Protein Tyrosine Phosphatase-1B Dephosphorylation of the Insulin Receptor Occurs in a Perinuclear Endosome Compartment in Human Embryonic Kidney 293 Cells

Protein tyrosine phosphatase-1B (PTP-1B) is a negative regulator of insulin signaling. It is thought to carry out this role by interacting with and dephosphorylating the activated insulin receptor (IR). However, little is known regarding the nature of the cellular interaction between these proteins, especially because the IR is localized to the plasma membrane and PTP-1B to the endoplasmic reticulum. Using confocal microscopy and fluorescence resonance energy transfer (FRET), the interaction between PTP-1B and the IR was examined in co-transfected human embryonic kidney 293 cells. Biological activities were not significantly affected for either PTP-1B or the IR with the fusion of W1B-green fluorescent protein (GFP) to the N terminus of PTP-1B (W1B-PTP-1B) or the fusion of Topaz-GFP to the C terminus of the IR (Topaz-IR). FRET between W1B and Topaz was monitored in cells transfected with either wild type PTP-1B (W1B-PTP-1B) or the substrate-trapping form PTP-1BD181A (W1B-PTP-1BD181A) and Topaz-IR. Co-expression of W1B-PTP-1B with Topaz-IR resulted in distribution of Topaz-IR to the plasma membrane, but no FRET was obtained upon insulin treatment. In contrast, co-expression of W1B-PTP-1BD181A with Topaz-IR caused an increase in cytosolic Topaz-IR fluorescence and, in some cells, a significant basal FRET signal, suggesting that PTP-1B is interacting with the IR during its synthesis. Stimulation of these cells with insulin resulted in a rapid induction of FRET that increased over time and was localized to a perinuclear spot. Co-expression of Topaz-IR with a GFP-labeled RhoB endosomal marker and treatment of the cells with insulin identified a perinuclear endosome compartment as the site of localization. Furthermore, the insulin-induced FRET could be prevented by the treatment of the cells with a specific PTP-1B inhibitor. These results suggest that PTP-1B appears not only to interact with and dephosphorylate the insulin-stimulated IR in a perinuclear endosome compartment but is also involved in maintaining the IR in a dephosphorylated state during its biosynthesis. Protein tyrosine phosphatase-1B (PTP-1B) is a negative regulator of insulin signaling. It is thought to carry out this role by interacting with and dephosphorylating the activated insulin receptor (IR). However, little is known regarding the nature of the cellular interaction between these proteins, especially because the IR is localized to the plasma membrane and PTP-1B to the endoplasmic reticulum. Using confocal microscopy and fluorescence resonance energy transfer (FRET), the interaction between PTP-1B and the IR was examined in co-transfected human embryonic kidney 293 cells. Biological activities were not significantly affected for either PTP-1B or the IR with the fusion of W1B-green fluorescent protein (GFP) to the N terminus of PTP-1B (W1B-PTP-1B) or the fusion of Topaz-GFP to the C terminus of the IR (Topaz-IR). FRET between W1B and Topaz was monitored in cells transfected with either wild type PTP-1B (W1B-PTP-1B) or the substrate-trapping form PTP-1BD181A (W1B-PTP-1BD181A) and Topaz-IR. Co-expression of W1B-PTP-1B with Topaz-IR resulted in distribution of Topaz-IR to the plasma membrane, but no FRET was obtained upon insulin treatment. In contrast, co-expression of W1B-PTP-1BD181A with Topaz-IR caused an increase in cytosolic Topaz-IR fluorescence and, in some cells, a significant basal FRET signal, suggesting that PTP-1B is interacting with the IR during its synthesis. Stimulation of these cells with insulin resulted in a rapid induction of FRET that increased over time and was localized to a perinuclear spot. Co-expression of Topaz-IR with a GFP-labeled RhoB endosomal marker and treatment