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Phosphatidylinositol 3-Kinase p85 Adaptor Function in T-cells

CO-STIMULATION AND REGULATION OF CYTOKINE TRANSCRIPTION INDEPENDENT OF ASSOCIATED p110*
Open AccessPublished:October 25, 2001DOI:https://doi.org/10.1074/jbc.M107648200
      Phosphatidylinositol 3-kinase (PI3K) is a key regulator of a variety of cellular functions from cytoskeletal organization, vesicular trafficking, and cell proliferation to apoptosis. The enzyme complex is comprised of an 85-kDa adaptor (p85) coupled to a 110-kDa catalytic subunit (p110). While the function of PI3K has been largely attributed to the generation of D-3 lipids, an unanswered question has been whether p85 with a number of motifs (SH2, SH3, BcR homology (BH) region) can generate independent intracellular signals. In this study, we demonstrate that p85 lacking p110 (Δp85) can activate NFAT transcription in T-cell hybridomas and normal splenocytes. This up-regulatory effect was unaffected by inhibition of PI 3-kinase, and cooperated specifically with Rac1, but not related family members. Stimulation correlated with Rac1 binding and was lost with the deletion of the BH domain. Lastly, the CD28-Δp85 chimera also cooperated with TcR/CD3 to provide co-signals that enhanced IL-2 transcription. Our findings identify for the first time p85 as an adaptor that operates independently of the classic PI 3-kinase catalytic pathway and further shows that this pathway can provide co-signals in the regulation of T-cell function.
      PI3K
      phosphatidylinositol 3-kinase
      IL
      interleukin
      HA
      hemagglutinin
      Ab
      antibody
      mAb
      monoclonal Ab
      GST
      glutathioneS-transferase
      BcR
      B-cell receptor
      TcR
      T-cell receptor
      BH
      BcR homology domain
      NFAT
      nuclear factor of activated T-cells
      Phosphatidylinositol 3-kinases (PI 3-kinases; PI3K)1 is a key enzyme involved in regulating multiple mammalian cell functions such as cell growth, vesicular trafficking, cytoskeletal organization, proliferation, and apoptosis (
      • Kapeller R.
      • Cantley L.C.
      ,
      • Hiles I.
      • Otsu M.
      • Volinia S.
      • Fry M.
      • Gout I.
      • Dhand R.
      • Panayotou G.
      • Ruiz-Larrea F.
      • Thompson A.
      • Totty N.
      • Hsuan J.
      • Courtneidge S.
      • Parker P.
      • Waterfield M.
      ,
      • Schu P.V.
      • Takegawa K.
      • Fry M.J.
      • Stack J.H.
      • Waterfield M.D.
      • Emr S.D.
      ,
      • Odorizzi G.M.B.
      • Emr S.D.
      ,
      • Vanhaesbroeck B.
      • Leevers S.J.
      • Ahmadi K.
      • Timms J.
      • Katso R.
      • Driscoll P.C.
      • Woscholski R.
      • Parker P.J.
      • Waterfield M.D.
      ). PI3Ks are heterodimer molecules composed of a p85α, β, adapter subunits complexed to p110α, β, or γ catalytic subunits. p110 is both a serine kinase and a lipid kinase that phosphorylates the D-3 position of phosphatidylinositol (PI), phosphatidylinositol 4-phosphate (PI 4-P), and phosphatidylinositol 4,5-bisphosphate (PI 4,5-P2) to generate phosphatidylinositol 3-phosphate (PI 3-P), phosphatidylinositol 3,4-bisphosphate (PI 3,4-P2) and phosphatidylinositol 3,4,5-trisphosphate (PI 3,4,5-P3), respectively (
      • Kapeller R.
      • Cantley L.C.
      ,
      • Hiles I.
      • Otsu M.
      • Volinia S.
      • Fry M.
      • Gout I.
      • Dhand R.
      • Panayotou G.
      • Ruiz-Larrea F.
      • Thompson A.
      • Totty N.
      • Hsuan J.
      • Courtneidge S.
      • Parker P.
      • Waterfield M.
      ,
      • Rameh L.E.
      • Cantley L.C.
      ,
      • Dhand R.I.
      • Hiles I.
      • Panayotou G.
      • Roche S.
      • Fry M.J.
      • Gout I.
      • Totty N.F.
      • Truong O.
      • Vincendo P.
      • Yonezawa K.
      • Kasuga M.
