FcϵRI-mediated Induction of Nuclear Factor of Activated T-cells

Nuclear factor of activated T-cells (NFAT) is a transcriptional activator that binds to the interleukin-2 promoter and is believed to be responsible for T-cell-specific interleukin-2 gene expression. Here we demonstrate using electrophoretic mobility shift assays that nuclear NFAT can be induced in the rat basophilic leukemia (RBL-2H3) mast cell line and rat bone marrow-derived mast cells upon cross-linkage of the high affinity receptor (FcϵRI) for immunoglobulin E (IgE). Receptor-dependent activation of NFAT was mimicked by the combination of the protein kinase C activator phorbol myristate acetate and the calcium ionophore ionomycin. The induced binding activity was specific for the NFAT recognition motif because competition with nonradioactive NFAT oligonucleotide abolished the DNA binding activity, whereas nonradioactive oligonucleotides recognized by the transcription factors NFκB, glucocorticoid receptors, and TFIID did not. An oligonucleotide representing the AP-1 recognition sequence also blocked the NFAT DNA binding activity, as did a combination of anti-Fos and anti-Jun antibodies. Using electrophoretic mobility shift assays, AP-1-binding proteins were found to be induced in RBL-2H3 cells under the same conditions as was the NFAT binding activity. Together these data suggest that the NFAT complex in mast cells contains Fos and Jun proteins as does NFAT in T-cells. The appearance of nuclear NFAT binding activity was dependent in part upon calcium mobilization, as buffering the antigen-induced calcium rise with intracellular BAPTA strongly inhibited NFAT activation. Prevention of calcium influx with external EGTA also inhibited NFAT activation, indicating that release of calcium from internal stores was insufficient for sustained activation of mast cell NFAT. Cyclosporin A, a potent inhibitor of the calmodulin-dependent phosphatase calcineurin, blocked the induction of NFAT-DNA binding activity, implicating calcineurin as a key signaling enzyme in this pathway. These results suggest that NFAT is present in the mast cell line RBL-2H3 and in primary bone marrow-derived mast cells, is similar in subunit composition to the T-cell NFAT, and may play a role in calcium-dependent signal transduction in mast cells. Nuclear factor of activated T-cells (NFAT) is a transcriptional activator that binds to the interleukin-2 promoter and is believed to be responsible for T-cell-specific interleukin-2 gene expression. Here we demonstrate using electrophoretic mobility shift assays that nuclear NFAT can be induced in the rat basophilic leukemia (RBL-2H3) mast cell line and rat bone marrow-derived mast cells upon cross-linkage of the high affinity receptor (FcϵRI) for immunoglobulin E (IgE). Receptor-dependent activation of NFAT was mimicked by the combination of the protein kinase C activator phorbol myristate acetate and the calcium ionophore ionomycin. The induced binding activity was specific for the NFAT recognition motif because competition with nonradioactive NFAT oligonucleotide abolished the DNA binding activity, whereas nonradioactive oligonucleotides recognized by the transcription factors NFκB, glucocorticoid receptors, and TFIID did not. An oligonucleotide representing the AP-1 recognition sequence also blocked the NFAT DNA binding activity, as did a combination of anti-Fos and anti-Jun antibodies. Using electrophoretic mobility shift assays, AP-1-binding proteins were found to be induced in RBL-2H3 cells under the same conditions as was the NFAT binding activity. Together these data suggest that the NFAT complex in mast cells contains Fos and Jun proteins as does NFAT in T-cells. The appearance of nuclear NFAT binding activity was dependent