53BP2S, interacting with insulin receptor substrates, modulates insulin signaling.
It is well known that insulin receptor substrates (IRS) act as a mediator for signal transduction of insulin, insulin-like growth factors, and several cytokines. To identify proteins that interact with IRS and modulate IRS-mediated signals, we performed yeast two-hybrid screening with IRS-1 as bait. Out of 109 cDNA-positive clones identified from a human placental cDNA library, two clones encoded 53BP2, p53-binding protein 2 (53BP2S), a short form splicing variant of the apoptosis-stimulating protein of p53 that possesses Src homology region 3 domain, and ankyrin repeats domain, and had been reported to interact with p53, Bcl-2, and NF-kappaB. Interaction of 53BP2S with IRS-1 was confirmed by glutathione S-transferase pull-down and co-immunoprecipitation assays in COS-7 cells and 3T3-L1 adipocytes. The Src homology region 3 domain and ankyrin repeats domain of 53BP2S were responsible for its interaction with IRS-1, whereas the phosphotyrosine binding domain and a central domain (amino acid residues 750-861) of IRS-1 were required for its interaction with 53BP2S. In CHO-C400 cells, expression of 53BP2S reduced insulin-stimulated IRS-1 tyrosine phosphorylation with a concomitant enhancement of IRS-2 tyrosine phosphorylation. In addition, the amount of the phosphatidylinositol 3-kinase regulatory p85 subunit associated with tyrosine-phosphorylated proteins, and activation of Akt was enhanced by 53BP2S expression. Although 53BP2S also enhanced Akt activation in 3T3-L1 adipocytes, insulin-induced glucose transporter 4 translocation was markedly inhibited in accordance with reduction of insulin-induced AS160 phosphorylation. Together these data demonstrate that 53BP2S interacts and modulates the insulin signals mediated by IRSs.
Hakuno
Fumihiko
F
Department of Animal Sciences, Graduate School of Agriculture and Life Sciences, the University of Tokyo, Tokyo, Japan.
Kurihara
Shigekazu
S
Watson
Robert T
RT
Pessin
Jeffrey E
JE
Takahashi
Shin-Ichiro
S
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D017398:Alternative Splicing; D000818:Animals; D017209:Apoptosis; D016466:CHO Cells; D019556:COS Cells; D002352:Carrier Proteins / Q000502:physiology*; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D007328:Insulin / Q000378:metabolism*; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D010750:Phosphoproteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D015398:Signal Transduction; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; C526292:IRS1 protein, human; C526297:IRS2 protein, human; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; C526295:Irs1 protein, mouse; C526298:Irs2 protein, mouse; D010750:Phosphoproteins; D051057:Proto-Oncogene Proteins c-akt