BACKGROUND: The nuclear factor of activated T cells (NFAT) family of transcription factors plays a key role in rapidly inducing IL4 gene expression in effector T cells. OBJECTIVE: Because human basophils secrete high levels of IL-4, we have examined whether specific NFAT species are expressed in these cells and whether Fc(epsilon)RI-mediated activation affects their subcellular localization and transcriptional function. METHODS: Intracellular NFAT protein was identified by using 2-color flow cytometry; gene expression was done with RT-PCR. Subcellular localization of NFAT was assessed by means of Western blotting. Electrophoretic mobility shift assays assessed NFAT involvement in IL-4 transcription. RESULTS: Basophils constitutively expressed high levels of NFAT2. In contrast, NFAT1 (NFATp), which is found in most leukocytes, was not seen in basophils. Low-level staining for NFAT4 was detected but was variably expressed among donor cells. Likewise, NFAT2 mRNA was constitutively expressed in basophils, and message for NFAT4 was seen in 3 of 5 preparations, whereas that for NFAT1 was found in only 1 of 5 preparations. NFAT2 protein accumulated in the nuclei of basophils activated for 1 hour with anti-IgE, and this was inhibited with the addition of FK506. A protein-DNA complex was formed with nuclear lysates from basophils and an IL-4 promoter NFAT consensus probe, with greater binding intensities detected in lysates of activated cells. An antibody to NFAT2 reduced the formation of the complex, whereas no effects were seen with antibodies to NFAT1, NFAT4, or unrelated transcription factors. CONCLUSIONS: The selective and specific expression of NFAT2 in basophils is unique among leukocytes. This transcription factor also appears to play a critical role in the Fc(epsilon)RI-mediated production of IL-4 in these cells.
BACKGROUND: The nuclear factor of activated T cells (NFAT) family of transcription factors plays a key role in rapidly inducing IL4 gene expression in effector T cells. OBJECTIVE: Because human basophils secrete high levels of IL-4, we have examined whether specific NFAT species are expressed in these cells and whether Fc(epsilon)RI-mediated activation affects their subcellular localization and transcriptional function. METHODS: Intracellular NFAT protein was identified by using 2-color flow cytometry; gene expression was done with RT-PCR. Subcellular localization of NFAT was assessed by means of Western blotting. Electrophoretic mobility shift assays assessed NFAT involvement in IL-4 transcription. RESULTS: Basophils constitutively expressed high levels of NFAT2. In contrast, NFAT1 (NFATp), which is found in most leukocytes, was not seen in basophils. Low-level staining for NFAT4 was detected but was variably expressed among donor cells. Likewise, NFAT2 mRNA was constitutively expressed in basophils, and message for NFAT4 was seen in 3 of 5 preparations, whereas that for NFAT1 was found in only 1 of 5 preparations. NFAT2 protein accumulated in the nuclei of basophils activated for 1 hour with anti-IgE, and this was inhibited with the addition of FK506. A protein-DNA complex was formed with nuclear lysates from basophils and an IL-4 promoter NFAT consensus probe, with greater binding intensities detected in lysates of activated cells. An antibody to NFAT2 reduced the formation of the complex, whereas no effects were seen with antibodies to NFAT1, NFAT4, or unrelated transcription factors. CONCLUSIONS: The selective and specific expression of NFAT2 in basophils is unique among leukocytes. This transcription factor also appears to play a critical role in the Fc(epsilon)RI-mediated production of IL-4 in these cells.
Authors: Trong Le; Jody Tversky; Kristin L Chichester; Anja P Bieneman; Shau-Ku Huang; Robert A Wood; John T Schroeder Journal: J Allergy Clin Immunol Date: 2008-10-09 Impact factor: 10.793
Authors: Houda Z Elloumi; Nitsan Maharshak; Kavitha N Rao; Taku Kobayashi; Hyungjin S Ryu; Marcus Mühlbauer; Fengling Li; Christian Jobin; Scott E Plevy Journal: PLoS One Date: 2012-03-27 Impact factor: 3.240