in part upon calcium mobilization, as buffering the antigen-induced calcium rise with intracellular BAPTA strongly inhibited NFAT activation. Prevention of calcium influx with external EGTA also inhibited NFAT activation, indicating that release of calcium from internal stores was insufficient for sustained activation of mast cell NFAT. Cyclosporin A, a potent inhibitor of the calmodulin-dependent phosphatase calcineurin, blocked the induction of NFAT-DNA binding activity, implicating calcineurin as a key signaling enzyme in this pathway. These results suggest that NFAT is present in the mast cell line RBL-2H3 and in primary bone marrow-derived mast cells, is similar in subunit composition to the T-cell NFAT, and may play a role in calcium-dependent signal transduction in mast cells. Nuclear factor of activated T-cells (NFAT)1( 1The abbreviations used are: NFATnuclear factor of activated T-cellsNFATpnuclear factor of activated T-cells, preexistingNFATcnuclear factor of activated T-cells, cytosolicFcϵRIreceptor with high affinity for IgEILinterleukinEMSAelectrophorectic mobility shift assayRBLrat basophilic leukemiaRBMMCrat bone marrow-derived mast cellsPKCprotein kinase CPMAphorbol 12-myristate 13-acetateBSAbovine serum albuminTNP-BSAtrinitrophenylated bovine serum albuminBAPTA-AM1,2-bis(2-aminophenoxy)eth-ane-N,N,N′,N′-tetraacetic acidTBETris-borate-EDTA bufferCsAcyclosporin A.) is a transcription complex believed to mediate the final step in the signal transduction pathway linking T-cell receptor engagement with the expression of the interleukin-2 (IL-2) gene (for reviews, see Ullman et al., 1990Ullman K.S. Northrop J.P. Verweij C.L. Crabtree G.R. Annu. Rev. Immunol. 1990; 8: 421-452Crossref PubMed Scopus (491) Google Scholar and Rao, 1994Rao A. Immunol. Today. 1994; 15: 274-281Abstract Full Text PDF PubMed Scopus (490) Google Scholar). Although selective expression of NFAT is thought to account for the T-cell specificity of IL-2 gene expression, recent evidence suggests the possibility that NFAT may be one in a family of related transcription factors that regulate the transcription of cytokine genes in other leukocytes. The present work was undertaken to determine whether NFAT-related activity can be induced in mast cells via cross-linkage of the receptor with high affinity for IgE (FcϵRI). nuclear factor of activated T-cells nuclear factor of activated T-cells, preexisting nuclear factor of activated T-cells, cytosolic receptor with high affinity for IgE interleukin electrophorectic mobility shift assay rat basophilic leukemia rat bone marrow-derived mast cells protein kinase C phorbol 12-myristate 13-acetate bovine serum albumin trinitrophenylated bovine serum albumin 1,2-bis(2-aminophenoxy)eth-ane-N,N,N′,N′-tetraacetic acid Tris-borate-EDTA buffer cyclosporin A. Although the exact molecular make-up of the NFAT complex is unknown, there is evidence that it is composed of a lymphocyte-specific cytoplasmic component (or components) present in unactivated cells, designated NFATp/c, and a ubiquitous, inducible nuclear component containing Fos and Jun proteins (Jain et al., 1992Jain J. McCaffrey P.G. Valge-Archer V.E. Rao A. Nature. 1992; 356: 801-804Crossref PubMed Scopus (429) Google Scholar; Ullman et al., 1990Ullman K.S. Northrop J.P. Verweij C.L. Crabtree G.R. Annu. Rev. Immunol. 1990; 8: 421-452Crossref PubMed Scopus (491) Google Scholar. Cross-linkage of the T-cell receptor results in two intracellular signals required for the assembly of active NFAT in the nucleus: activation of protein kinase C and mobilization of cytosolic calcium. The activation of PKC is necessary for the transcription of Fos and Jun genes (Jain et al., 1992Jain J. McCaffrey P.G. Valge-Archer V.E. Rao A. Nature. 