CD28 mediates transcriptional upregulation of the interleukin-2 (IL-2) promoter through a composite element containing the CD28RE and NF-IL-2B AP-1 sites.
Mutagenesis studies have demonstrated the requirement for the CD28-responsive element (CD28RE) within the interleukin-2 (IL-2) promoter for transcriptional upregulation by CD28. Here, we demonstrate that CD28 responsiveness is conferred by a composite element containing both the CD28RE and the NF-IL-2B AP-1 sites (RE/AP). Mutations at either site within the RE/AP composite element abolish activity. The RE/AP composite element is a site for signal integration within the IL-2 promoter, since its activation is dependent on at least two separate signalling pathways being activated, through the T-cell receptor, CD28, and/or phorbol myristate acetate. Activation is maximal when all three signals occur simultaneously. By using a panel of CD28 cytoplasmic domain mutants, it was found that the transcriptional activation of the RE/AP composite element correlates exactly with the pattern of IL-2 secretion induced by these mutants upon stimulation. Similar to the upregulation of IL-2 secretion, the transcriptional upregulation of the RE/AP composite element by CD28 is FK506 insensitive. The pattern of activation of the RE/AP composite element is different from that observed for either an NFAT or consensus AP-1 site, implying that RE/AP represents a unique element. Using gel shift analysis, we demonstrate that stimulation by CD28 induces the association of the NF-kappaB family member c-Rel to the CD28RE within the RE/AP composite element. The transcriptional upregulation of IL-2 by CD28 appears, therefore, to be mediated through the RE/AP composite element, involving the association of c-Rel with the CD28RE.
Shapiro
V S
VS
Department of Medicine, Howard Hughes Medical Institute, University of California, San Francisco 94143, USA.
Truitt
K E
KE
Imboden
J B
JB
Weiss
A
A
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D018106:CD28 Antigens / Q000502:physiology*; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000502:physiology; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics; D020569:Proto-Oncogene Proteins c-rel; D018808:Transcription Factor AP-1 / Q000502:physiology*; D014158:Transcription, Genetic; D015854:Up-Regulation
D018106:CD28 Antigens; D004268:DNA-Binding Proteins; D007376:Interleukin-2; D046911:Macromolecular Substances; D011518:Proto-Oncogene Proteins; D020569:Proto-Oncogene Proteins c-rel; D018808:Transcription Factor AP-1