      • Courtneidge S.A.
      • Waterfield M.D.
      ). By contrast, while the p85 subunit has no catalytic activity, it has proline-rich sequences and domains that include a BcR homology domain (BH domain), two SH2 domains, proline-rich motifs, an inter-SH2 region, and an SH3 domain (
      • Zheng Y.S.
      • Bagrodia S.
      • Cerione R.A.
      ,
      • Tolias K.F.
      • Cantley L.C.
      • Carpenter C.L.
      ). Proline sequences can bind to the SH3 domains of Src kinases and other PI 3-kinase complexes (
      • Prasad K.V.S.
      • Janssen O.
      • Kapeller R.
      • Raab M.
      • Cantley L.C.
      • Rudd C.E.
      ,
      • Pleiman C.M.
      • Hertz W.M.
      • Cambier J.C.
      ). The BH domain binds to the Rho family members Rac1 and Cdc42, while inter-SH2 region binds to the N-terminal region of p110 (
      • Hu P.
      • Schlessinger J.
      ,
      • Beeton C.A.
      • Das P.
      • Waterfield M.S.
      • Shepherd P.R.
      ). Further, it is well established that p85 SH2 domains bind to phosphorylated YMXM motifs of various proteins such as middle T antigen and receptors for growth factors platelet-derived growth factor, epidermal growth factor, and CSF-1 (
      • Fantl W.J.
      • Escobedo J.A.
      • Martin G.A.
      • Turck C.W.
      • delRosarion M.
      • McCormick F.
      • Williams L.T.
      ,
      • Chan T.O.
      • Rittenhouse S.E.
      • Tsichlis P.N.
      ,
      • Songyang Z.
      • Shoelson S.E.
      • Chanduri M.
      • Gish G.
      • Pawson T.
      • haser W.G.
      • King F.
      • Roberts T.
      • Ratnofsky S.
      • Lechleider R.J.
      • Neel B.G.
      • Birge R.B.
      • Fajardo J.E.
      • Chou M.M.
      • Hanafusa H.
      • Schaffhausen B.
      • Cantley L.C.
      ). In certain instances, SH2, SH3, and the BH domain binding to ligand can up-regulate p110 catalytic activity (
      • Zheng Y.S.
      • Bagrodia S.
      • Cerione R.A.
      ,
      • Pleiman C.M.
      • Hertz W.M.
      • Cambier J.C.
      ,
      • Carpenter C.L.
      • Auger K.R.
      • Chanudhuri M.
      • Yoakim M.
      • Schaffhausen B.
      • Shoelson S.
      • Cantley L.C.
      ). By generating D-3 lipids, which bind the inner face of the plasma membrane, PI 3-kinase facilitates recruitment of pleckstrin homology (PH) domain-carrying proteins such as PDK1 (phosphatidyl 3,4,5-trisphosphate-dependent protein kinase-1) and AKT (protein kinase B). In this context, the lipid kinase has been found to regulate protein-serine kinases, PDK1 and AKT (
      • Chan T.O.
      • Rittenhouse S.E.
      • Tsichlis P.N.
      ,
      • Gold M.R.
      • Ingham R.J.
      • McLeod S.J.
      • Christian S.L.
      • Scheid M.P.
      • Duronio V.
      • Santos L.
      • Matsuuchi L.
      ,
      • Franke F.
      • Yang S.I.
      • Chan T.O.
      • Datta K.
      • Kazlauskas A.
      • Morrison D.K.
      • Kaplan D.R.
      • Tsichlis P.N.
      ,
      • Genot E.M.
      • Arrieumerlou C.
      • Ku G.
      • Burgering B.M.
      • Weiss A.
      • Kramer I.M.
      ,
      • Toker A.
      ,
      • Vanhaesebroeck B.
      • Alessi D.R.
      ). D-3 lipid binding to AP-2 complexes can also facilitate receptor down-modulation (
      • Franke F.
      • Yang S.I.
      • Chan T.O.
      • Datta K.
      • Kazlauskas A.
      • Morrison D.K.
      • Kaplan D.R.
      • Tsichlis P.N.
      ,
      • Rapoport I.
      • Miyazaki M.
      • Boll W.
      • Duckworth B.
      • Cantley L.C.
      • Shoelson S.
      • Kirchhausen T.
      ).
      T-cell receptor (TcR) and B-cell receptor (BcR) ligation lead to the activation of PI3K and the generation of PI 3,4-P2 and PI 3,4,5-P3 in immune cells (
      • Rudd C.E.