of the cells with insulin identified a perinuclear endosome compartment as the site of localization. Furthermore, the insulin-induced FRET could be prevented by the treatment of the cells with a specific PTP-1B inhibitor. These results suggest that PTP-1B appears not only to interact with and dephosphorylate the insulin-stimulated IR in a perinuclear endosome compartment but is also involved in maintaining the IR in a dephosphorylated state during its biosynthesis. The insulin receptor is a heterotetramer plasma membrane protein that consists of two extracellular α subunits and two membrane-spanning intracellular β subunits that contain inherent tyrosine kinase domains (1Saltiel A.R. Kahn C.R. Nature. 2001; 414: 799-806Crossref PubMed Scopus (3973) Google Scholar). The receptor is initially synthesized as a single polypeptide α-β proreceptor that undergoes processing in the endoplasmic reticulum (ER). 1The abbreviations used are: ER, endoplasmic reticulum; BRET, bioluminescence resonance energy transfer; EBNA, Epstein-Barr virus nuclear antigen; EGFR, epidermal growth factor receptor; ECFP, enhanced cyan fluorescent protein; EYFP, enhanced yellow fluorescent protein; FRET, fluorescence resonance energy transfer; GFP, green fluorescent protein; HEK, human embryonic kidney; IR, insulin receptor; PBS, phosphate-buffered saline; PDGFR, platelet-derived growth factor receptor; PTP-1B, protein tyrosine phosphatse-1B. While it is in the ER, the proreceptor dimerizes and undergoes disulfide bond formation, glycosylation, and proteolysis before transfer to the plasma membrane (2Olson T.S. Bamberger M.J. Lane M.D. J. Biol. Chem. 1987; 263: 7342-7351Abstract Full Text PDF Google Scholar, 3Lu K. Guidotti G. Mol. Biol. Cell. 1996; 7: 679-691Crossref PubMed Scopus (23) Google Scholar, 4Bass J. Chiu G. Argon Y. Steiner D.F. J. Cell Biol. 1998; 141: 637-646Crossref PubMed Scopus (95) Google Scholar). Binding of insulin to the receptor on the plasma membrane results in activation of the β-subunit tyrosine kinase activity, and, once activated, autophosphorylation on three critical tyrosine residues in the kinase activation loop occurs, resulting in full activation of the tyrosine kinase activity (1Saltiel A.R. Kahn C.R. Nature. 2001; 414: 799-806Crossref PubMed Scopus (3973) Google Scholar). The activated IR then begins to phosphorylate its various substrates to initiate the insulin signaling cascade but, at the same time, undergoes internalization into endosomes, where it remains activated (5Di Guglielmo G.M. Drake P.G. Baass P.C. Authier F. Posner B.I. Bergeron J.M. Mol. Cell. Biochem. 1998; 182: 59-63Crossref PubMed Scopus (111) Google Scholar, 6Bevan A.P. Seabright P.J. Tikerpae J. Posner B.I. Smith G.D. Siddle K. Mol. Cell. Endocrinol. 2000; 164: 145-157Crossref PubMed Scopus (8) Google Scholar). However, acidification of the endosome lumen causes insulin dissociation from the receptor, which prevents further activation. The receptor then undergoes dephosphorylation, at which time it is then sorted back to the plasma membrane (5Di Guglielmo G.M. Drake P.G. Baass P.C. Authier F. Posner B.I. Bergeron J.M. Mol. Cell. Biochem. 1998; 182: 59-63Crossref PubMed Scopus (111) Google Scholar). Identification of the protein tyrosine phosphatases (PTPs) involved in the dephosphorylation of the activated IR is of critical interest, because inhibitors directed toward these PTPs could be used in the treatment of insulin resistance and Type 2 diabetes. Using a mouse gene knockout, PTP-1B was shown to be a crucial phosphatase involved in the in vivo dephosphorylation of the activated IR (7Elchebly M. Payette P. Michaliszyn E. Cromlish W. Collins L. Loy A.L. Normandin D. Cheng A. Himms-Hagen J. Ramachandran C. Gresser M.J. Tremblay M.L. Kennedy B.P. Science. 