synergistic activation of a family of phosphoinositide 3-kinase via g-protein coupled and tyrosine...

Chemistry and Physics of Lipids

98 (1999) 79–86

Synergistic activation of a family of phosphoinositide 3-kinasevia G-protein coupled and tyrosine kinase-related receptors

Toshiaki Katada a,*, Hiroshi Kurosu a, Taro Okada a, Takahiro Suzuki a,Noriko Tsujimoto a, Shunsuke Takasuga a, Kenji Kontani a, Osamu Hazeki a,

Michio Ui b

a Department of Physiological Chemistry, Graduate School of Pharmaceutical Sciences, Uni6ersity of Tokyo, 7-3-1 Hongo,Bunkyo-ku, Tokyo 113-0033, Japan

b Ui Laboratory, the Institute of Physical and Chemical Research, Hirosawa 2-1, Wako-shi 351-01, Japan

Abstract

Phosphoinositide 3-kinase (PI 3-kinase) is a key signaling enzyme implicated in a variety of receptor-stimulated cellresponses. Stimulation of receptors possessing (or coupling to) protein-tyrosine kinase activates heterodimeric PI3-kinases, which consist of an 85-kDa regulatory subunit (p85) containing Src-homology 2 (SH2) domains and a110-kDa catalytic subunit (p110a or p110b). Thus, this form of PI 3-kinases could be activated in vitro by aphosphotyrosyl peptide containing a YMXM motif that binds to the SH2 domains of p85. Receptors coupling toabg-trimeric G proteins also stimulate the lipid kinase activity of a novel p110g isoform, which is not associated withp85, and thereby is not activated by tyrosine kinase receptors. The activation of p110g PI 3-kinase appears to bemediated through the bg subunits of the G protein (Gbg). In addition, rat liver heterodimeric PI 3-kinases containingthe p110b catalytic subunit are synergistically activated by the phosphotyrosyl peptide plus Gbg. Such enzymaticproperties were also observed with a recombinant p110b/p85a expressed in COS-7 cells. In contrast, anotherheterodimeric PI 3-kinase consisting of p110a and p85 in the same rat liver, together with a recombinant p110a/p85a,was not activated by Gbg, though their activities were stimulated by the phosphotyrosyl peptide. Synergisticactivation of PI 3-kinase by the stimulation of the two major receptor types was indeed observed in intact cells, suchas chemotactic peptide (N-formyl–Met–Leu–Phe) plus insulin (or Fcg II) receptors in differentiated THP-1 andCHO cells and adenosine (A1) plus insulin receptors in rat adipocytes. Thus, PI 3-kinase isoforms consisting of p110bcatalytic and SH2-containing (p85 or its related) regulatory subunits appeared to function as a ‘cross-talk’ enzymebetween the two signal transduction pathways mediated through tyrosine kinase and G protein-coupled receptors.

Abbre6iations: fMLP, N-formyl-Met-Leu-Phe; Ga, a subunits of GTP-binding proteins; Gbg, bg subunits of GTP-bindingproteins; p85, 85-kDa regulatory subunit of PI 3-kinase; p110, 110-kDa catalytic subunit of PI 3-kinase; PI, phosphoinositide;PtdIns(3)P, phosphatidylinositol 3-monophosphate; PtdIns(4)P, phosphatidylinositol 4-monophosphate; PtdIns(3,4)P2, phos-phatidylinositol 3,4-bisphosphate; PtdIns(4,5)P2, phosphatidylinositol 4,5-bisphosphate; PtdIns(3,4,5)P3, phosphatidylinositol 3,4,5-triphosphate; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; SH2, Src homology 2; SH3, Src homology 3.

* Corresponding author. Tel.: +81-3-3812-2111 (ext. 4750); fax: +81-3-3815-9604.E-mail address: katada@ mol.f.u-tokyo.ac.jp (T. Katada)

0009-3084/99/$ - see front matter © 1999 Elsevier Science Ireland Ltd. All rights reserved.

