Essential role of insulin receptor substrate 1 in differentiation of brown adipocytes.
The most widely distributed members of the family of insulin receptor substrate (IRS) proteins are IRS-1 and IRS-2. These proteins participate in insulin and insulin-like growth factor 1 signaling, as well as the actions of some cytokines, growth hormone, and prolactin. To more precisely define the specific role of IRS-1 in adipocyte biology, we established brown adipocyte cell lines from wild-type and IRS-1 knockout (KO) animals. Using differentiation protocols, both with and without insulin, preadipocyte cell lines derived from IRS-1 KO mice exhibited a marked decrease in differentiation and lipid accumulation (10 to 40%) compared to wild-type cells (90 to 100%). Furthermore, IRS-1 KO cells showed decreased expression of adipogenic marker proteins, such as peroxisome proliferator-activated receptor gamma (PPARgamma), CCAAT/enhancer-binding protein alpha (C/EBPalpha), fatty acid synthase, uncoupling protein-1, and glucose transporter 4. The differentiation deficit in the KO cells could be reversed almost completely by retrovirus-mediated reexpression of IRS-1, PPARgamma, or C/EBPalpha but not the thiazolidinedione troglitazone. Phosphatidylinositol 3-kinase (PI 3-kinase) assays performed at various stages of the differentiation process revealed a strong and transient activation in IRS-1, IRS-2, and phosphotyrosine-associated PI 3-kinase in the wild-type cells, whereas the IRS-1 KO cells showed impaired phosphotyrosine-associated PI 3-kinase activation, all of which was associated with IRS-2. Akt phosphorylation was reduced in parallel with the total PI 3-kinase activity. Inhibition of PI 3-kinase with LY294002 blocked differentiation of wild-type cells. Thus, IRS-1 appears to be an important mediator of brown adipocyte maturation. Furthermore, this signaling molecule appears to exert its unique role in the differentiation process via activation of PI 3-kinase and its downstream target, Akt, and is upstream of the effects of PPARgamma and C/EBPalpha.
Fasshauer
M
M
Research Division, Joslin Diabetes Center, and Department of Medicine, Harvard Medical School, Boston, Massachusetts 02215, USA.
Klein
J
J
Kriauciunas
K M
KM
Ueki
K
K
Benito
M
M
Kahn
C R
CR
D017667:Adipocytes / Q000166:cytology* / Q000201:enzymology / Q000378:metabolism; D002001:Adipose Tissue, Brown / Q000166:cytology* / Q000201:enzymology / Q000378:metabolism; D000818:Animals; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D004789:Enzyme Activation; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D018345:Mice, Knockout; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010750:Phosphoproteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases; D021122:Protein Subunits; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D011972:Receptor, Insulin / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D015398:Signal Transduction; D013234:Stem Cells / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection
D022763:CCAAT-Enhancer-Binding Protein-alpha; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; D020928:Mitogen-Activated Protein Kinases; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins; D019000:Phosphotyrosine; D017346:Protein Serine-Threonine Kinases; D021122:Protein Subunits; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D011972:Receptor, Insulin; D018160:Receptors, Cytoplasmic and Nuclear; D014157:Transcription Factors; C526295:Irs1 protein, mouse; C526298:Irs2 protein, mouse