COOPERATION OF BINDING-SITES FOR STAT6 AND NF-KAPPA-B REL IN THE IL-4-INDUCED UP-REGULATION OF THE HUMAN IGE GERMLINE PROMOTER/

Ig heavy chain class switching is directed by cytokines inducing trans cription from unreaarranged C-H genes. Subsequently, such primed cells can undergo switch recombination to express the selected new isotype, In the case of IgE class switching, IL-4 activates the IgE germline p romoter by inducing the interaction of the transcription factor STAT6 (IL-4STAT) with a responsive DNA element in the proximal region of the promoter, This study describes the characterization of two additional cis-acting elements that interact with members of the NF kappa B/rel transcription factor family in an IL-4-independent fashion, Electropho retic mobility shift assays show that the nucleoprotein complex formed an the upstream site (NF kappa B1) contains the classical F50/p65 het erodimer. The complex on the proximal site (NF kappa B2) appears to be composed Of p50 and relB. IgE germline promoter reporter gene constru cts carrying point mutations in the NF kappa 2 site were largely unres ponsive to IL-4 stimulation in transient transfection experiments, whi le plasmids with similar mutations in the NF kappa B1 site responded t o cytokine stimulation better than the wild-type promoter, The NF kapp a B2 effect was dependent on the presence of the STAT6 binding site, d emonstrating that the NF kappa B2 motif is necessary but not sufficien t for mediating cytokine up-regulation, in addition, the combination o f a NF kappa B/rel binding site and the STAT6 response element conferr ed IL-4 inducibility to a heterologous minimal promoter, while the ind ividual sites had no effect. The available data suggest that the NF ka ppa B2 nucleoprotein complex may cooperate with DNA-bound STAT6 to ach ieve IL-4-dependent activation of the human IgE germline gene.