PII : S 0009 -3084 (99 )00020 -1

T. Katada et al. / Chemistry and Physics of Lipids 98 (1999) 79–8680

© 1999 Elsevier Science Ireland Ltd. All rights reserved.

Keywords: G protein; Phosphoinositide 3-kinase; Signal transduction; Tyrosine kinase

1. Introduction

Phosphoinositide 3-kinase (PI 3-kinase) is a keysignaling enzyme implicated in the regulation of abroad array of biological responses including re-ceptor-stimulated mitogenesis, oxidative burst,membrane ruffling, and glucose uptake (Tokerand Cantley, 1997; Vanhaesebroeck et al., 1997a).The activation of PI 3-kinase results in an increasein cellular levels of D-3 phosphorylated phospho-inositides, such as PtdIns(3)P, PtdIns(3,4)P2, andPtdIns(3,4,5)P3 (Fig. 1). These products, however,do not serve as the substrates of phospholipase C(Serunian et al., 1989) and thus have been pro-posed to act as second messengers. In this regard,recent studies have indicated that PtdIns(3,4)P2

can directly activate certain protein kinase C iso-forms (Toker et al., 1994) and PtdIns(3,4,5)P3 iscapable of binding to the PH domain of Grp1, aguanine nucleotide exchange factor of the smallGTP-binding protein ARF1 and the PH domainof PKB/Akt (Hammonds-Odie et al., 1996; Klar-lund et al., 1997; Stephens et al., 1998).

As shown in Fig. 2, at least two types of PI3-kinase, in terms of mode of activation, havebeen described in mammalian cells (Fry, 1994).One is stimulated by membrane-bound receptorsactivating tyrosine kinase, while the other is underthe direct control of heterotrimeric GTP-bindingproteins. The well-known former type has beenstructurally characterized as a heterodimer con-sisting of a 110-kDa catalytic subunit (p110) andan 85-kDa regulatory subunit (p85); the regula-tory subunit contains one SH3 and two SH2domains. Stimulation of tyrosine kinase receptorsby extracellular signals results in phosphorylationof specific tyrosine residues located in the YMXMmotifs of the receptors or adaptor molecules, suchas insulin-receptor substrate-1. These phosphory-lated proteins bind to the SH2 domains of p85and stimulate the lipid kinase activity. The stimu-latory effect of these proteins can thus be mim-icked in vitro by a synthetic tyrosine-phosphorylated peptide possessing the YMXMmotif (Backer et al., 1992; Carpenter et al., 1993).Although several subtypes of p85 and p110 (a, band d) have been identified (Hiles et al.,

Fig. 1. Substrates and products of the reaction catalyzed by PI 3-kinase.

T. Katada et al. / Chemistry and Physics of Lipids 98 (1999) 79–86 81

Fig. 2. Various types of PI 3-kinases and their possible involvement in signal transduction pathways mediated through tyrosinekinase-related and G protein-coupled receptors.

1992; Hu et al., 1993; Vanhaesebroeck et al.,1997b), differences in their functions have notbeen well described.

In addition to the tyrosine phosphorylation-de-pendent activation of PI 3-kinase, the bg-subunitsof G proteins also stimulate the lipid kinase activ-ity (Thomason et al., 1994; Kurosu et al., 1995;Stoyanov et al., 1995; Okada et al., 1996;Stephens et al., 1997). One report showed that apartially purified PI 3-kinase, which was immuno-chemically distinct from p110a and not associatedwith p85, is activated by Gbg (Stephens et al.,1994). Thus, a novel catalytic subunit of PI 3-ki-nase, designated as p110g, has been cloned andshown to be activated in vitro by both the a- andbg-subunits of G proteins (Stoyanov et al., 1995).Receptor-induced translocation of p110g to a cy-toskeletal fraction has also been reported (Zhanget al., 1995). This isozyme lacks the binding site top85 and thus dose not interact with the regulatorysubunit. More recently, Stephens et al. (1997)reported a heterodimeric PI 3-kinase consisting ofa p110g-related 120-kDa (or 117-kDa) catalyticsubunit and a 101-kDa adaptor protein which isdistinct from p85. PI 3-kinase activity of the novelheterodimer was markedly stimulated by Gbg butnot by a tyrosine-phosphorylated peptide.

