scholarly journals Mechanism of feedback regulation of insulin receptor substrate-1 phosphorylation in primary adipocytes

2005 ◽  
Vol 388 (2) ◽  
pp. 713-720 ◽  
Author(s):  
Ingeborg HERS ◽  
Jeremy M. TAVARÉ
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Mammalian Target Of Rapamycin ◽  
Feedback Regulation ◽  
Serine Phosphorylation ◽  
Insulin Receptor Substrate ◽  
Insulin Receptor Substrate 1 ◽  
P70s6 Kinase ◽  
Mitogen Activated Protein ◽  
Phosphoinositide 3 Kinase

Serine and threonine phosphorylation of IRS-1 (insulin receptor substrate-1) has been reported to decrease its ability to be tyrosine-phosphorylated by the insulin receptor. Insulin itself may negatively regulate tyrosine phosphorylation of IRS-1 through a PI3K (phosphoinositide 3-kinase)-dependent feedback pathway. In the present study, we examined the regulation and role of IRS-1 serine phosphorylation in the modulation of IRS-1 tyrosine phosphorylation in physiologically relevant cells, namely freshly isolated primary adipocytes. We show that insulin-stimulated phosphorylation of Ser312 and Ser616 in IRS-1 was relatively slow, with maximal phosphorylation achieved after 20 and 5 min respectively. The effect of insulin on phosphorylation of both these sites required the activation of PI3K and the MAPKs (mitogen-activated protein kinases) ERK1/2 (extracellular-signal-regulated kinase 1 and 2), but not the activation of mTOR (mammalian target of rapamycin)/p70S6 kinase, JNK (c-Jun N-terminal kinase) or p38MAPK. Although inhibition of PI3K and ERK1/2 both substantially decreased insulin-stimulated phosphorylation of Ser312 and Ser616, only wortmannin enhanced insulin-stimulated tyrosine phosphorylation of IRS-1. Furthermore, inhibition of mTOR/p70S6 kinase, JNK or p38MAPK had no effect on insulin-stimulated IRS-1 tyrosine phosphorylation. The differential effect of inhibition of ERK1/2 on insulin-stimulated IRS-1 phosphorylation of Ser312/Ser616 and tyrosine indicates that these events are independent of each other and that phosphorylation of Ser312/Ser616 is not responsible for the negative regulation of IRS-1 tyrosine phosphorylation mediated by PI3K in primary adipocytes.

scholarly journals Mechanisms of inhibition of lipolysis by insulin, vanadate and peroxovanadate in rat adipocytes

1999 ◽  
Vol 339 (2) ◽  
pp. 281-289 ◽  
Author(s):  
Isabelle CASTAN ◽  
Jonny WIJKANDER ◽  
Vincent MANGANIELLO ◽  
Eva DEGERMAN
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Regulatory Subunit ◽  
Insulin Receptor Substrate ◽  
Insulin Receptor Substrate 1 ◽  
Rat Adipocytes ◽  
Lipolytic Action ◽  
Dependent Protein ◽  
Phosphodiesterase Type ◽  
Phosphoinositide 3 Kinase

Vanadate and peroxovanadate (pV), potent inhibitors of tyrosine phosphatases, mimic several of the metabolic actions of insulin. Here we compare the mechanisms for the anti-lipolytic action of insulin, vanadate and pV in rat adipocytes. Vanadate (5 mM) and pV (0.01 mM) inhibited lipolysis induced by 0.01–1 µM isoprenaline, vanadate being more and pV less efficient than insulin (1 nM). A loss of anti-lipolytic effect of pV was observed by increasing the concentration of isoprenaline and/or pV. pV induced tyrosine phosphorylation of the insulin receptor and insulin receptor substrate-1 to a greater extent than insulin, whereas vanadate affected these components little if at all. In addition, only a higher concentration (0.1 mM) of pV induced the tyrosine phosphorylation of p85, the 85 kDa regulatory subunit of phosphoinositide 3-kinase (PI-3K). Vanadate activated PI-3K-independent (in the presence of 10 nM isoprenaline) and PI-3K-dependent (in the presence of 100 nM isoprenaline) anti-lipolytic pathways, both of which were found to be independent of phosphodiesterase type 3B (PDE3B). pV (0.01 mM), like insulin, activated PI-3K- and PDE3B-dependent pathways. However, the anti-lipolytic pathway of 0.1 mM pV did not seem to require insulin receptor substrate-1-associated PI-3K and was found to be partly independent of PDE3B. Vanadate and pV (only at 0.01 mM), like insulin, decreased the isoprenaline-induced activation of cAMP-dependent protein kinase. Overall, these results underline the complexity and the diversity in the mechanisms that regulate lipolysis.

scholarly journals YMXM motifs and signaling by an insulin receptor substrate 1 molecule without tyrosine phosphorylation sites.

1996 ◽  
Vol 16 (8) ◽  
pp. 4147-4155 ◽  
Author(s):  
M G Myers ◽  
Y Zhang ◽  
G A Aldaz ◽  
T Grammer ◽  
E M Glasheen ◽  
...  
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Binding Sites ◽  
Insulin Receptor Substrate ◽  
Phosphorylation Sites ◽  
Mitogenic Response ◽  
Insulin Receptor Substrate 1 ◽  
Mitogen Activated Protein ◽  
High Insulin ◽  
P70 S6k

Tyrosine phosphorylation of insulin receptor substrate 1 (IRS-1) by the activated receptors for insulin, IGF-1, and various cytokines creates binding sites for signaling proteins with Src homology 2 domains (SH2 proteins). Determining the role of specific SH2 proteins during insulin signaling has been difficult because IRS-1 possesses as many as 18 potential tyrosine phosphorylation sites, several of which contain redundant motifs. Using 32D cells, which contain no endogenous IRS proteins, we compared the signaling ability of an IRS-1 molecule in which 18 potential tyrosine phosphorylation sites were replaced by phenylalanine (IRS-1(F18)) with two derivative molecules which retained three YMXM motifs (IRS-1(3YMXM)) or the two COOH-terminal SHP2-Fyn binding sites (IRS-1(YCT)). During insulin stimulation, IRS-1(F18) failed to undergo tyrosine phosphorylation or mediate activation of the phosphotidylinositol (PI) 3'-kinase or p70(s6k); IRS-1(YCT) was tyrosine phosphorylated but also failed to mediate these signaling events. Neither IRS-1(3YMXM) nor IRS-1(YCT) mediated activation of mitogen-activated protein kinases. IRS-1(F18) and IRS-1(YCT) partially mediated similar levels of insulin-stimulated mitogenesis at high insulin concentrations, however, suggesting that IRS-1 contains phosphotyrosine-independent elements which effect mitogenic signals, and that the sites in IRS-l(YCT) do not augment this signal. IRS-1(3YMXM) mediated the maximal mitogenic response to insulin, although the response to insulin was more sensitive with wild-type IRS-1. By contrast, the association of IRS-1(3YMXM) with PI 3'-kinase was more sensitive to insulin than the association with IRS-1. Thus, the binding of SH2 proteins (such as PI 3'-kinase) by YMXM motifs in IRS-1 is an important element in the mitogenic response, but other elements are essential for full mitogenic sensitivity.

scholarly journals Regulation of insulin receptor substrate-1 by mTORC2 (mammalian target of rapamycin complex 2)

2013 ◽  
Vol 41 (4) ◽  
pp. 896-901 ◽  
Author(s):  
Michael A. DeStefano ◽  
Estela Jacinto
Keyword(s):  
Growth Factors ◽  
Insulin Receptor ◽  
Insulin Signalling ◽  
Mammalian Target Of Rapamycin ◽  
Serine Phosphorylation ◽  
Target Of Rapamycin ◽  
Insulin Receptor Substrate ◽  
Insulin Receptor Substrate 1 ◽  
Cullin 7 ◽  
Mtor Complex 1

mTOR (mammalian target of rapamycin) responds to the presence of nutrients, energy and growth factors to link cellular metabolism, growth and proliferation. The rapamycin-sensitive mTORC (mTOR complex) 1 activates the translational regulator S6K (S6 kinase), leading to increased protein synthesis in the presence of nutrients. On the other hand, the rapamycin-insensitive mTORC2 responds to the presence of growth factors such as insulin by phosphorylating Akt to promote its maturation and allosteric activation. We recently found that mTORC2 can also regulate insulin signalling at the level of IRS-1 (insulin receptor substrate-1). Whereas mTORC1 promotes IRS-1 serine phosphorylation that is linked to IRS-1 down-regulation, we uncovered that mTORC2 mediates its degradation. In mTORC2-disrupted cells, inactive IRS-1 accumulated despite undergoing phosphorylation at the mTORC1-mediated serine sites. Defective IRS-1 degradation was due to attenuated expression of the CUL7 (Cullin 7) ubiquitin ligase substrate-targeting sub-unit Fbw8. mTORC2 and Fbw8 co-localize at the membrane where mTORC2 phosphorylates Ser86 to stabilize Fbw8 and promotes its cytosolic localization upon insulin stimulation. Under conditions of chronic insulin exposure, inactive serine-phosphorylated IRS-1 and Fbw8 co-localize to the cytosol where the former becomes ubiquitylated via CUL7/Fbw8. Thus mTORC2 negatively feeds back to IRS-1 via control of Fbw8 stability and localization. Our findings reveal that, in addition to persistent mTORC1 signalling, increased mTORC2 signals can promote insulin resistance due to mTORC2-mediated degradation of IRS-1.

