Interleukin‐2 promoter activity in Epstein‐Barr virus‐transformed B lymphocytes is controlled by nuclear factor‐ϰB

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Abstract
The regulation of interleukin (IL)‐2 gene expression has been investigated mainly in T lymphocytes, the predominant producers of IL‐2. However, B cells can also synthesize IL‐2. In the present study we analyzed the control of IL‐2 promoter activity in Epstein‐Barr virus (EBV)‐transformed B cell clones which are capable of secreting IL‐2 at a low level after stimulation with phorbol 12‐myristate 13‐acetate and the Ca2+ ionophore ionomycin. Transient transfections using reporter constructs with multiples of transcription factor binding sites from the IL‐2 promoter [distal nuclear factor (NF)‐AT, proximal NF‐AT, AP‐1/Octamer (UPS) or NF‐ϰB (TCEd) sites] were performed. In EBV‐transformed B clones, the ϰB site exerted the strongest inducible activity; the NF‐AT binding sites showed either no or only weak activity compared to Jurkat T cells. An IL‐2 promoter bearing a defective NF‐ϰB site was completely inactive in EBV‐B cells, white it still had activity in Jurkat T cells. In seven EBV‐B cell clones or lines differing in their capacity to secrete IL‐2, the activity of the IL‐2 promoter correlated well with the status of IL‐2 secretion. Similarly, a human immunodeficiency virus promoter, whose activity is controlled through ϰB factors, was found to be active in the IL‐2‐producing EBV‐B cells, but inactive in the non‐IL‐2‐producing cells. Electrophoretic mobility shift assays using protein extracts from EBV‐B cells and the IL‐2 NF‐ϰB probe revealed the constitutive generation of ϰB complexes in IL‐2‐secreting cells consisting mainly of heterodimeric p50/p65 complexes. A weaker ϰB complex formation and faster‐migrating complexes were detected in non‐IL‐2‐secreting cells. These results demonstrate that the IL‐2 NF‐ϰB site is indispensable for the activity of the IL‐2 promoter in EBV‐transformed B cells, whereas other transcription factors appear to be less important for IL‐2 expression in these cells.