Translocation of IRS-1 to Cytosol Attenuates Insulin-Stimulated Glucose Transport without Affecting PI3-Kinase Activity~!2010-03-23~!2010-05-10~!2010-06-18~!

Junko Kawahara,Isao Usui,Atsuko Takano,Kazuyuki Tobe,Kazuyuki Hiratani,Yukiko Kanatani,Masashi Kobayashi,Tetsuro Haruta,Tatsuhito Uno,Minoru Iwata

doi:10.2174/1876524601003010014

Abstract

IRS-1 is a major substrate for insulin receptor tyrosine kinase. It is reported that intracellular translocation of serine-phosphorylated IRS-1 from low density microsome (LDM) fraction to cytosol attenuates its ability to transmit insulin signaling to the downstream molecules. In this study, we examined which insulin signal and action were affected by translocation of IRS-1 in 3T3-L1 adipocytes. Adenovirus-mediated overexpression of constitutively active PI3-kinase (p110CAAX) induces translocation of IRS-1 to cytosol without increasing IRS-1tyrosine phosphorylation. IRS-1 protein localized in cytosol fraction in p110CAAX-expressing cells maintained the ability to be tyrosine-phosphorylated by short term insulin treatment. Long term treatment with insulin for 4 to 8 h decreased tyrosine phosphorylation of IRS-1, PI3kinase activity, Akt phosphorylation and glucose uptake by second stimulation with insulin. Pretreatment with rapamycin, a specific mTOR inhibitor, increased the protein level of IRS-1 in LDM fraction and restored the attenuated insulin signaling and glucose uptake after long term insulin treatment. On the other hand, pretreatment with lactacystin, a specific proteasomal inhibitor, increased the protein level of serine-phosphorylated IRS-1 in cytosol fraction. In this condition, insulin signaling from IRS-1 to Akt was restored, but glucose uptake was not. Taken together, we conclude that localization of IRS-1 in LDM fraction is necessary for insulin-stimulated glucose uptake, while IRS-1, once serinephosphorylated and translocated to cytosol, fails to stimulate glucose uptake despite its intact ability to be tyrosine phosphorylated and to transmit insulin signaling to Akt level.

Highlights

Insulin receptor tyrosine kinase is activated in response to insulin stimulation
We previously reported that the activation of a pathway sensitive to rapamycin, an inhibiter of mammalian target of rapamycin, was involved in serine/threonine phosphorylation and intracellular translocation of Insulin receptor substrate (IRS)-1 from low-density microsome (LDM) to cytosol after prolonged exposure of 3T3-L1 adipocytes to insulin
To evaluate how mammalian target of rapamycin (mTOR)-dependent serine/threonine phosphorylation and translocation of IRS-1 after long term insulin exposure affects the early step of insulin signals, we first compared the effects of either rapamycin, which inhibits serine/threonine phosphorylation and translocation of IRS-1

Summary

Introduction

Insulin receptor tyrosine kinase is activated in response to insulin stimulation. Activation of phosphatidyl inositol (PI) 3kinase, recruited to the tyrosine-phosphorylated IRS-1, mediates various metabolic actions of insulin, such as translocation of GLUT4 followed by stimulation of glucose transport [1, 2]. Several recent studies have reported that serine/threonine phosphorylation of IRS-1 is highly associated to the impaired insulin signaling [3,4,5,6,7,8,9,10,11]. Some studies reported that serine phosphorylation of IRS-1 impairs insulin-stimulated tyrosine phosphorylation of IRS-1 [3,4,5,6,7,8]. Serine/threonine phosphorylation of IRS-1 is linked with altered intracellular localization of IRS-1, which interferes with the interaction of IRS-1 with insulin receptor, resulting in the decreased

Results

Discussion

Conclusion