scholarly journals Serine Phosphorylation Proximal to Its Phosphotyrosine Binding Domain Inhibits Insulin Receptor Substrate 1 Function and Promotes Insulin Resistance

2004 ◽  
Vol 24 (21) ◽  
pp. 9668-9681 ◽  
Author(s):  
Yan-Fang Liu ◽  
Avia Herschkovitz ◽  
Sigalit Boura-Halfon ◽  
Denise Ronen ◽  
Keren Paz ◽  
...  
Keyword(s):  
Insulin Resistance ◽  
Insulin Receptor ◽  
Insulin Signaling ◽  
Insulin Treatment ◽  
Binding Domain ◽  
Serine Phosphorylation ◽  
Insulin Receptor Substrate ◽  
Control Mechanisms ◽  
Insulin Receptor Substrate 1 ◽  
Phosphotyrosine Binding Domain

ABSTRACT Ser/Thr phosphorylation of insulin receptor substrate (IRS) proteins negatively modulates insulin signaling. Therefore, the identification of serine sites whose phosphorylation inhibit IRS protein functions is of physiological importance. Here we mutated seven Ser sites located proximal to the phosphotyrosine binding domain of insulin receptor substrate 1 (IRS-1) (S265, S302, S325, S336, S358, S407, and S408) into Ala. When overexpressed in rat hepatoma Fao or CHO cells, the mutated IRS-1 protein in which the seven Ser sites were mutated to Ala (IRS-17A), unlike wild-type IRS-1 (IRS-1WT), maintained its Tyr-phosphorylated active conformation after prolonged insulin treatment or when the cells were challenged with inducers of insulin resistance prior to acute insulin treatment. This was due to the ability of IRS-17A to remain complexed with the insulin receptor (IR), unlike IRS-1WT, which underwent Ser phosphorylation, resulting in its dissociation from IR. Studies of truncated forms of IRS-1 revealed that the region between amino acids 365 to 430 is a main insulin-stimulated Ser phosphorylation domain. Indeed, IRS-1 mutated only at S408, which undergoes phosphorylation in vivo, partially maintained the properties of IRS-17A and conferred protection against selected inducers of insulin resistance. These findings suggest that S408 and additional Ser sites among the seven mutated Ser sites are targets for IRS-1 kinases that play a key negative regulatory role in IRS-1 function and insulin action. These sites presumably serve as points of convergence, where physiological feedback control mechanisms, which are triggered by insulin-stimulated IRS kinases, overlap with IRS kinases triggered by inducers of insulin resistance to terminate insulin signaling.

scholarly journals Insulin receptor substrate 1 rescues insulin action in CHO cells expressing mutant insulin receptors that lack a juxtamembrane NPXY motif.

1995 ◽  
Vol 15 (9) ◽  
pp. 4711-4717 ◽  
Author(s):  
D Chen ◽  
D J Van Horn ◽  
M F White ◽  
J M Backer
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Cho Cells ◽  
Glycogen Synthesis ◽  
Insulin Receptors ◽  
Insulin Receptor Substrate ◽  
Insulin Stimulation ◽  
Insulin Receptor Substrate 1 ◽  
Npxy Motif ◽  
Stimulation Of

Insulin signals are mediated through tyrosine phosphorylation of specific proteins such as insulin receptor substrate 1 (IRS-1) and Shc by the activated insulin receptor (IR). Phosphorylation of both proteins is nearly abolished by an alanine substitution at Tyr-960 (A960) in the beta-subunit of the receptor. However, overexpression of IRS-1 in CHO cells expressing the mutant receptor (A960 cells) restored sufficient tyrosine phosphorylation of IRS-1 to rescue IRS-1/Grb-2 binding and phosphatidylinositol 3' kinase activation during insulin stimulation. Shc tyrosine phosphorylation and its binding to Grb-2 were impaired in the A960 cells and were unaffected by overexpression of IRS-1. Although overexpression of IRS-1 increased IRS-1 binding to Grb-2, ERK-1/ERK-2 activation was not rescued. These data suggest that signaling molecules other than IRS-1, perhaps including Shc, are critical for insulin stimulation of p21ras. Interestingly, overexpression of IRS-1 in the A960 cells restored insulin-stimulated mitogenesis and partially restored insulin stimulation of glycogen synthesis. Thus, IRS-1 tyrosine phosphorylation is sufficient to increase the mitogenic response to insulin, whereas insulin stimulation of glycogen synthesis appears to involve other factors. Moreover, IRS-1 phosphorylation is either not sufficient or not involved in insulin stimulation of ERK.

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