In contrast to these reports, two groups includ-ing the authors showed that Gbg is capable ofstimulating PI 3-kinase activity immuno-precipi-tated with anti-p85 antibodies (Thomason et al.,1994; Okada et al., 1996). Interestingly, insulin-in-duced accumulation of PtdIns(3,4,5)P3 in THP-1cells was synergistically enhanced by activation ofa pertussis toxin-sensitive G protein (Okada et al.,1996). This observation suggests the presence ofan isozyme that is regulated by both phosphoty-rosyl proteins and G proteins. In this review, wediscuss a PI 3-kinase isoform responsible for suchsynergistic activation and its physiological role incellular signal transduction.

2. Various types of PI 3-kinase and a proposedmodel for their activation mechanisms

When rat liver cytoplasmic proteins were ap-plied to a DEAE-Sepharose column and elutedwith a linear gradient of KCl, there were severalpeaks of PI 3-kinase activity (Kurosu et al., 1995).We found that the PI 3-kinase peak, which waseluted from the column with approximately 150mM KCl, was stimulated by Gbg when assayedwith PtdIns(4,5)P2 as a substrate (Kurosu et al.,


1997). The Gbg-sensitive PI 3-kinase was furtherpurified by means of sequential chromatographywith CM-sepharose, Sephacryl S-300/HR, Blue-Sepharose, and Mono Q columns. The MonoQ-purified PI 3-kinase fraction contained 110- and85-kDa proteins which were recognized with anti-p110b and p85 antibodies, respectively, and thedegree of the Gbg-stimulated activity was corre-lated with both protein amounts. There was, how-ever, no protein band immunoreacted with ananti-p110a specific antibody in the Mono Qfractions.

We next investigated whether a signal associ-ated with protein-tyrosine phosphorylation mayalso regulate the kinase activity, since the Gbg-sensitive fractions contained p85 regulatory sub-unit. A tyrosine-phosphorylated peptide,NGDY*MPMSPKS (Y* indicates phospho-ty-rosine), derived from the sequence of insulin-re-ceptor substrate-1 was used for the present kinaseassay. The phosphotyrosyl peptide produced onlya slight increase in the PI 3-kinase that waspurified on the Mono Q column. However, thecombination of Gbg with the phosphotyrosyl pep-tide caused marked activation of the PI 3-kinase.

The active fractions of the Mono Q columnwere next immunoprecipitated with an antibodyspecific for p110b. The proteins in the immuno-

precipitate were separated by SDS-PAGE andthen analyzed by immunoblotting with the anti-p85 antibody. As expected, an 85-kDa peptide canbe detected in the p110b immunoprecipitate. ThePI 3-kinase activity in the immunoprecipitate wasagain found to be stimulated by Gbg. Furtheraddition of the phosphopeptide to the assay mix-ture increased the Gbg-stimulated activity in theimmuno complex, as was the case in the MonoQ-purified PI 3-kinase. The activity in the immunocomplex was completely inhibited by the presenceof PI 3-kinase inhibitors, wortmannin (Ui et al.,1995) or LY294002 (Vlahos et al., 1994). Thus, theMono Q-purified PI 3-kinase, that is activated ina cooperative manner by Gbg and the phosphoty-rosyl peptide, appears to be a heterodimer consist-ing of p110b and p85 (or a related subunit). For acomparison, another heterodimeric PI 3-kinaseconsisting of p110a and p85 was also separatedfrom the same rat liver, and the p110a-subtypespecificity in the separated fraction was confirmedby immuno-blot analysis. In contrast to the resultswith p110b/p85, Gbg had no stimulatory effect onthe PI 3-kinase activity of the p110a/p85 PI 3-ki-nase fraction, though its activity was certainlystimulated by the phosphotyrosyl peptide.