1999; 283: 1544-1548Crossref PubMed Scopus (1926) Google Scholar, 8Klaman L.D. Boss O. Peroni O.D. Kim J.K. Martino J.L. Zabolotny J.M. Moghal N. Lubkin M. Kim Y.B. Sharpe A.H. Stricker-Krongrad A. Shulman G.I. Neel B.G. Kahn B.B. Mol. Cell. Biol. 2000; 20: 5479-5489Crossref PubMed Scopus (1130) Google Scholar). The results from the PTP-1B null mouse were key in establishing the role of PTP-1B in the negative regulation of insulin signaling (9Asante-Appiah E. Kennedy B.P. Am. J. Physiol. 2003; 284: E663-E670Crossref PubMed Scopus (168) Google Scholar). In addition to having enhanced insulin sensitivity, these mice were also resistant to diet-induced obesity. Moreover, insulin-induced tyrosine phosphorylation of the receptor was increased and prolonged in muscle and liver of these animals, suggesting that the role of PTP-1B is to dephosphorylate the activated IR (7Elchebly M. Payette P. Michaliszyn E. Cromlish W. Collins L. Loy A.L. Normandin D. Cheng A. Himms-Hagen J. Ramachandran C. Gresser M.J. Tremblay M.L. Kennedy B.P. Science. 1999; 283: 1544-1548Crossref PubMed Scopus (1926) Google Scholar). Although these studies have highlighted the importance of PTP-1B as a negative regulator in insulin signaling, it is conceivable that these results could possibly be due to an indirect effect of PTP-1B on insulin signaling and not due to a direct interaction between PTP-1B and the IR. PTP-1B has been shown to be located on the cytoplasmic surface of the ER (10Frangioni Neel B.G. Cell. Full Text PDF PubMed Scopus Google the IR is located at the plasma where and these two interact the has not been N. D. 2003; PubMed Scopus Google on the interaction between PTP-1B and the IR in cells bioluminescence resonance energy transfer these could that PTP-1B and the IR where in the this interaction was not where PTP-1B and the IR interact the have PTP-1B and the IR to of the green fluorescent protein (GFP) and fluorescence resonance energy transfer to the interaction A. A. L. Biochem. 20: Full Text PDF PubMed Scopus Google Scholar). FRET is a and for in the of A. A. L. Biochem. 20: Full Text PDF PubMed Scopus Google Scholar). this that PTP-1B with the IR in a specific Furthermore, this interaction be by the addition of a specific PTP-1B inhibitor. for W1B and Topaz in were obtained from and and were from The was from and were by and human PTP-1B were from and the IR for the α and β subunits were obtained from and The was by was obtained from Cell were obtained from were obtained from and was from The fusion the was by P. J. Cell 2001; PubMed Scopus Google Scholar). The PTP-1B was synthesized at of the be in M. C. C. Y. 2001; Google were by of W1B and Topaz in W1B was from by that an site was at the and an site at its with of the The was into and then into and was to an site at its and a site at its and into with and the PTP-1B was to the of W1B in to a fusion protein a between the and the W1B-PTP-1B substrate-trapping D. A. PubMed Scopus Google at was to the as a of W1B-PTP-1B in cells, the fusion was from and into the in the wild type or were from with and and into the in The Topaz was from and and into the site of The of the human IR was from by L. E. M. Cell. Full Text PDF PubMed Scopus Google and the was by to the site and to an site and the at the The resulting Topaz-IR fusion protein a between the of the human IR and the of of the was by and of W1B-PTP-1B for cells the were in at to an of by the addition of and for an 2 of cells was with PBS, in of and the addition of of the cells were at with Cell was then by at for and of protein was by for a further at to The was and with of and of protein at with the was by and the were then with and in 2 and activity was as Y. Kennedy B.P. E. Biochem. 