      ,
      • Ward S.G.
      • June C.H.
      • Olive D.
      ,
      • Marshall A.J.
      • Nirro H.
      • Yun T.J.
      • Clark E.A.
      ,
      • Lafont V.
      • Astoul E.
      • Laurence A.
      • Liautard J.
      • Cantrell D.
      ). p85α/β isoforms are expressed (
      • Skolnik E.Y.
      • Margolis B.
      • Mohammadi M.
      • Lowenstein E.
      • Fischer R.
      • Drepps A.
      • Ullrich A.
      • Schlessinger J.
      ,
      • Nolte R.T.
      • Eck M.J.
      • Schlessinger J.
      • Shoelson S.E.
      • Harrison S.C.
      ) and can be recruited to the TcR complex by a combination of TcR zeta chain (
      • Exley M.
      • Varticovski L.
      • Peter M.
      • Sancho J.
      • Terhorst C.
      ) and p56lck/p59fyn-T SH3 domain binding (
      • Prasad K.V.S.
      • Janssen O.
      • Kapeller R.
      • Raab M.
      • Cantley L.C.
      • Rudd C.E.
      ,
      • Prasad K.V.S.
      • Cai Y.C.
      • Raab M.
      • Duckworth B.
      • Cantley L.
      • Shoelson S.E.
      • Rudd C.E.
      ). TcR and BcR ligation can induce tyrosine phosphorylation of p85/p110 and activate serine kinases, protein kinase B, and ribosomal S6 kinase (
      • Lafont V.
      • Astoul E.
      • Laurence A.
      • Liautard J.
      • Cantrell D.
      ). The loss of p85α expression in mice results in a defective B-cell function and differentiation (
      • Fruman D.A.
      • Snapper S.B.
      • Yballe C.M.
      • Davidson L.
      • Yu J.Y.
      • Alt F.W.
      • Cantley L.C.
      ).
      In T-cells, CD28 provides a crucial second signal in TcR-regulated cytokine production and proliferation (
      • Bluestone J.
      ,
      • Linsley P.
      ,
      • Thompson C.B.
      ). However, despite its central role, little is known regarding the molecular basis of co-stimulation. In this sense, the co-receptor acts as a major site of PI3K recruitment due to classic p85 SH2 domain binding to a cytoplasmic YMNM motif (
      • Prasad K.V.S.
      • Cai Y.C.
      • Raab M.
      • Duckworth B.
      • Cantley L.
      • Shoelson S.E.
      • Rudd C.E.
      ,
      • Truitt K.E.
      • Hicks C.M.
      • Imboden J.B.
      ,
      • Pages F.
      • Ragueneau M.
      • Klasen S.
      • Battifora M.
      • Couez D.
      • Sweet R.
      • Truneh A.
      • Ward S.G.
      • Olive D.
      ). By binding to PH domains, D-3 lipids recruit various proteins to the inner face of the lipid bilayer. Nevertheless, studies have come to different conclusions on the contribution of PI 3-kinase to CD28-mediated co-signaling (
      • Rudd C.E.
      ). Mutations that disrupt the motif, or selectively disrupt PI 3-kinase binding attenuate signaling (
      • Pages F.
      • Ragueneau M.
      • Klasen S.
      • Battifora M.
      • Couez D.
      • Sweet R.
      • Truneh A.
      • Ward S.G.
      • Olive D.
      ,
      • Cai Y.-C.
      • Cefai D.
      • Schneider H.
      • Raab M.
      • Nabavi N.
      • Rudd C.E.
      ,
      • Cefai D.
      • Cai Y.-C.
      • Hu H.
      • Rudd C.
      ), while the same mutants showed no defects in Jurkat cells (
      • Lu Y.
      • Phillips C.A.
      • Trevillyan J.M.
      ,
      • Truitt K.E.
      • Shi J.
      • Gibson S.
      • Segal L.G.
      • Mills G.B.
      • Imboden J.B.
      ,
      • Hutchcroft J.E.
      • Franklin D.P.
      • Tsai B.
      • Harrison-Findik D.
      • Vartocovski L.
      • Bierer B.
      ). One explanation for this discrepancy is the finding that Jurkat cells lack phosphoinositide phosphatase PTEN, an enzyme that removes the phosphate moiety on the 3rd position of the inositol ring (
      • Shan X.
