Requirement for 3-phosphoinositide-kependent dinase-1 (PDK-1) in insulin-induced glucose uptake in immortalized brown adipocytes.
To provide insight into the physiological importance of 3-phosphoinositide-dependent kinase-1 (PDK-1) in the metabolic actions of insulin, we have generated mice that harbor a PDK-1 gene containing LoxP sites (PDK-1(lox/lox) mice) and established immortalized brown preadipocyte cell lines both from these animals and from wild-type mice. Exposure to appropriate hormonal inducers resulted in the differentiation of >80% of the immortalized brown preadipocytes derived from both types of mice into mature adipocytes. Introduction of the Cre recombinase with the use of adenovirus-mediated gene transfer induced a dose-dependent decrease in the abundance of PDK-1 in PDK-1(lox/lox) adipocytes but not in the wild-type cells. In Cre-expressing PDK-1(lox/lox) adipocytes in which the abundance of PDK-1 was reduced by approximately 85%, the insulin-induced phosphorylation both of Akt on threonine 308 and of p70 S6 kinase on threonine-389 was markedly inhibited. The phosphorylation both of Akt on serine 473 and of p42 and p44 isoforms of mitogen-activated protein kinase induced by insulin was not affected by Cre expression, indicating that the latter specifically inhibits PDK-1-dependent signaling. Both glucose uptake and the translocation of glucose transporter 4 to the plasma membrane induced by insulin as well as glucose uptake induced by a constitutively active form of phosphoinositide 3-kinase were also greatly inhibited by Cre expression in PDK-1(lox/lox) adipocytes. Phosphorylation of AMP-activated protein kinase and glucose uptake induced by 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) were not affected by Cre expression in PDK-1(lox/lox) adipocytes. These results indicate that PDK-1 is essential for insulin-induced glucose uptake in adipocytes.
Sakaue
Hiroshi
H
Division of Diabetes and Digestive and Kidney Diseases, Department of Clinical Molecular Medicine, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Chuo-ku, Kobe 650-0017, Japan.
Nishizawa
Akihiko
A
Ogawa
Wataru
W
Teshigawara
Kiyoshi
K
Mori
Toshiyuki
T
Takashima
Yasuhiro
Y
Noda
Tetsuo
T
Kasuga
Masato
M
D064413:3-Phosphoinositide-Dependent Protein Kinases; D000256:Adenoviridae / Q000235:genetics; D017667:Adipocytes / Q000378:metabolism; D002001:Adipose Tissue, Brown / Q000378:metabolism*; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport; D002454:Cell Differentiation; D002460:Cell Line; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D017353:Gene Deletion; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D005947:Glucose / Q000378:metabolism / Q000493:pharmacokinetics*; D051275:Glucose Transporter Type 4; D006579:Heterozygote; D007150:Immunohistochemistry; D007328:Insulin / Q000378:metabolism; D019426:Integrases / Q000378:metabolism; D020935:MAP Kinase Signaling System; D051379:Mice; D008856:Microscopy, Fluorescence; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D009124:Muscle Proteins; D010766:Phosphorylation; D020033:Protein Isoforms; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism* / Q000502:physiology*; D021381:Protein Transport; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D012265:Ribonucleotides / Q000494:pharmacology; D012694:Serine / Q000737:chemistry; D015398:Signal Transduction; D013912:Threonine / Q000737:chemistry; D014764:Viral Proteins / Q000378:metabolism
D051275:Glucose Transporter Type 4; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins; D012265:Ribonucleotides; C495693:Slc2a4 protein, mouse; D014764:Viral Proteins; D013912:Threonine; D000620:Aminoimidazole Carboxamide; D012694:Serine; D064413:3-Phosphoinositide-Dependent Protein Kinases; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; C045073:Cre recombinase; D019426:Integrases; C031143:AICA ribonucleotide; D005947:Glucose