2003; Scopus (23) Google Scholar). Cell and 293 cells were in with and in a with and cells were at a of cells were transfected with of the and the the was and with growth were growth for confocal microscopy and FRET were as that cells were transfected in were at a of of a and for to The cells were transfected with of the and growth as of the were to the cells were transfected by the same but and of 293 Cell transfected 293 cells were in with from to at with the cells were on the was and the cells were with were by the cells into of and a Cell was by of the for at The were used for or were further for In the the were to a protein of and of was with of at with of protein was then to the and the were for a further at with The were then by and with by two with were by on The for of and were the the were at were in and with the in the of by Protein were by the and FRET were transfected and on as insulin cells were with and with for at three with PBS, the cells were with to the and a confocal of cells were with an of a of and a Topaz was an of a of and a The for FRET were an of a of and a were a were and FRET was as in where and were the of the the FRET, and Topaz a is the of W1B into the FRET and was the of Topaz into the FRET were no from W1B Topaz or a and were by of cells W1B or Topaz and the of in the FRET and that in the W1B or Topaz of was and a of was the to were for of and of Protein of to PTP-1B and the IR was used to the cellular between PTP-1B and IR. that PTP-1B activity to at its N W1B-PTP-1B was and from cells, and its to was W1B-PTP-1B phosphatase activity with to and which could be by treatment with a PTP-1B E. C. P. Ramachandran C. Y. Kennedy B.P. G. PubMed Scopus Google not increase the of PTP-1B and IR for of cellular examined the substrate-trapping form of PTP-1B (W1B-PTP-1BD181A) D. A. PubMed Scopus Google Scholar). The of W1B-PTP-1BD181A and Topaz-IR co-transfected into 293 cells was examined and the of an with and which to the of the W1B-PTP-1B fusion with and and the of a to the of the β-subunit of the IR to The also the of a in addition to with the and β-subunit and also with an the α of the IR not that the to the form of the The of the IR in transfected 293 cells has been J. A. J. Biol. Chem. Full Text PDF PubMed Google Scholar). These also that the β-subunit and the α-β tyrosine in the of J. A. J. Biol. Chem. Full Text PDF PubMed Google Scholar). also results in this in which a significant of basal phosphorylation was on for the and the treatment with the of phosphorylation on the fusion protein increased at was no in the phosphorylation of the IR is with the in which insulin treatment increased the tyrosine phosphorylation of the β-subunit but no on the of phosphorylation J. A. J. Biol. Chem. Full Text PDF PubMed Google Scholar). the addition of the to PTP-1B and IR not as the protein could be from with an In the W1B-PTP-1B with the fusion protein and the form of the However, treatment of the cells with insulin only resulted in an increase in the of the fusion protein that was in with W1B-PTP-1BD181A of W1B-PTP-1B and Topaz-IR in 293 in cells, the A. A. L. Biochem. 20: Full Text PDF PubMed Scopus Google Scholar). distribution of is shown in where a fusion protein with a was used P. J. Cell 2001; PubMed Scopus Google Scholar). the and FRET the However, fusion of W1B to either wild type and or the substrate-trapping of PTP-1B and results in distribution of the W1B fluorescence to a of the the cellular distribution for in cells was and this distribution is with the of PTP-1B with the ER N. D. 2003; PubMed Scopus Google Scholar). for the of W1B-PTP-1B to the ER was obtained W1B-PTP-1BD181A was co-transfected with a ER W1B-PTP-1B and distribution and of the Topaz-IR co-transfected with the wild type W1B-PTP-1B resulted in the of the Topaz-IR fluorescence to the plasma membrane and was and no FRET was with wild type W1B-PTP-1B either in the or of insulin and In contrast, of Topaz-IR with the substrate-trapping form of PTP-1B, resulted in a significant increase in Topaz-IR intracellular fluorescence