Activating signal cointegrator 1 (ASC-1) harbors an autonomous transactivation domain that contains a putative zinc finger motif which provides binding sites for basal transcription factors TBP and TFIIA, transcription integrators steroid receptor coactivator 1 (SRC-1) and CBP-p300, and nuclear receptors, as demonstrated by the glutathione S-transferase pull-down assays and the yeast two-hybrid tests. The ASC-1 binding sites involve the hinge domain but not the C-terminal AF2 core domain of nuclear receptors. Nonetheless, ASC-1 appears to require the AF2-dependent factors to function (i.e., CBP-p300 and SRC-1), as suggested by the ability of ASC-1 to coactivate nuclear receptors, either alone or in cooperation with SRC-1 and p300, as well as its inability to coactivate a mutant receptor lacking the AF2 core domain. By using indirect immunofluorescence, we further show that ASC-1, a nuclear protein, is localized to the cytoplasm under conditions of serum deprivation but is retained in the nucleus when it is serum starved in the presence of ligand or coexpressed CBP or SRC-1. These results suggest that ASC-1 is a novel coactivator molecule of nuclear receptors which functions in conjunction with CBP-p300 and SRC-1 and may play an important role in establishing distinct coactivator complexes under different cellular conditions.The nuclear receptor superfamily is a group of ligand-dependent transcriptional regulatory proteins that function by binding to specific DNA sequences named hormone response elements in the promoters of target genes (reviewed in reference 36). The superfamily includes receptors for a variety of small hydrophobic ligands, such as steroids, T3, and retinoids, as well as a large number of related proteins that do not have known ligands, referred to as orphan nuclear receptors. Functional analysis of nuclear receptors has shown that there are two major activation domains. The N-terminal domain (AF1) contains a ligand-independent activation function, whereas the extreme C-terminal helix of the ligand binding domain (LBD) serves as an integral component of the ligand-dependent transactivation function AF2, which also includes several essential helixes in broad parts of the LBD. This C-terminal AF2 core region, which is relatively highly conserved among nuclear receptors, undergoes an allosteric change upon ligand binding, and deletion or point mutations in this region often impair transcriptional activation without changing ligand and DNA binding affinities (36).Transcriptional activation of nuclear receptors appears to involve at least two separate processes: derepression and activation. Repression is mediated in part by the interaction of unliganded receptors with corepressors such as the nuclear receptor corepressor (N-CoR) (8) and SMRT (20). However, ligand binding triggers the dissociation of these corepressors and the concomitant recruitment of coactivators. These putative receptor-interacting coactivators include RIP-140 and RIP-160 (5, 6), ERAP-140 and ERAP-160 (17), TIF1 (28), TRIP1 (30), ...