No abstract

Molecular cloning of the polypeptide component of the Rel-related human p75 nucleoprotein complex has revealed its identity with the 65-kDa (p65) subunit of NF-KB.Functional analyses of chimeric proteins composed of NF-KB p65 C-terminal sequences linked to the DNA-binding domain of the yeast GAL4 polypeptide have indicated that the rinal 101 amino acids of NF-#cB p65 comprise a potent transcriptional activation domain. Transient transfection of human T cells with an expression vector encoding NF-KB p65, but not NF-tcB p50, produced marked transcriptional activation of a basal promoter containing duplicated KB enhancer motifs from the long terminal repeat of type 1 human immunodeficiency virus.These stimulatory effects of NF-acB p65 were synergistically enhanced by coexpression of NF-ucB p50 but were completely inhibited by coexpression of the v-rel oncogene product. Together, these functional studies demonstrate that NF-KB p65 is a transactivating subunit of the heterodimeric NF-mcB complex and serves as one cellular target for v-Rel-mediated transcriptional repression.The NF-KB transcription factor complex plays a central role in the activation of type 1 human immunodeficiency virus (HIV-1) and many immunologically relevant cellular genes (1). Nuclear NF-KB DNA-binding activity is constitutively expressed in mature B cells (2) and induced in T cells by phorbol esters (3, 4), tumor necrosis factor a (5, 6), and the Tax protein of the type I human T-cell leukemia virus (HTLV-I) (7-9). Nuclear NF-KB expression is regulated in part at a posttranslational level (3) involving its dissociation from the cytoplasmic inhibitor IKB (10, 11). This dissociation reaction is apparently catalyzed by the phosphorylation of IKB, which in turn allows the rapid translocation of NF-KB to the nucleus (10-12).Sequence analyses of human and murine cDNAs encoding the DNA-binding subunit (NF-KB p50) (13,14) and the IKB-binding subunit (NF-KB p65) (15, 16) of NF-KB have revealed extensive homology between the N-terminal 300 amino acids of these two polypeptides, the v-Rel oncoprotein, and the dorsal gene product, a ventral morphogen in Drosophila (17). v-Rel specifically engages the KB enhancer and acts as a dominant negative repressor of NF-KB in mature T cells (18)(19)(20). However, the molecular basis for the transcriptional activation mediated by the NF-KB p5O/p65 complex or transcriptional repression mediated by v-Rel has remained unknown.Although gel retardation analyses with purified NF-KB originally suggested that only NF-KB p50 had intrinsic DNAbinding activity (21,22), recent DNA/protein crosslinking studies with human T cell extracts have demonstrated four Rel-related KB-specific proteins that are expressed in a temporally biphasic manner (pS5/p75, early; pSO/p85, late) (7, 18, 23). The p55 and p50 adducts contain NF-KB p50 and a closely related protein, respectively (18), while the p85 adduct is composed of the c-rel protooncogene product (c-Rel) (18), which has been shown to contain a functional transactivation domai...