In a of the Topaz-IR and W1B-PTP-1BD181A cells, was as as a significant basal of FRET The FRET in these cells to have an ER type of on the results from in which a of cells examined cells were of cells FRET in the of FRET between W1B-PTP-1B and basal FRET in cells W1B-PTP-1BD181A and Topaz-IR is with the of the Topaz-IR by W1B-PTP-1BD181A as it is in the enhanced basal interaction between the substrate-trapping form of PTP-1B and the IR was also by to the interaction between the IR and PTP-1B N. D. 2003; PubMed Scopus Google Scholar). the basal as of the cells FRET, the W1B-PTP-1BD181A and not However, treatment of these cells with insulin resulted in a of the of the cells a FRET as in where cells cells were examined insulin a FRET The insulin-induced FRET as as insulin treatment as a perinuclear and increased in and over time C and insulin the FRET was and in a perinuclear The cellular of this insulin-induced FRET is to that of the compartment A. M. Mol. Cell Biol. PubMed Scopus Google Scholar, 2000; PubMed Scopus Google Scholar). that and back to the plasma membrane this compartment A. M. Mol. Cell Biol. PubMed Scopus Google Scholar). to results in to the in appears as a fluorescent in a perinuclear 2000; PubMed Scopus Google Scholar, J. Cell. Physiol. 2001; PubMed Scopus Google Scholar, L. M. M. J. Cell Biol. PubMed Scopus Google Scholar). the IR and PTP-1B to an endosomal perinuclear the Topaz-IR and W1B-PTP-1BD181A were co-transfected with RhoB to various In the of Topaz-IR fluorescence to the plasma membrane and is with which to endosome P. A. J. Cell Biol. PubMed Scopus Google Scholar). insulin is significant internalization of the Topaz-IR fluorescence as as a of fluorescence to a perinuclear endosomal where is an with Topaz-IR W1B-PTP-1BD181A in either the or of is and, insulin treatment to a perinuclear endosome its with W1B-PTP-1BD181A is with the endosome insulin treatment. 293 cells co-transfected with Topaz-IR and the endosome and with or insulin for The fluorescence shown is in Topaz-IR fluorescence is shown in and appears 293 cells co-transfected with W1B-PTP-1BD181A and and with or insulin for W1B-PTP-1BD181A fluorescence is shown in fluorescence is shown in and appears FRET between W1B-PTP-1BD181A and Topaz-IR with a PTP-1B the FRET between W1B-PTP-1BD181A and Topaz-IR was to PTP-1B co-transfected 293 cells were with a and PTP-1B is a M. C. C. Y. 2001; Google that has an of on PTP-1B and is with PTPs on for 293 cells W1B-PTP-1BD181A and Topaz-IR were with PTP-1B for to insulin of insulin was a in the of Topaz-IR cells were with the PTP-1B Topaz-IR to be localized to the plasma membrane and a with distribution is of the Topaz-IR with wild type W1B-PTP-1B and not W1B-PTP-1BD181A and Topaz-IR were by treatment with the PTP-1B because the of cells FRET FRET in the of were not with treatment. However, the insulin-induced FRET was cells were with inhibitor. In the insulin-induced FRET was in a by treatment of the cells with of the PTP-1B These results suggest that the PTP-1B to the site of W1B-PTP-1BD181A and its interaction with W1B-PTP-1BD181A and Topaz-IR in a basal this interaction is then resistant to the inhibitor. PTP-1B as a negative regulator of insulin signaling by dephosphorylating the activated IR. Although is for PTP-1B in this it this out in the is because PTP-1B in the ER, and the IR is located in the plasma this interaction occurs, confocal microscopy and FRET were used to this interaction the FRET has a for in cells, because it is only the A. A. L. Biochem. 20: Full Text PDF PubMed Scopus Google Scholar, A. M. F. Biol. