Analysis of insulin signalling by RNAi-based gene silencing

被引:64
作者
Zhou, QL [1 ]
Park, JG [1 ]
Jiang, ZY [1 ]
Holik, JJ [1 ]
Mitra, P [1 ]
Semiz, S [1 ]
Guilherme, A [1 ]
Powelka, AM [1 ]
Tang, X [1 ]
Virbasius, J [1 ]
Czech, MP [1 ]
机构
[1] Univ Massachusetts, Sch Med, Program Mol Med, Worcester, MA 01605 USA
关键词
gene silencing; glucose; insulin signalling; phosphoinositide 3-kinase (PI3K); phospholipase C gamma; protein kinase C gamma/xi siRNA;
D O I
10.1042/BST0320817
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Using siRNA-mediated gene silencing in cultured adipocytes, we have dissected the insulin-signalling pathway leading to translocation of GLUT4 glucose transporters to the plasma membrane. RNAi (RNA interference)-based depletion of components in the putative TC10 pathway (CAP, CrkII and c-Cbl plus Cbl-b) or the phospholipase Cgamma pathway failed to diminish insulin signalling to GLUT4. Within the phosphoinositide 3-kinase pathway, loss of the 5'-phosphatidylinositol 3,4,5-trisphosphate phosphatase SHIP2 was also without effect, whereas depletion of the 3'-phosphatase PTEN significantly enhanced insulin action. Downstream of phosphatidylinositol 3,4,5-trisphosphate and PDK1, silencing the genes encoding the protein kinases Akt1/PKBalpha, or CISK(SGK3) or protein kinases Clambda/xi had little or no effect, but loss of Akt2/PKBbeta significantly attenuated GLUT4 regulation by insulin. These results show that Akt2/PKBbeta is the key downstream intermediate within the phosphoinositide 3-kinase pathway linked to insulin action on GLUM in cultured adipocytes, whereas PTEN is a potent negative regulator of this pathway.
引用
收藏
页码:817 / 821
页数:5
相关论文
共 35 条
[1]   Isoform-specific regulation of insulin-dependent glucose uptake by Akt/protein kinase B [J].
Bae, SS ;
Cho, H ;
Mu, J ;
Birnbaum, MJ .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2003, 278 (49) :49530-49536
[2]   Glucose transporter recycling in response to insulin is facilitated by myosin Myo1c [J].
Bose, A ;
Guilherme, A ;
Robida, SI ;
Nicoloro, SMC ;
Zhou, QL ;
Jiang, ZY ;
Pomerleau, DP ;
Czech, MP .
NATURE, 2002, 420 (6917) :821-824
[3]   Regulated transport of the glucose transporter glut4 [J].
Bryant, NJ ;
Govers, R ;
James, DE .
NATURE REVIEWS MOLECULAR CELL BIOLOGY, 2002, 3 (04) :267-277
[4]   PHOSPHATIDYLINOSITOL 3-KINASE ACTIVATION IS REQUIRED FOR INSULIN STIMULATION OF PP70 S6 KINASE, DNA-SYNTHESIS, AND GLUCOSE-TRANSPORTER TRANSLOCATION [J].
CHEATHAM, B ;
VLAHOS, CJ ;
CHEATHAM, L ;
WANG, L ;
BLENIS, J ;
KAHN, CR .
MOLECULAR AND CELLULAR BIOLOGY, 1994, 14 (07) :4902-4911
[5]   Insulin-stimulated GLUT4 translocation requires the CAP-dependent activation of TC10 [J].
Chiang, SH ;
Baumann, CA ;
Kanzaki, M ;
Thurmond, DC ;
Watson, RT ;
Neudauer, CL ;
Macara, IG ;
Pessin, JE ;
Saltiel, AR .
NATURE, 2001, 410 (6831) :944-948
[6]   Akt1/PKBα is required for normal growth but dispensable for maintenance of glucose homeostasis in mice [J].
Cho, H ;
Thorvaldsen, JL ;
Chu, QW ;
Feng, F ;
Birnbaum, MJ .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2001, 276 (42) :38349-38352
[7]   Insulin resistance and a diabetes mellitus-like syndrome in mice lacking the protein kinase Akt2 (PKBβ) [J].
Cho, H ;
Mu, J ;
Kim, JK ;
Thorvaldsen, JL ;
Chu, QW ;
Crenshaw, EB ;
Kaestner, KH ;
Bartolomei, MS ;
Shulman, GI ;
Birnbaum, MJ .
SCIENCE, 2001, 292 (5522) :1728-1731
[8]   The lipid phosphatase SHIP2 controls insulin sensitivity [J].
Clément, S ;
Krause, U ;
Desmedt, F ;
Tanti, JF ;
Behrends, J ;
Pesesse, X ;
Sasaki, T ;
Penninger, J ;
Doherty, M ;
Malaisse, W ;
Dumont, JE ;
Le Marchand-Brustel, Y ;
Erneux, C ;
Hue, L ;
Schurmans, S .
NATURE, 2001, 409 (6816) :92-97
[9]  
CZECH MP, 1993, J BIOL CHEM, V268, P9187
[10]  
CZECH MP, 1995, ANNU REV NUTR, V15, P441, DOI 10.1146/annurev.nu.15.070195.002301