1992; 356: 801-804Crossref PubMed Scopus (429) Google Scholar; Northrop et al., 1993Northrop J.P. Ullman K.S. Crabtree G.R. J. Biol. Chem. 1993; 268: 2917-2923Abstract Full Text PDF PubMed Google Scholar), and an increase in intracellular calcium is required for the translocation of NFATp into the nucleus (Flanagan et al., 1991Flanagan W.M. Corthesy B. Bram R.J. Crabtree G.R. Nature. 1991; 352: 803-807Crossref PubMed Scopus (954) Google Scholar). The translocation of NFATp from the cytoplasm to the nucleus is thought to be controlled either directly or indirectly by the calcium-dependent protein phosphatase calcineurin (Clipstone and Crabtree, 1992Clipstone N. Crabtree G.R. Nature. 1992; 357: 695-697Crossref PubMed Scopus (1477) Google Scholar). The role of calcineurin in this signaling pathway has been further substantiated by the fact that a constitutively active form of calcineurin can bypass the calcium requirement for IL-2 transcriptional activation (O'Keefe et al., 1992O'Keefe S.J. Tamura J. Kincaid R.L. Tocci M.J. O'Neill E.A. Nature. 1992; 357: 692-694Crossref PubMed Scopus (788) Google Scholar). The immunosuppressive drugs cyclosporin A (CsA) and FK506 block transcription of the IL-2 gene by inhibiting the phosphatase activity of calcineurin and preventing the intracellular translocation of NFATp (McCaffrey et al., 1993aMcCaffrey P.G. Perrino B.A. Soderling T.R. Rao A. J. Biol. Chem. 1993; 268: 3747-3752Abstract Full Text PDF PubMed Google Scholar; Clipstone and Crabtree, 1992Clipstone N. Crabtree G.R. Nature. 1992; 357: 695-697Crossref PubMed Scopus (1477) Google Scholar; McCaffrey et al., 1993bMcCaffrey P.G. Luo C. Kerppola T.K. Jain J. Badalian T.M. Ho A.M. Burgeon E. Lane W.S. Lambert J.N. Curran T. Verdine G.L. Rao A. Hogan P.G. Science. 1993; 262: 750-754Crossref PubMed Scopus (379) Google Scholar; Mattila et al., 1990Mattila P.S. Ullman K.S. Fiering S. Emmel E.A. McCutcheon M. Crabtree G.R. Herzenberg L.A. EMBO J. 1990; 9: 4425-4433Crossref PubMed Scopus (291) Google Scholar). Once assembled in the nucleus, the NFAT complex initiates transcription by binding to two sequence-specific binding sites located in the enhancer region of the IL-2 promoter (Shaw et al., 1988Shaw J. Utz P.J. Durand D.B. Toole J.J. Emmel E.A. Crabtree G.R. Science. 1988; 241: 202-205Crossref PubMed Scopus (10) Google Scholar). The signal transduction pathway emanating from the FcϵRI in mast cells shares many relevant features with the T cell receptor-initiated pathway, including an increase in intracellular calcium (Beaven et al., 1984) and activation of PKC, which in turn modulates the expression of AP-1 binding proteins such as Jun and Fos (Lewin et al., 1993; Baranes and Razin, 1991). Antigen-activated mast cells produce cytokines such as IL-3, GM-CSF, TNF-α, IL-4, IL-5, IL-6, and IL-1 (Kaye et al., 1992; Wodnar-Filipowicz et al., 1989; Plaut et al., 1989), and NFAT binding sites have been identified in the regulatory regions of several of these genes (reviewed in Rao, 1994). The NFAT binding site located in the intergenic enhancer region of the IL-3/GM-CSF genes has been shown to bind NFAT from activated T-cells (Cockerill et al., 1993), and CsA inhibits the induction of DNA binding activity directed to this site in Jurkat cells treated with PMA and ionomycin. CsA blocks the FcϵRI-dependent induction of IL-1β, TNF-α, and IL-6 mRNA in mouse bone marrow-derived mast cells, apparently through the mediation of some of the same immunophilin proteins present in T-cells (Hultsch et al., 1991Hultsch T. Albers M.W. Schreiber S.L. Hohman R.J. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 6223-6229Crossref Scopus (114) Google Scholar; Kaye et al., 1992Kaye R.E. Fruman D.A. Bierer B.E. Albers M.W. Zydowsky L.D. Ho S.I. Jin Y. Castells M.C. Schreiber S.L. Walsh C.T. Burakoff S.J. Austen K.F. Katz H.R. Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 8542-8546Crossref PubMed Scopus (62) Google Scholar). This CsA sensitivity suggests a possible role for calcineurin in the FcϵRI-mediated expression of cytokine genes in mast cells. Given the presence of NFAT binding sites in the regulatory regions of cytokines induced by FcϵRI cross-linkage, a key question is, does NFAT mediate transduction of signals from the FcϵRI to the nucleus? Here we show that antigenic stimulation through the Fcϵ receptor induces NFAT DNA binding activity in two types of rat mast cells. The presumed NFAT complex in mast cells, like that in T-cells, contains AP-1 proteins in association with (an)other species conferring specificity for the NFAT recognition motif. Induction of NFAT binding activity by the FcϵRI was mimicked by simultaneous treatment with the PKC activator PMA and the calcium ionophore ionomycin, required calcium mobilization, and was blocked by the calcineurin inhibitor, CsA. The presence of NFAT in mast cells and its activation via the FcϵRI suggest that it may mediate the terminal steps of signal transduction via the FcϵRI, namely the transcription of one or more cytokine genes. The RBL-2H3 (RBL) cell line was maintained in 75-cm2 flasks as monolayers in Eagle's minimum essential medium (Earle's salts) supplemented with 20% fetal bovine serum (Life and as et al., The cells were in a in a the cells were by and to flasks a cell of of bone marrow-derived mast cells were from bone of in the presence of cell factor and rat of the and and molecular of this cell A. M. and for activation were activation was the cells were the by the medium with medium containing IgE from the et al., J. Immunol. PubMed Scopus Google Scholar). the of the cells were from the flasks by the monolayers with containing and the cells in the same for The cells were from the to a and with Eagle's minimum essential medium supplemented with and The cells were in the same and for This of the NFAT activity induced by the cell were activated by of the directly to the cell bovine serum albumin was used a final of and PMA and were used and CsA were by of CsA in of containing of The was to with and of this were directly into the cell the The cells were with CsA activation. cells were in the of the cells were for in medium containing the calcium cell activation. EGTA was a of to medium and the to with This an of of cells were and in medium containing was directly to the cell as a in to a final of Nuclear were as et al., 1990Mattila P.S. Ullman K.S. Fiering S. Emmel E.A. McCutcheon M. Crabtree G.R. Herzenberg L.A. EMBO J. 1990; 9: 4425-4433Crossref PubMed Scopus (291) Google with the cells were used the the cells were and in steps were in a with the to and A, and the cells were by and in of buffer A The cells were in of buffer A and to for The cell was and this of the cells were as by and were and in of buffer C and of The was for to nuclear proteins and for An of was to the proteins were by for and in of buffer C. The protein were and protein were using a binding assay The were in mobility shift assays were as et al., 1990Mattila P.S. Ullman K.S. Fiering S. Emmel E.A. McCutcheon M. Crabtree G.R. Herzenberg L.A. EMBO J. 1990; 9: 4425-4433Crossref PubMed Scopus (291) Google Scholar). for NFAT were in a containing of and of nuclear for AP-1 were in a of of and of nuclear proteins et al., 1990Mattila P.S. Ullman K.S. Fiering S. Emmel E.A. McCutcheon M. Crabtree G.R. Herzenberg L.A. EMBO J. 1990; 9: 4425-4433Crossref PubMed Scopus (291) Google Scholar). oligonucleotides and anti-Fos and anti-Jun and were of and containing oligonucleotides or were for the of of oligonucleotides with The for NFAT was upon the NFAT recognition site in the promoter for IL-2 et al., E.A. Verweij C.L. Durand D.B. E. Crabtree G.R. Science. 