DOWN-REGULATION OF NF-KAPPA-B PROTEIN-LEVELS IN ACTIVATED HUMAN-LYMPHOCYTES BY 1,25-DIHYDROXYVITAMIN-D-3

The effect of 1,25-dihydroxyvitamin D-3 [1,25(OH)(2)D-3], a steroid hormone with immunomodulating properties, on nuclear factor kappa B (NF-kappa B) proteins was examined in in vitro activated normal human lymphocytes by Western blot analysis. Over a 72-hr period of activation, the expression of the 50-kDa NF-kappa B, p50, and its precursor, p105, was increased progressively. When cells were activated in the presence of 1,25(OH)(2)D-3, the levels of the mature protein as well as its precursor were decreased. The effect of the hormone on the levels of p50 was demonstrable in the cytosolic and nuclear compartments; it required between 4 and 8 hr and was specific, as 25-hydroxyvitamin D-3 and 24,25-dihydroxyvitamin D-3 were ineffective, Besides p50, 1,25(OH)(2)D-3 decreased the levels of another NF-kappa B protein, namely c-rel. In addition, 1,25(OH)(2)D-3 decreased the abundance of a specific DNA-protein complex formed upon incubation of nuclear extracts from activated lymphocytes with a labeled NF-kappa B DNA binding motif. Further, 1,25(OH)(2)D-3 inhibited the transcriptional activity of NF-kappa B in Jurkat cells transiently transfected with a construct containing four tandem repeats of the NF-kappa B binding sequence of the immunoglobulin kappa light chain gene linked to the chloramphenicol acetyltransferase reporter gene, These observations demonstrate directly that there is de novo synthesis of NF-kappa B during human lymphocyte activation and suggest that this process is hormonally regulated.