Fos and Jun form a tight heterodimeric complex that activates transcription by AP1 sites. We have recognized that two adjacent regions of the Jun A1 activation domain are conserved in the Fos protein, and we refer to these two homologous regions as homology box 1 (HOB1) and homology box 2 (HOB2). Using GAIA chimeras, we show that the HOB1/HOB2 region of Fos and Jun is an independent activation domain in which HOB1 and HOB2 act cooperatively to activate transcription. This cooperativity is retained after the replacement of

Transcriptional activation in eukaryotes involves protein-protein interactions between regulatory transcription factors and components of the basal transcription machinery. Here we show that c-Fos, but not a related protein, Fra-1, can bind the TATA-box-binding protein (TBP) both in vitro and in vivo and that c-Fos can also interact with the transcription factor IID complex. High-affinity binding to TBP requires c-Fos activation modules which cooperate to activate transcription. One of these activation modules contains a TBP-binding motif (TBM) which was identified through its homology to TBP-binding viral The c-Fos oncoprotein is the product of an immediate-early response gene which, together with c-Jun, can activate transcription of promoters bearing AP-1-binding sites (28). The c-Fos and c-Jun proteins have a homologous bZIP DNAbinding domain which allows them to form a complex via the leucine zipper and bind DNA via the basic motif (12,18,27,35,45,46,53). In addition, c-Fos and c-Jun have a number of independently acting homologous activation domains (1,2,5,7,50). One of these contains two homologous motifs (HOB1 and HOB2) which cannot activate transcription independently but do so cooperatively when combined (50). The HOBW motif contains a recognition site for MAP kinase whose presence is essential for the activity of HOB1. In c-Jun, this site is phosphorylated by a MAP kinase-like activity both in vitro and in vivo (6, 21, 37), suggesting that phosphorylation of HOB1 is essential for its function.The c-Fos protein is a member of a family of transcription factors which are intermittently similar along their lengths. They include Fra-1, Fra-2, FosB, and FosBSF. These c-Fosrelated proteins can all bind to c-Jun and related proteins JunB and JunD, forming heterodimers which can bind to AP-1 sites. It is unclear how these c-Fos-c-Jun-related heterodimers differ in function. Discrimination may come at the level of DNA binding (44) Initiation of transcription by RNA polymerase II can be mediated by a set of basal factors which assemble at cis-acting elements such as the TATA box and the initiator (Inr) element (41). Initiation of transcription from either one of these elements can be augmented by regulatory transcription factors bound to the promoter. These activators are thought to function by forming protein-protein interactions with basal transcription factors (14). The study of viral transactivators such as VP16, ElA, Zta, and IE2 has shown that contact with the general factors TATA-box-binding protein (TBP) and transcription factor IIB (TFIIB) can be an important step in activation (9,17,23,24,30,31,40). Several cellular transcription factors, PU.1, p53, c-Myc, and E2F, have domains which bind TBP in vitro (15,16,19,49), and recently, the c-Rel protein has been shown to form a functional contact with TBP in vivo (26,54). Direct contact with TBP is not, however, the only mechanism by which regulators activate transcription. The Spl protein, for example, can associate with other components of the basal trans...

The c‐Fos protein has three activation modules at its C‐terminus, two of which contain motifs (HOB1 and HOB2) which are also present in the activation domains of c‐Jun. Here we show the existence of two additional activation modules at the N‐terminus of c‐Fos, one of which contains a second HOB1 motif (HOB1‐N). The N‐terminus also contains an inhibitor domain (ID1) which silences HOB1 activity. GAL4 fusion experiments showed that ID1 can specifically silence HOB1‐containing activation domains from c‐Fos or c‐Jun when linked in cis, but will not affect other distinct activation domains. The c‐Fos related protein, FosB, also contains an inhibitor domain. Mutagenic and deletion analyses identify an inhibitor motif (IM1) conserved between c‐Fos and FosB, which is required for inhibitor function. Mutagenesis of IM1 enhances the ability of c‐Fos to activate an AP1 bearing promoter. Finally, squelching experiments suggest that c‐Fos ID1 binds a limiting protein involved in inhibition. These results demonstrate the existence of a new class of inhibitor domain within transcriptional activators, which acts in a sequence specific manner to inhibit a subset of activation domains.

The c-Fos and c-Jun proteins bind an AP1 site and activate transcription synergistically. These two proteins have a common activation domain which has two co-operating motifs, HOB1 and HOB2. The HOB1 motif of c-Jun includes S73 which is required for Ha-Ras-induced super-activation and phosphorylation by MAP kinase-like enzymes. Since c-Fos HOB1 has a conserved Thr residue (T232) analogous to c-Jun S73 we have proposed that c-Fos HOB1 will be regulated in the same way as c-Jun HOB1. Here we show that the HOB1-containing activation domain of c-Fos is stimulated by Ha-Ras in vivo and phosphorylated by a MAP kinase family member in vitro and that mutating T232 to Ala abolishes both functions. Collectively these results suggest that phosphorylation of the HOB1 motif increases its activation capacity. To provide direct evidence for this we change the context of c-Fos T232 to a PKA recognition site, and show that HOB1 activity is now stimulated by the catalytic subunit of PKA. This 'PKA specificity' experiment represents a novel and powerful way to analyse phosphorylation events involved in a variety of biological functions.