      • Czar M.J.
      • Bunnell S.C.
      • Liu P.
      • Liu Y.
      • Schwartzberg P.L.
      • Wange R.L.
      ). Cells with constitutively high levels of PI 3,4-P2and PI 3,4,5-P3 would be less dependent on CD28-associated PI 3-kinase for co-signaling. Other studies using inhibitors of PI 3-kinase catalytic activity have also yielded mixed results, and in certain instances showed a potentiation of IL-2 transcription (
      • Lu Y.
      • Phillips C.A.
      • Trevillyan J.M.
      ,
      • Hausdorff S.F.
      • Fingar D.C.
      • Morioka K.
      • Garza L.A.
      • Whiteman E.L.
      • Summers S.A.
      ,
      • Ueda Y.
      • Levine B.L.
      • Huang M.L.
      • Freeman G.J.
      • Nadler L.M.
      • June C.
      • Ward S.G.
      ,
      • Ward S.G.
      • Wilson A.
      • Turner L.
      • Westwick J.
      • Sansom D.M.
      ). Recent in vivo re-constitution studies of YMNM mutants in mice have confirmed the central importance of the motif in CD28-mediated graft versus host responses (
      • Harada Y.
      • Tokushima M.
      • Matsumoto Y.
      • Ogawa S.
      • Otsuka M.
      • Hayashi K.
      • Weiss B.D.
      • June C.H.
      • Abe R.
      ) and the induction of BcL-XL (
      • Okkenhaug K.
      • Wu L.
      • Garza K.
      • La Rose J.
      • Khoo W.
      • Odermatt B.
      • Mak T.
      • Ohashi P.
      • Rottapel R.
      ). The major downstream target of the kinase, AKT or PKB has recently been implicated in CD28 regulation of IL-2, but not of Th2 cytokines (
      • Kane L.
      • Andres P.
      • Howland K.
      • Abbas A.
      • Weiss A.
      ).
      Given the uncertainty regarding the role of PI 3-kinase in T-cell signaling, we re-examined the possible role of the p85 adaptor alone on the activation process. PI-3 kinase has been reported to bind to the small GTP-binding proteins Rac1 and Cdc42 (
      • Zheng Y.S.
      • Bagrodia S.
      • Cerione R.A.
      ,
      • Tolias K.F.
      • Cantley L.C.
      • Carpenter C.L.
      ,
      • Exton J.H.
      ). Although evidence suggestive of a role of p85-mediated function exists (
      • Gringhuis S.I.
      • deLeij L.F.
      • Wayman G.A.
      • Tokumitsu H.
      • Vellenga E.
      ,
      • Jimenez C.
      • Portela R.A.
      • Mellado M.
      • Rodriguez-Frade J.M.
      • Collard J.
      • Serrano A.
      • Martinez A.C.
      • Avila J.
      • Carrera A.C.
      ,
      • Jascur T.
      • Gilman J.
      • Mustelin T.
      ), a direct demonstration of p85-mediated regulation of cell function independent of p110 has remained elusive. In this study, we show that membrane localized p85 lacking an ability to associate with p110 (p85Δ) potently up-regulates interleukin-2 transcription in Jurkat and normal peripheral T-cells. Rac1 synergistically cooperated with p85, an effect ablated by the loss of the BH domain and its binding to the GTP-binding protein. Specificity was observed by the fact that cooperativity was not inhibited by wortmannin inhibition of lipid kinase activity and that other members of the Rho family, Rho and Rac2 failed to cooperate with p85. Further, co-ligation of CD28-p85Δ with anti-CD3 was found to cooperate to provide co-stimulation in the up-regulation of IL-2 transcription in normal T-cells. Our findings demonstrate for the first time that p85 can generate signals independent of binding to p110 that lead to enhanced gene transcription and co-stimulation.

      DISCUSSION

      PI 3-kinase plays a central role in the regulation of multiple cellular events (
      • Kapeller R.
      • Cantley L.C.
      ,
      • Vanhaesbroeck B.
      • Leevers S.J.
      • Ahmadi K.
      • Timms J.
      • Katso R.
      • Driscoll P.C.
      • Woscholski R.
      • Parker P.J.
      • Waterfield M.D.
      ). Many of these effects are due to the production of D-3 lipids that act to recruit proteins to the membranes of cells. However, a major question has been whether the p85 subunit can itself act as an adaptor that is coupled to other signaling pathways. One alternate pathway is the binding of p85 to small GTPases such as Rac and Cdc42 (
      • Zheng Y.S.