In addition to the p110b/p85 isoform, we founda 110-kDa (or 100-kDa)/46-kDa heterodimeric

Fig. 3. Effects of Gbg and the phosphotyrosyl peptide on recombinant p110a/p85a, p110b/p85a, and p110g PI 3-kinases. Theepitope-tagged p110 (a, b or g) and/or p85a were co-expressed in COS-7 cells and subjected to immunoprecipitation with amonoclonal antibody against Myc-tag. The immunoprecipitated samples were assayed for the PI 3-kinase activity in the absence orpresence of the phosphotyrosyl peptide (100 mM) and/or Gbg (0.5 mM).


Fig. 4. Various types of PI 3-kinase and a possible model for their activation mechanisms.

form of PI-3 kinase, which had not been tightlyretained on the column (Kurosu et al., 1995). Thisheterodimeric (p110/p46) PI-3 kinase shared ex-actly the same enzymatic properties with thep110b/p85 isoform; the lipid kinase activity of theheterodimer form was stimulated by both Gbgand the phosphotyrosyl peptide and synergisti-cally by the presence of the two activators. Thep110 catalytic and p46 regulatory subunits ap-peared to be the p110b isoform and the spliceform of p85a (Fruman et al., 1994), respectively,based on immunological and amino-terminal se-quence analyses (unpublished results).

To confirm that PI 3-kinase containing thep110b catalytic subunit certainly functions as thetarget isoform for synergistic activation by Gbgand phosphotyrosyl peptide, we expressed Myc-tagged p110a/p85a, p110b/p85a or p110g in COS-7 cells with their cDNAs. As shown in the middlepanel of Fig. 3, PI 3-kinase activity of the recom-binant p110b/p85a was stimulated by both Gbgand the phosphotyrosyl peptide and synergisti-cally by the presence of the two activators. Theactivity of p110a/p85a was increased by the phos-photyrosyl peptide but was unaffected by Gbgregardless of the presence or absence of the phos-photyrosyl peptide (Fig. 3, left panel). In contrast,p110g PI 3-kinase was only activated by Gbg

(Fig. 3, right panel). These results indicate thatthe Gbg-sensitivity of the heterodimer PI 3-kinasewas mediated by p110b.

Various types of PI 3-kinase and a possiblemodel for their activation mechanisms are illus-trated in Fig. 4. In this review, we characterizedbiochemically and immunologically the target PI3-kinase of Gbg-induced stimulation. Immunolog-ical analysis suggested that the rat liver Gbg-sensi-tive PI 3-kinase is a heterodimer consisting ofp110b and p85 (or its related subunit). In fact,Gbg could stimulate an epitope-tagged p110b/p85a PI 3-kinase, which was expressed in COS-7cells (Fig. 3). Both the purified and the recombi-nant p110b/p85 PI 3-kinases were stimulated notonly by Gbg but also by a synthetic phosphotyro-syl peptide (containing YMXM motif) that bindsto the SH2 domain of p85. A surprising feature ofthe p110b/p85 type is its synergistic activation inthe presence of the two effectors. The actions ofthe two effectors are considered to be specific,since (i) Gbg-induced activation could be inhib-ited by the GDP-bound form of Gai-2, and (ii) anonphosphorylated peptide with the same se-quence had no stimulatory effect (Kurosu et al.,1997). On the other hand, heterodimeric PI 3-ki-nases containing p110a isoform as their catalyticsubunits appeared to be totally insensitive to Gbg-


induced activation, indicating that this form maybe selectively involved in the tyrosine kinase-me-diated signal transduction.

