Costimulation of peripheral blood T cell activation by human endothelial cells. Enhanced IL-2 transcription correlates with increased c-fos synthesis and increased Fos content of AP-1.
Endothelial cells (EC) act as APC for resting PBL in vitro, and may have important roles in vivo in the pathogenesis of allograft rejection and delayed hypersensitivity. We previously reported that human umbilical vein EC provide costimulatory signals to PHA-stimulated PBL via CD2:lymphocyte function-associated Ag-3 and an unidentified ligand pair, resulting in a three- to eight-fold enhancement of IL-2 production. The physiologic relevance of this increase was demonstrated by the proliferative advantage provided by EC to PBL suboptimally stimulated with mAb OKT3. We now report that EC costimulation causes increased levels of IL-2 mRNA as a result of increased IL-2 transcription in PBL. We therefore examined the effects of EC on T cell nuclear factors known to regulate IL-2 transcription, including c-jun and c-fos-two components of the transcription factor AP-1, NFAT, and others. PBL constitutively express c-jun transcripts, and the level of c-jun mRNA is not altered by PHA activation in the absence or presence of EC. In contrast, c-fos mRNA is absent from resting T cells and is induced on PHA activation. EC alone do not induce c-fos mRNA but augment the level of c-fos mRNA in PHA-activated T cells by 3- to 10-fold. This effect is largely independent of the CD2:lymphocyte function-associated Ag-3 pathway. Gel-shift analysis reveals the constitutive presence of nuclear factors in resting PBL that bind to the proximal AP-1 site of the IL-2 promoter and that contain immunoreactive c-Jun but not c-Fos protein. In contrast, AP-1 from PHA-activated cells contains c-Jun and low levels of c-Fos. Strikingly, costimulation with EC results in a dramatic increase (up to 15-fold) in the c-Fos content of AP-1. Levels of other nuclear factors involved in IL-2 regulation were not altered by EC, although NFAT-DNA complexes migrated at a slightly different mobility. In summary, our data suggest that changes in the composition of transcription factor AP-1 is a key molecular mechanism for increasing IL-2 transcription and may underlie the phenomenon of costimulation by EC.
Hughes
C C
CC
Molecular Cardiobiology Program, Boyer Center for Molecular Medicine, Yale University School of Medicine, New Haven, CT 06536.
Pober
J S
JS
D001483:Base Sequence; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology*; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics*; D008213:Lymphocyte Activation; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D016760:Proto-Oncogene Proteins c-fos / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D016755:Proto-Oncogene Proteins c-jun / Q000032:analysis* / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D014158:Transcription, Genetic
D007376:Interleukin-2; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D012333:RNA, Messenger