      • Bagrodia S.
      • Cerione R.A.
      ,
      • Tolias K.F.
      • Cantley L.C.
      • Carpenter C.L.
      ). In this context, the cytoplasmic YMNM motif of CD28 binds to PI3K and is required in multiple systems for optimal cytokine release (
      • Pages F.
      • Ragueneau M.
      • Klasen S.
      • Battifora M.
      • Couez D.
      • Sweet R.
      • Truneh A.
      • Ward S.G.
      • Olive D.
      ,
      • Cai Y.-C.
      • Cefai D.
      • Schneider H.
      • Raab M.
      • Nabavi N.
      • Rudd C.E.
      ,
      • Harada Y.
      • Tokushima M.
      • Matsumoto Y.
      • Ogawa S.
      • Otsuka M.
      • Hayashi K.
      • Weiss B.D.
      • June C.H.
      • Abe R.
      ) or BcL-XL expression (
      • Okkenhaug K.
      • Wu L.
      • Garza K.
      • La Rose J.
      • Khoo W.
      • Odermatt B.
      • Mak T.
      • Ohashi P.
      • Rottapel R.
      ). Despite this, variable results have been obtained with the use of inhibitors of the enzyme (
      • Lu Y.
      • Phillips C.A.
      • Trevillyan J.M.
      ,
      • Hausdorff S.F.
      • Fingar D.C.
      • Morioka K.
      • Garza L.A.
      • Whiteman E.L.
      • Summers S.A.
      ,
      • Ueda Y.
      • Levine B.L.
      • Huang M.L.
      • Freeman G.J.
      • Nadler L.M.
      • June C.
      • Ward S.G.
      ,
      • Ward S.G.
      • Wilson A.
      • Turner L.
      • Westwick J.
      • Sansom D.M.
      ). For this reason, we examined the possibility that the p85 adaptor protein might itself act to generate signals in the context of CD28 mediated co-stimulation. Our findings demonstrate that p85Δ, either as a myristoylated protein (mp85Δ), or as a receptor chimera had potent stimulatory effects on IL-2 transcriptional activity. The expression of p85Δ had no obvious effect on endogenous p85 or p110 levels (data not shown). Furthermore, this stimulatory effect was not inhibited by inhibition of PI3K catalytic activity with wortmannin (Fig. 3). Specific synergy was observed with Rac1, where neither Rho nor Rac2 cooperated with p85Δ (Fig. 2), and the loss of the BH domain resulted in a concordant loss of Rac1 binding and transcription (Fig.4). Importantly, our findings also show that p85Δ signaling can operate in normal T-cells, thus eliminating the concern that the pathway only operates in transformed cell lines. Endogenous p85-Rac1 complexes could also be identified in T-cells (Fig. 6C). The major downstream target of this pathway was identified as NFAT as shown by its sensitivity to CsA, and the enhancement of transcription with a NFAT-restricted TNFα reporter. Overall, our findings demonstrate that p85 can operate as an adaptor protein that interacts with Rac1 in the regulation of NFAT-regulated cytokine transcription.
      Our finding of p85Δ-Rac1 regulation of IL-2 transcription has direct relevance to the ability of CD28 to mediate co-stimulation in T-cells. Because CD28 represents the primary site of PI3K recruitment in T-cells (
      • Prasad K.V.S.
      • Cai Y.C.
      • Raab M.
      • Duckworth B.
      • Cantley L.
      • Shoelson S.E.
      • Rudd C.E.
      ,
      • Truitt K.E.
      • Hicks C.M.
      • Imboden J.B.
      ,
      • Pages F.
      • Ragueneau M.
      • Klasen S.
      • Battifora M.
      • Couez D.
      • Sweet R.
      • Truneh A.
      • Ward S.G.
      • Olive D.
      ), a key question is whether the receptor might engage the CD28-Rac1 pathway. Although the importance of the YMNM motif has now been documented in several systems (
      • Pages F.
      • Ragueneau M.
      • Klasen S.
      • Battifora M.
      • Couez D.
      • Sweet R.
      • Truneh A.
      • Ward S.G.
      • Olive D.
      ,
      • Cai Y.-C.
      • Cefai D.
      • Schneider H.
      • Raab M.
      • Nabavi N.
      • Rudd C.E.
      ,
      • Cefai D.