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, T.S. P. A. 2001; PubMed Scopus Google Scholar, A. Mol. Biol. Cell. PubMed Scopus Google Scholar). The and have been used and for FRET because of significant in and resistance to P. J. Cell 2001; PubMed Scopus Google Scholar). Using this have that PTP-1B not only with the IR an endosomal compartment in an but also to interact in an in the ER, PTP-1B is to in the of the IR to the plasma The of PTP-1B in the tyrosine phosphorylation state of the IR during has been J. A. J. Biol. Chem. Full Text PDF PubMed Google Scholar). that of the IR as as of various receptor tyrosine in 293 cells resulted in a significant of receptor tyrosine phosphorylation J. A. J. Biol. Chem. Full Text PDF PubMed Google Scholar). phosphorylation not only on the β-subunit but also on the with insulin caused a increase in β-subunit tyrosine phosphorylation effect on the was the PTP-1B was co-transfected with the IR, the phosphorylation in the and of insulin results Topaz-IR was with PTP-1BD181A dephosphorylate the the and β-subunit a basal of phosphorylation in the of The of processing of the IR for the increased intracellular Topaz-IR also for the FRET in a of the cells due to the Topaz-IR It is also that the of Topaz-IR as it the ER to the plasma membrane by W1B-PTP-1BD181A results in the of the in the ER, for the FRET in some cells. the FRET results from a between W1B-PTP-1BD181A and it be resistant to by a PTP-1B which the addition of the PTP-1B no effect on the FRET In contrast, the addition of the PTP-1B W1B-PTP-1BD181A from interacting with resulting in the distribution of Topaz-IR to the plasma membrane in these cells. the of the insulin-induced FRET be due to PTP-1B an interaction with the activated IR. It is not only of the cells the FRET, because it be that PTP-1B with the IR during then a of cells have a FRET 293 cells an PTP-1B that be to to the processing of Topaz-IR. in the cells with the FRET, Topaz-IR is at a the of the PTP-1B, for increased with In contrast, co-expression of Topaz-IR with W1B to wild type PTP-1B results in the processing of the IR and of Topaz-IR to the plasma to the interaction between the IR and the substrate-trapping of PTP-1B, also an basal that to PTP-1B interacting with the IR during N. D. 2003; PubMed Scopus Google Scholar). In either the of the ER from PTP-1B or treatment of cells with resulted in a significant in basal BRET, which a role for the PTP-1B in the of the IR. of the tyrosine kinase activity of the IR activated, resulting in autophosphorylation to the receptor, which then into (5Di Guglielmo G.M. Drake P.G. Baass P.C. Authier F. Posner B.I. Bergeron J.M. Mol. Cell. Biochem. 1998; 182: 59-63Crossref PubMed Scopus (111) Google Scholar, 6Bevan A.P. Seabright P.J. Tikerpae J. Posner B.I. Smith G.D. Siddle K. Mol. Cell. Endocrinol. 2000; 164: 145-157Crossref PubMed Scopus (8) Google Scholar). These the of insulin and the internalization not only for the IR to phosphorylate substrates that from at the plasma membrane but also for the insulin (5Di Guglielmo G.M. Drake P.G. Baass P.C. Authier F. Posner B.I. Bergeron J.M. Mol. Cell. Biochem. 1998; 182: 59-63Crossref PubMed Scopus (111) Google Scholar, 6Bevan A.P. Seabright P.J. Tikerpae J. Posner B.I. Smith G.D. Siddle K. Mol. Cell. Endocrinol. 2000; 164: 145-157Crossref PubMed Scopus (8) Google Scholar). in the the dissociation of insulin from the receptor further receptor activation. some dephosphorylation occurs, and the receptor is sorted to back to the plasma membrane or to the for In this insulin-induced FRET between W1B-PTP-1BD181A and Topaz-IR was as as insulin suggesting that the interaction between IR and PTP-1B IR activation and These results in with the time for the activation and dephosphorylation of the IR, as as with for the interaction between the IR and PTP-1B that be of insulin addition (5Di Guglielmo G.M. Drake P.G. Baass P.C. Authier F. Posner B.I. Bergeron J.M. Mol. Cell. Biochem. 