Stephens et al. (1994) previously showed that aPI 3-kinase activity distinct from the p110/p85heterodimer was activated by Gbg but was unaf-fected by a phosphotyrosyl peptide. In agreementwith their results, the authors have observed thepresence in THP-1 cells of a PI 3-kinase activitythat is regulated solely by Gbg (Okada et al.,1996). Since there was no protein band im-munoreactive to anti-p85 antibody in those frac-tions, the Gbg-sensitive activities are consideredto be different from the p110b/p85 described inthis study. It is unlikely that the previously re-ported enzyme is a monomer of the p110b subunitfreed from p85, since the enzyme migrates as anapparent molecular mass of 200 000 on a gel-filtration column (Stephens et al., 1994). Instead,the p110g isoform may be responsible for theGbg-sensitive PI 3-kinases not activated by thephosphotyrosyl peptide, since this PI 3-kinase hasbeen recently reported to contain a p101 regula-tory subunit, in addition to the p110g catalyticone (Stephens et al., 1997). Thus, there are at leasttwo species of the catalytic subunits, p110b andp110g, that are Gbg-sensitive.

3. Synergistic activation of PI 3-kinase by thestimulation of G-protein coupled and tyrosinekinase-related receptors

Recent studies have revealed that a tyrosinekinase-associated PI 3-kinase plays an importantrole in cellular signaling mediated by pertussistoxin-sensitive G proteins (Stephens et al., 1993;Ptasznik et al., 1995; Hawes et al., 1996; Lopez-Ilasaca et al., 1997). For examples, an increase inPI 3-kinase activity was observed in immuno-pre-cipitated fractions with anti-phosphotyrosine andanti-src-type protein tyrosine kinase antibodiesupon stimulation of G protein-coupled receptors(Stephens et al., 1993; Ptasznik et al., 1995), andMAP kinase activation induced by Gbg was at-tenuated by the introduction of a dominant nega-tive p85 mutant (Hawes et al., 1996).Furthermore, we previously reported in mono-cytic THP-1 cells that stimulation of chemotacticfMLP receptors, which activate pertussis toxin-sensitive G proteins, potentiated the insulin-in-duced and thus tyrosine kinase-mediatedaccumulation of PtdIns(3,4,5)P3 (Okada et al.,1996).

Such synergistic activation of PI 3-kinase by thetwo groups of membrane receptors has also beenobserved in several cell types, and increased Pt-

Fig. 5. Synergistic activation of PI 3-kinase by G protein-coupled and tyrosine kinase receptors in various types of cells.


dIns(3,4,5)P3 formation appears to be responsiblefor the cell responses (Fig. 5). In the THP-1 cells,PtdIns(3,4,5)P3 formation synergistically stimu-lated by insulin plus fMLP appeared to result inthe activation of protein kinase B and S6 kinase(unpublished results). In addition, PtdIns(3,4,5)P3

formation induced by the combination of fMLP-and FcgII-receptor stimulation potentiated super-oxide formation in differentiated THP-1 cells (In-oue et al., 1997; Tsujimoto et al., 1997). In CHOcells expressing both insulin and fMLP receptors,insulin-induced actin rearrangement (membraneruffling) was enhanced by the simultaneous addi-tion of fMLP (unpublished results). Moreover,insulin-induced glucose uptake was markedly en-hanced by the stimulation of adenosine (A1) re-ceptors in rat adipocytes (unpublished results).These results suggested that p110b/p85 (or theirrelated) PI 3-kinase is indeed regulated down-stream of a pertussis toxin-sensitive G protein-coupled receptor. Thus, the synergistic activationof PI 3-kinase appears to function in the intactcell systems.

Acknowledgements

This work was supported in part by researchgrants from the Scientific Research Fund of theMinistry of Education, Science, Sports, and Cul-ture of Japan, and from the ‘Research for theFuture’ Program from the Japan Society for thePromotion of Science (JSPS-RFTF 96L00505).

