Induction of ref-1 ensures AP-1 activation in intracellular oxidative environment of IL-2-stimulated BA/F3beta cells.
Our previous study of interleukin-2 (IL-2) signaling found that redox factor-1 (Ref-1) mRNA was upregulated by IL-2. In this study, we further studied the function of Ref-1 in the potential redox regulation of IL-2 signaling in BA/F3beta cells. Western blot analysis confirmed that IL-2 stimulation increases Ref-1 protein. Flow cytometric assay by using 2',7'-dichlorofluorescin diacetate indicated that IL-2 stimulation results in an oxidative shift of intracellular environment. However, IL-2-induced activator protein-1 (AP-1) is oxidation-sensitive. Gel shift assays of nuclear extracts immunodepleted of Ref-1 protein demonstrated that IL-2-induced AP-1 DNA binding is dependent on the presence of Ref-1. This was further confirmed by the restoration of AP-1 DNA binding upon the re-addition of immunoprecipitated Ref-1. Additionally, reporter gene assays showed that AP-1 transcriptional activity was enhanced by the overexpression of Ref-1 and attenuated by the introduction of antisense Ref-1. These results suggest that the induction of Ref-1 ensures AP-1 activation in the intracellular oxidative environment of IL-2-stimulated BA/F3beta cells.
Yan
M
M
Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 320 Yue Yang Road, Shanghai, 200031, China.
Xu
W
W
Lu
L
L
Sun
L
L
Liu
X
X
Zheng
Z
Z
D001483:Base Sequence; D019757:Carbon-Oxygen Lyases / Q000096:biosynthesis*; D002460:Cell Line; D017931:DNA Primers; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D007376:Interleukin-2 / Q000494:pharmacology*; D018384:Oxidative Stress; D011485:Protein Binding; D018808:Transcription Factor AP-1 / Q000378:metabolism*
D017931:DNA Primers; D007376:Interleukin-2; D018808:Transcription Factor AP-1; D019757:Carbon-Oxygen Lyases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase