordingly, the increased expression of dSESE-MAPKK protein during differentiation resulted in the increased basal MAPK activity in clone 219 adipocytes and, to a lesser extent, in clone 233 adipocytes. In contrast to clone 233 and parental adipocytes, basal 2-deoxyglucose uptake was enhanced fourfold in clone 219 adipocytes, in accordance with increased expression of GLUT1 mRNA and protein. Whereas GLUT4 mRNA was similarly expressed in all of the adipocytes, GLUT4 protein appeared to decrease in clone 219 adipocytes. More importantly, subcellular fractionation studies showed that the localization of both GLUT1 and GLUT4 in the plasma membranes (PMs) was markedly increased in the basal state in clone 219 adipocytes compared with that in clone 233 and parental adipocytes, in which both glucose transporters were preferentially located in intracellular compartments. Consequently, insulin-induced translocation of GLUT1 was abolished in clone 219 adipocytes, although the remaining intracellular GLUT4 was still responsive to insulin stimulation, which led to the movement to the PM. As combined effects on the situation of GLUT1 and GLUT4, the foldness of insulin stimulation of glucose transport based on the basal activity was reduced in cells expressing constitutively active MAPKK. These results imply that chronic activation of MAPK could be one of the mechanisms for insulin resistance.
authors.affiliation: Department of Medicine and Clinical Science, Graduate School of Medicine, Kyoto University, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Yamamoto
authors.forename: Y
authors.initials: Y
authors.lastname: Yoshimasa
authors.forename: M
authors.initials: M
authors.lastname: Koh
authors.forename: J
authors.initials: J
authors.lastname: Suga
authors.forename: H
authors.initials: H
authors.lastname: Masuzaki
authors.forename: Y
authors.initials: Y
authors.lastname: Ogawa
authors.forename: K
authors.initials: K
authors.lastname: Hosoda
authors.forename: H
authors.initials: H
authors.lastname: Nishimura
authors.forename: Y
authors.initials: Y
authors.lastname: Watanabe
authors.forename: G
authors.initials: G
authors.lastname: Inoue
authors.forename: K
authors.initials: K
authors.lastname: Nakao
mesh_terms: D016475:3T3 Cells; D017667:Adipocytes / Q000166:cytology / Q000378:metabolism*; D019943:Amino Acid Substitution; D000818:Animals; D001692:Biological Transport; D002454:Cell Differentiation; D002462:Cell Membrane / Q000378:metabolism; D017353:Gene Deletion; D015870:Gene Expression; D005947:Glucose / Q000378:metabolism; D051272:Glucose Transporter Type 1; D051275:Glucose Transporter Type 4; D007425:Intracellular Membranes / Q000378:metabolism; D051379:Mice; D020929:Mitogen-Activated Protein Kinase Kinases / Q000235:genetics / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009124:Muscle Proteins; D014018:Tissue Distribution; D014981:Xenopus
chemical_list: D051272:Glucose Transporter Type 1; D051275:Glucose Transporter Type 4; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; C495678:Slc2a1 protein, mouse; C495693:Slc2a4 protein, mouse; D020928:Mitogen-Activated Protein Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; D005947:Glucose