1989; PubMed Scopus Google Scholar). was for the binding assay by the oligonucleotides and in the presence of The for NFκB, and were as oligonucleotides with the and These oligonucleotides were with as a the binding were in for The were and NFAT activity in the Jurkat T-cell line has been (Shaw et al., 1988Shaw J. Utz P.J. Durand D.B. Toole J.J. Emmel E.A. Crabtree G.R. Science. 1988; 241: 202-205Crossref PubMed Scopus (10) Google Scholar; et al., 1988Shaw J. Utz P.J. Durand D.B. Toole J.J. Emmel E.A. Crabtree G.R. Science. 1988; 241: 202-205Crossref PubMed Scopus (10) Google Scholar; Emmel et al., E.A. Verweij C.L. Durand D.B. E. Crabtree G.R. Science. 1989; PubMed Scopus Google Clipstone and Crabtree, 1992Clipstone N. Crabtree G.R. Nature. 1992; 357: 695-697Crossref PubMed Scopus (1477) Google Scholar), we used this cell line as a in the present whether NFAT binding activity is present in activated mast cells, we RBL-2H3 cells and either through the FcϵRI or with PMA and ionomycin. show that in NFAT binding activity was induced in cells and a with PMA and and and also a stimulation via the Fcϵ receptor with the and stimulation the appearance of a that to a similar to that of the induced in Jurkat cells treated with PMA and ionomycin. through the Fcϵ receptor induced NFAT activity, with that induced by the cells with PMA and determine the specificity of the NFAT DNA binding activity induced in cells, oligonucleotides containing recognition were in to the NFAT oligonucleotide the NFAT DNA binding activity present in the nuclear from and Jurkat cells the presence of NFκB, and oligonucleotides the NFAT binding activity in either or Jurkat nuclear the NFAT DNA binding activity induced in cells by stimulation via the FcϵRI was specific for the NFAT recognition motif. stimulation of cells via the FcϵRI induces expression of the AP-1 transcription complex and Razin, E. 1991; PubMed Google Scholar; et al., which in T-cells with to form the NFAT transcription we the that AP-1 part of this complex in mast cells. shown in AP-1 oligonucleotides the NFAT in and in Jurkat cells as as the NFAT oligonucleotides did present the same This is with the as with T-cells, AP-1 part of the NFAT complex in cells. further this we in the presence of to the AP-1 proteins Fos and Using nuclear proteins from cells through the FcϵRI that either the Fos or the Jun the NFAT binding activity and the appearance of a of mobility and used in the Fos and Jun the NFAT DNA binding activity induced by FcϵRI cross-linkage through the FcϵRI, NFAT binding activity was to and the activity for to A of NFAT binding activity be in cells. This activity did the increase in NFAT induced by cell activation. evidence that AP-1 proteins to the NFAT transcriptional complex present in cells, we the of appearance of AP-1 activity cell activation through the that AP-1 activity in nuclear in a similar as did NFAT, AP-1 binding activity was present to activation. A induction was by a increase in AP-1 activity to The of AP-1 that of the NFAT the of used in the AP-1 this suggests that the of appearance of AP-1 does the of appearance of NFAT DNA binding activity. determine whether the appearance of NFAT in the nucleus of cells is cells were activated in the presence of EGTA to buffer calcium to cells were with the cell for conditions to block the calcium rise by FcϵRI A. The with EGTA did by NFAT binding activity the present in the cells in medium and cells in the presence of for to the and for the did the appearance of NFAT the of the and of cells with did the induction of NFAT binding activity by and cells with in the presence of EGTA induced NFAT binding