References

Backer, J.M., Myers, M.G. Jr., Shoelson, S.E., et al., 1992.Phosphatidylinositol 3%-kinase is activated by associationwith IRS-1 during insulin stimulation. EMBO J. 11, 3469–3479.

Carpenter, C.L., Auger, K.R., Chanudhuri, M., et al., 1993.Phosphoinositide 3-kinase is activated by phosphopeptidesthat bind to the SH2 domains of the 85-kDa subunit. J. Biol.Chem. 268, 9478–9483.

Fruman, D.A., Cantley, L.C., Carpenter, C.L., 1994. Structuralorganization and alternative splicing of the murine phospho-inositide 3-kinase p85a gene. Genomics 37, 113–121.

Fry, M.J., 1994. Structure, regulation and function of phospho-inositide 3-kinases. Biochim. Biophys. Acta 1226, 237–268.

Hammonds-Odie, L.P., Jackson, T.R., Profit, A.A., et al., 1996.Identification and cloning of centaurin-a. A novel phos-phatidylinositol 3,4,5-trisphosphate-binding protein fromrat brain. J. Biol. Chem. 271, 18859–18868.

Hawes, B.E., Luttrell, L.M., van Biesen, T., Lefkowitz, R.J.,1996. Phosphatidylinositol 3-kinase is an early intermediatein the Gbg-mediated mitogen-activated protein kinase sig-naling pathway. J. Biol. Chem. 271, 12133–12136.

Hiles, I.D., Otsu, M., Volinia, S., et al., 1992. Phosphatidylinos-itol 3-kinase: structure and expression of the 110 kd catalyticsubunit. Cell 70, 419–429.

Hu, P., Mondino, A., Skolnik, E. Y., Schlessinger, J., 1993.Cloning of a novel, ubiquitously expressed human phos-phatidylinositol 3-kinase and identification of its binding siteon p85. Mol. Cell. Biol. 13, 7677–7688.

Inoue, S., Kontani, K., Tsujimoto, N., et al., 1997. Protein-ty-rosine phosphorylation by IgG1 subclass CD38 monoclonalantibodies is mediated through stimulation of the FcgIIreceptors in human myeloid cell lines. J. Immunol. 159,5226–5232.

Klarlund, J.K., Guilherme, A., Holik, J.J., Virbasius, J.V.,Chawla, A., Czech, M.P., 1997. Signaling by phosphoinosi-tide-3,4,5-trisphosphate through proteins containing pleck-strin and Sec7 homology domains. Science 275, 1927–1930.

Kurosu, H., Hazeki, O., Kukimoto, I., et al., 1995. Radiolabel-ing of catalytic subunits of PI 3-kinases with 17b-hydroxy-16a-[125I]iodowortmannin: identification of theGbg-sensitive isoform as a complex composed of 46-kDaand 100-kDa subunits. Biochem. Biophys. Res. Commun.216, 655–661.

Kurosu, H., Maehama, T., Okada, T., et al., 1997. Het-erodimeric phosphoinositide 3-kinase consisting of p85 andp110b is synergistically activated by the bg subunits of Gproteins and phosphotyrosyl peptide. J. Biol. Chem. 272,24252–24256.

Lopez-Ilasaca, M., Crespo, P., Pellici, P.G., Gutkind, J.S.,Wetzker, R., 1997. Linkage of G protein-coupled receptorsto the MAPK signaling pathway through PI 3-kinase g.Science 275, 394–397.

Okada, T., Hazeki, O., Ui, M., Katada, T., 1996. Synergisticactivation of PtdIns 3-kinase by tyrosine-phosphorylatedpeptide and bg-subunits of GTP-binding proteins. Biochem.J. 317, 475–480.

