HCF-1 functions as a coactivator for herpes simplex virus VP16 and a number of mammalian transcription factors. Mature HCF-1 is composed of two subunits generated by proteolytic cleavage of a larger precursor at six centrally-located HCF PRO repeats. The resulting N-and Cterminal subunits remain tightly associated via two complementary pairs of self-association domains: termed SAS1N-SAS1C and SAS2N-SAS2C. Additional HCF proteins have been identified in mammals (HCF-2) and Caenorhabditis elegans (CeHCF). Both contain wellconserved SAS1 domains but do not undergo proteolytic processing. Thus, the significance of the cleavage and self-association of HCF-1 remains enigmatic. Here, we describe the isolation of the Drosophila HCF homologue (dHCF) using a genetic screen based on conservation of the SAS1 interaction. The N-terminal β-propeller domain of dHCF supports VP16-induced complex formation and is more similar to mammalian HCF-1 than other homologues. We show that fulllength dHCF expressed in Drosophila cells undergoes proteolytic cleavage giving rise to tightly associated N-and C-terminal subunits. As with HCF-1, the SAS1N and SAS1C elements of dHCF are separated by a large central region, however, this sequence lacks obvious homology to the HCF PRO repeats required for HCF-1 cleavage. The conservation of HCF processing in insect cells argues that formation of separate N-and C-terminal subunits is important for HCF function.HCF-1, also known as C1 factor, is a broadly-expressed nuclear protein involved in the regulation of cellular and viral gene expression. HCF-1 was first identified through its role in transcriptional activation by the herpes simplex virus (HSV) VP16 protein (Gerster and Roeder, 1988;Kristie et al., 1989). Association of HCF-1 with VP16 (also known as α-TIF or Vmw65) leads to the assembly of a multiprotein-DNA complex that includes the POU protein Oct-1 and the TAATGARAT element, a VP16-responsive DNA sequence present in all viral immediate-early (IE) gene promoters (reviewed in (Herr, 1998)). Nuclear localization of VP16 is dependent on association with HCF-1, which contributes a nuclear localization signal (NLS) located at its C-terminus (LaBoissière et al., 1999;Scarr et al., 2000;Wilson et al., 2000). In addition, HCF-1 contains its own activation domain and this is required for optimal transactivation by the VP16-induced complex ( The arrangement of functional domains in human HCF-1 is shown in Figure 1A. HCF-1 is translated as a 230-300-kDa precursor and processed into N-and C-terminal subunits through autocatalytic proteolytic cleavage at a series of six HCF PRO repeats located near the center of the precursor polypeptide (Wilson et al., 1993b;Kristie et al., 1995;Wilson et al., 1995b;Vogel and Kristie, 2000). The resulting subunits remain stably associated via two matched pairs of interaction domains: termed SAS1N-SAS1C and SAS2N-SAS2C . The SAS1C domain corresponds to a pair of fibronectin type 3 (Fn3) repeats, which interact with a 43-residue sequence (SAS1N) that is locate...