activity, that induced in the presence of external and of calcium influx with EGTA did NFAT induction as as did intracellular which the rise in intracellular calcium to release from internal stores and influx the intracellular release apparently can a NFAT a sustained induction influx of calcium. determine whether the calcium-dependent protein calcineurin is in the induction of NFAT binding activity in mast cells, we activated cells with in the presence of the calcineurin inhibitor CsA from to CsA and antigen-induced NFAT binding activity, whereas CsA the activity and blocked nuclear NFAT induction Using specific DNA binding as a activation of cells by cross-linkage of the Fcϵ receptor induced NFAT activity similar to that in Jurkat cells with PMA and NFAT was in nuclear antigenic with the activity for This is similar to the of appearance of nuclear NFAT activity in Jurkat T-cells with PMA and (Shaw et al., 1988Shaw J. Utz P.J. Durand D.B. Toole J.J. Emmel E.A. Crabtree G.R. Science. 1988; 241: 202-205Crossref PubMed Scopus (10) Google Scholar), the appearance of NFAT activity in T-cells with (McCaffrey et al., 1993bMcCaffrey P.G. Luo C. Kerppola T.K. Jain J. Badalian T.M. Ho A.M. Burgeon E. Lane W.S. Lambert J.N. Curran T. Verdine G.L. Rao A. Hogan P.G. Science. 1993; 262: 750-754Crossref PubMed Scopus (379) Google Scholar). cells a of mast cells, the possibility that the presence of NFAT activity in these cells is an of the this we also a mast cell from rat bone These cells nuclear NFAT activity inducible under the same activation used with cells in with et al., 1988Shaw J. Utz P.J. Durand D.B. Toole J.J. Emmel E.A. Crabtree G.R. Science. 1988; 241: 202-205Crossref PubMed Scopus (10) Google Scholar), NFAT activity was in nuclear from cells. This activity cells were from the flasks and was dependent upon the presence of calcium. Nuclear from cells with and to medium that did NFAT DNA binding activity. NFAT activity to a cells were in medium for to antigenic have in intracellular in cells to an that calcium influx cell may be responsible for NFAT activity directly that the induction of NFAT activity by cross-linkage of the FcϵRI is to an of cytosolic of the the by buffering external with EGTA the NFAT activity induced by of cells with under conditions found to block the antigen-induced calcium inhibited induction of NFAT activity by the appearance of NFAT DNA binding activity in the nucleus of cells be part of a signaling pathway like that to mediate the translocation of in T-cells. some nuclear NFAT activity influx of was suggests that may be from intracellular stores to of the active NFAT complex that sustained activity role does calcium play in the induction of mast cell NFAT The of a potent calcineurin inhibitor, to block NFAT induction suggests that as with T-cells, calcium-dependent activation of this phosphatase is a key step in activation of NFAT via the NFAT activity in cells was by CsA a of and was by CsA and was blocked This is to the for CsA of NFAT activity with Jurkat cells with PMA and ionomycin. Emmel et al., E.A. Verweij C.L. Durand D.B. E. Crabtree G.R. Science. 1989; PubMed Scopus Google Scholar found that CsA of and inhibited the induction of NFAT activity in Jurkat cells, and activity was present CsA. in Jurkat cells, Mattila et al., 1990Mattila P.S. Ullman K.S. Fiering S. Emmel E.A. McCutcheon M. Crabtree G.R. Herzenberg L.A. EMBO J. 1990; 9: 4425-4433Crossref PubMed Scopus (291) Google Scholar found of NFAT binding activity with a in NFAT binding with and CsA. the of CsA for inhibiting NFAT induction in cells is similar to that found in Jurkat cells. mouse bone marrow-derived mast cells in IL-3, CsA has been shown to antigen-induced of and IL-6 mRNA of and (Kaye et al., 1992Kaye R.E. Fruman D.A. Bierer B.E. Albers M.W. Zydowsky L.D. Ho S.I. Jin Y. Castells M.C. Schreiber S.L. Walsh C.T. Burakoff S.J. Austen K.F. Katz H.R. Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 8542-8546Crossref PubMed Scopus (62) Google Scholar). results that CsA inhibits the induction of NFAT activity a similar suggests that NFAT may play a role in the transcriptional activation of these genes in mast cells. AP-1 and NFAT activity in the nucleus with a similar antigenic stimulation of cells A and An NFAT nuclear complex in T-cells is thought to in part upon the that AP-1 oligonucleotides binding of this complex to the NFAT recognition whereas the has binding of AP-1 to the AP-1 recognition motif (Jain et al., 1992Jain J. McCaffrey P.G. Valge-Archer V.E. Rao A. Nature. 1992; 356: 801-804Crossref PubMed Scopus (429) Google Scholar; Northrop et al., 1993Northrop J.P. Ullman K.S. Crabtree G.R. J. Biol. Chem. 1993; 268: 2917-2923Abstract Full Text PDF PubMed Google Scholar). The with cells suggest that the NFAT complex also contains AP-1 in bind to the NFAT recognition motif et al., 1990Ullman K.S. Northrop J.P. Verweij C.L. Crabtree G.R. Annu. Rev. Immunol. 1990; 8: 421-452Crossref PubMed Scopus (491) Google Scholar; Jain et al., 1992Jain J. McCaffrey P.G. Valge-Archer V.E. Rao A. Nature. 1992; 356: 801-804Crossref PubMed Scopus (429) Google Scholar). we show that to the AP-1 proteins and in block NFAT DNA evidence as in T-cells, these proteins to the mast cell NFAT complex a of et al., C.L. C. Crabtree G.R. J. Biol. Chem. 1990; Full Text PDF PubMed Google Scholar), transcription by the NFAT site in the IL-2 promoter to and and to a of cells in the mast cells DNA binding activity specific for this site in also was in of Verweij et al., C.L. C. Crabtree G.R. J. Biol. Chem. 1990; Full Text PDF PubMed Google Scholar and et al., S. J. Biol. Chem. 1993; 268: Full Text PDF PubMed Google in the transcriptional activity was Given that NFAT activity, and that other were as were the T-cells used for the specificity of other cell types may as found that stimulation via relevant or with PMA and ionomycin, induces NFAT activity. recent that assembly of the NFAT is to more and more McCaffrey et al., 1993bMcCaffrey P.G. Luo C. Kerppola T.K. Jain J. Badalian T.M. Ho A.M. Burgeon E. Lane W.S. Lambert J.N. Curran T. Verdine G.L. Rao A. Hogan P.G. Science. 1993; 262: 750-754Crossref PubMed Scopus (379) Google Scholar a to the preexisting cytoplasmic component NFATp from T-cells. A protein from this was shown to bind directly the NFAT recognition motif from the IL-2 to with Fos and Jun proteins to form and to be constitutively in T-cells (McCaffrey et al., 1993bMcCaffrey P.G. Luo C. Kerppola T.K. Jain J. Badalian T.M. Ho A.M. Burgeon E. Lane W.S. Lambert J.N. Curran T. Verdine G.L. Rao A. Hogan P.G. Science. 1993; 262: 750-754Crossref PubMed Scopus (379) Google Scholar). to be of this A similar with T-cells a to the for NFAT Ullman et al., 1990Ullman K.S. Northrop J.P. Verweij C.L. Crabtree G.R. Annu. Rev. Immunol. 1990; 8: 421-452Crossref PubMed Scopus (491) Google Scholar a gene cytosolic which was from NFATp in were or in T-cells and NFATp were present PMA and induced transcription of NFATp et al., J.P. Ho L.A. A. Crabtree G.R. Nature. 1994; PubMed Scopus Google Scholar). be to determine which of the and inducible cytosolic in nuclear NFAT assembly in mast cells. The present results that two types of mast cells, RBL-2H3 cells and rat bone mast cells, in to NFAT DNA binding activity. with the role of AP-1 and the subunit composition of the mast cell NFAT complex key for cells a for of NFAT in that can be more through a relevant pathway, can T-cells. may whether mast cell NFAT the transcription of one or more cytokine genes in to FcϵRI