      • Cai Y.-C.
      • Hu H.
      • Rudd C.
      ,
      • Harada Y.
      • Tokushima M.
      • Matsumoto Y.
      • Ogawa S.
      • Otsuka M.
      • Hayashi K.
      • Weiss B.D.
      • June C.H.
      • Abe R.
      ), the use of inhibitors of the enzyme has yielded mixed results (
      • Lu Y.
      • Phillips C.A.
      • Trevillyan J.M.
      ,
      • Hausdorff S.F.
      • Fingar D.C.
      • Morioka K.
      • Garza L.A.
      • Whiteman E.L.
      • Summers S.A.
      ,
      • Ueda Y.
      • Levine B.L.
      • Huang M.L.
      • Freeman G.J.
      • Nadler L.M.
      • June C.
      • Ward S.G.
      ,
      • Ward S.G.
      • Wilson A.
      • Turner L.
      • Westwick J.
      • Sansom D.M.
      ,
      • Reif K.
      • Lucas S.
      • Cantrell D.
      ). In certain instances, wortmannin was even found to increase stimulation in a manner that is similar to that observed in our studies on p85Δ signaling (Fig. 3). Further, co-ligation of CD28-p85Δ and TcRζ/CD3 potentiated NFAT-mediated IL-2 transcription in both DC27.10 and normal murine splenocytes (Figs. 5 and 6). The level of co-stimulation occurred at levels comparable with that reported in other studies (
      • Krummel M.F.
      • Allison J.P.
      ,
      • Boise L.H.
      • Minn A.J.
      • Noel P.J.
      • June C.H.
      • Accavitti M.A.
      • Lindsten T.
      • Thompson C.B.
      ). Therefore, although our studies do not exclude a role for p110 and the generation of D-3 lipids in ensuring efficient T-cell signaling, they demonstrate that the role for PI3K in co-stimulation is more complex that previously appreciated. In this context, at least part of co-receptor signaling may be attributed to p85 cooperativity with Rac1. Further, the lack of an effect of inhibitors of PI 3-kinase on T-cell function may be an insufficient parameter for excluding a role for p85 in the regulation of a given function. Similarly, functional defects in p85-deficient mice may in part be related to the loss of p85-Rac signaling (
      • Fruman D.A.
      • Snapper S.B.
      • Yballe C.M.
      • Davidson L.
      • Yu J.Y.
      • Alt F.W.
      • Cantley L.C.
      ).
      PI3Ks have been implicated in the regulation of different types of cytoskeletal rearrangements that include ruffling and the disassembly of stress fibers. Similarly the induction of membrane ruffles by growth factors appears to require Rac activation (
      • Ridley A.J.
      • Paterson H.F.
      • Johnston C.L.
      • Diekmann D.
      • Hall A.
      ). Preliminary studies failed to show a detectable re-arrangement of the cytoskeleton (data not shown). Consistent with this, unlike in the case of RhoGAPs, BH domain binding does not activate the intrinsic GTPase activity of Cdc42 (
      • Zheng Y.S.
      • Bagrodia S.
      • Cerione R.A.
      ). Another possible target is PAK1, which is regulated by Rac1 in its regulation of NFAT function in T-cells (
      • Yablonski D.
      • Kane L.P.
      • Qian D.
      • Weiss A.
      ). Indeed, preliminary studies have shown cooperativity between p85Δ, Rac1, and PAK in the stimulation of IL-2 transcription (data not shown). Surprising was the specific synergy between p85 and Rac1 but not Rac2 (Fig. 2). The latter family member differs from Rac1 in the C terminus of the protein (
      • Courjal F.
      • Chuchana P.
      • Theillet C.
      • Fort P.
      ). This suggests that the two Rac family members differ fundamentally in their coupling to other proteins. Surprisingly, Cdc42 also failed to cooperate with p85 in potentiating transcription (data not shown). Lastly, the ubiquitous expression of Rac1 suggests a role for the p85-Rac1 pathway in the up-regulation of general gene activation (
      • Zheng Y.S.
      • Bagrodia S.
      • Cerione R.A.
      ,
      • Gringhuis S.I.
      • deLeij L.F.
      • Wayman G.A.
      • Tokumitsu H.
      • Vellenga E.
      ). Further studies will be needed to define downstream intermediates regulated by p85-Rac1 and the molecular basis for the distinction between the two Rac family members.

      REFERENCES

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