1998; 182: 59-63Crossref PubMed Scopus (111) Google Scholar, N. D. 2003; PubMed Scopus Google Scholar, G. Bergeron J.M. Posner B.I. J. Biol. Chem. Full Text PDF PubMed Google Scholar). The that the FRET over time insulin addition that W1B-PTP-1BD181A is Topaz-IR in the endosome its dephosphorylation and back to the plasma of IR dephosphorylation by has been shown to an of the IR in and of to the plasma membrane A.P. Seabright P.J. Tikerpae J. Posner B.I. Smith G.D. Siddle K. Mol. Cell. Endocrinol. 2000; 164: 145-157Crossref PubMed Scopus (8) Google Scholar). of Topaz-IR and treatment of cells with insulin results in of Topaz-IR with an endosomal protein that is in and but not in P. A. J. Cell Biol. PubMed Scopus Google Scholar). The of Topaz-IR with RhoB in a perinuclear endosome compartment and the that the insulin-induced FRET between W1B-PTP-1BD181A and Topaz-IR also in the same cellular suggest that PTP-1B with the IR in this perinuclear endosome It has been shown that receptor in 293 cells a perinuclear compartment distribution is to that for the insulin-induced FRET 2000; PubMed Scopus Google Scholar, J. Cell. Physiol. 2001; PubMed Scopus Google Scholar, L. M. M. J. Cell Biol. PubMed Scopus Google Scholar). Although that PTP-1B with the IR in this it is the The of the interaction that insulin a between and IR to but where this interaction was not N. D. 2003; PubMed Scopus Google Scholar). The that the form of the PTP-1B substrate-trapping a insulin-induced the cytosolic form and that the ER form of PTP-1B internalization of the IR for to Posner and have out on IR internalization and signaling (5Di Guglielmo G.M. Drake P.G. Baass P.C. Authier F. Posner B.I. Bergeron J.M. Mol. Cell. Biochem. 1998; 182: 59-63Crossref PubMed Scopus (111) Google Scholar, 6Bevan A.P. Seabright P.J. Tikerpae J. Posner B.I. Smith G.D. Siddle K. Mol. Cell. Endocrinol. 2000; 164: 145-157Crossref PubMed Scopus (8) Google Scholar, G. Bergeron J.M. Posner B.I. J. Biol. Chem. Full Text PDF PubMed Google Scholar). The results with studies on the activation of the IR and its internalization into also that PTP-1B was not in liver the activated IR G. Bergeron J.M. Posner B.I. J. Biol. Chem. Full Text PDF PubMed Google Scholar). Although some of PTP-1B with it is that PTP-1B is localized to the but, this be due to the that the ER and the endosome into the interaction between PTP-1B and the activated IR between the endosome and the FRET was also used to the interaction of PTP-1B with the epidermal and platelet-derived growth factor and that the interaction in on the surface of the ER P.J. A. Neel B.G. Science. PubMed Scopus Google Scholar). The PTP-1B and IR interaction that the PTP-1B to and the IR in a single perinuclear endosome in the interaction between PTP-1B and the IR with and could be to the used in the 293 a mouse or to in growth factor receptor and not from in endosomes, of the receptor to A. M. Mol. Cell Biol. PubMed Scopus Google Scholar). insulin from the IR in endosomes, and the receptor is (5Di Guglielmo G.M. Drake P.G. Baass P.C. Authier F. Posner B.I. Bergeron J.M. Mol. Cell. Biochem. 1998; 182: 59-63Crossref PubMed Scopus (111) Google Scholar, 6Bevan A.P. Seabright P.J. Tikerpae J. Posner B.I. Smith G.D. Siddle K. Mol. Cell. Endocrinol. 2000; 164: 145-157Crossref PubMed Scopus (8) Google Scholar). in the of the PTP-1B and receptor tyrosine kinase In the studies FRET between PTP-1B and the IR was as as receptor activation time were not FRET between and could be at but was at The rapid of the PTP-1B and IR interaction is with a N. D. 2003; PubMed Scopus Google Scholar). the direct and of FRET two in this as with the and the with a PTP-1B used in the In have shown that insulin-induced activation of the IR results in a rapid and direct interaction between PTP-1B and the IR in a perinuclear endosome compartment in 293 cells. However, with the and of these it be to in insulin especially from and and for and Cromlish for

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