Ptasznik, A., Traynor-Kaplan, A., Bokoch, G.M., 1995. Gprotein-coupled chemoattractant receptors regulate Lyntyrosine kinase.Shc adapter protein signaling complexes. J.Biol. Chem. 270, 19969–19973.

Serunian, L., Haber, M.T., Fukui, T., et al., 1989. Polyphospho-inositides produced by phosphatidylinositol 3-kinase arepoor substrates for phospholipases C from rat liver andbovine brain. J. Biol. Chem. 264, 17809–17815.

Stephens, L., Eguinoa, A., Corey, S., Jackson, T., Hawkins,P.T., 1993. Receptor stimulated accumulation of phos-phatidylinositol (3,4,5)-trisphosphate by G-protein medi-ated pathways in human myeloid derived cells. EMBO J. 12,2265–2273.

Stephens, L., Smrcka, A., Cooke, F.T., Jackson, T.R., Stern-weis, P.C., Hawkins, P.T., 1994. A novel phosphoinositide


3 kinase activity in myeloid-derived cells is activated by Gprotein bg subunits. Cell 77, 83–93.

Stephens, L., Eguinoa, A., Erdjument-Bromage, H., et al.,1997. The Gbg sensitivity of a PI3K is dependent upon atightly associated adaptor, p101. Cell 89, 105–114.

Stephens, L., Anderson, K., Stokoe, D., et al., 1998. Proteinkinase B kinases that mediate phosphatidylinositol 3,4,5-trisphosphate-dependent activation of protein kinase B.Science 279, 710–714.

Stoyanov, B., Volinia, S., Hanck, T., et al., 1995. Cloning andcharacterization of a G protein-activated human phospho-inositide-3 kinase. Science 269, 690–693.

Thomason, P.A., James, S.R., Casey, P.J., Downes, C.P.,1994. A G-protein bg-subunit-responsive phosphoinositide3-kinase activity in human platelet cytosol. J. Biol. Chem.269, 16525–16528.

Toker, A., Cantley, L.C., 1997. Signalling through the lipidproducts of phosphoinositide-3-OH kinase. Nature 387,673–676.

Toker, A., Meyer, M., Reddy, K.K., et al., 1994. Activation ofprotein kinase C family members by the novel polyphos-phoinositides PtdIns-3,4-P2 and PtdIns-3,4,5-P3. J. Biol.Chem. 269, 32358–32367.

Tsujimoto, N., Kontani, K., Inoue, S., et al., 1997. Potentia-tion of chemotactic peptide-induced superoxide generationby CD38 ligation in human myeloid cell lines. J. Biochem.121, 949–956.

Ui, M., Okada, T., Hazeki, K., Hazeki, O., 1995. Wortmanninas a unique probe for an intracellular signalling protein,phosphoinositide 3-kinase. Trends Biochem. Sci. 20, 303–307.

Vanhaesebroeck, B., Leevers, S.J., Panayotou, G., Waterfield,M.D., 1997. Phosphoinositide 3-kinases: a conserved fam-ily of signal transducers. Trends Biochem. Sci. 22, 267–272.

Vanhaesebroeck, B., Welham, M.J., Kotani, K., et al., 1997.p110d, A novel phosphoinositide 3-kinase in leukocytes.Proc. Natl. Acad. Sci. USA 94, 4330–4335.

Vlahos, C.J., Matter, W.F., Hui, K.Y., Brown, R.F., 1994. Aspecific inhibitor of phosphatidylinositol 3-kinase, 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one(LY294002). J. Biol. Chem. 269, 5241–5248.

Zhang, J., Zhang, J., Benovic, J.L., et al., 1995. Sequestrationof a G-protein bg subunit or ADP-ribosylation of Rho caninhibit thrombin-induced activation of platelet phospho-inositide 3-kinases. J. Biol. Chem. 270, 6589–6594.

.

synergistic activation of a family of phosphoinositide 3-kinase via g-protein coupled and tyrosine...

Documents