Yeast SUG1 was originally characterized as a transcriptional mediator for the GAL4 transactivator. A similar role in vertebrates was suggested by the ligandenhanced interaction between mammalian homologues of yeast SUG1 and the ligand-dependent activating domain (AF-2) of nuclear receptors. SUG1 was also shown to be a component of the PA700 regulatory complex of the 26 S proteasome and a member of a large family of putative ATPases. However, no catalytic function has yet been attributed to SUG1. We show here that SUG1 is a 3-5 DNA helicase whose activity is dependent on an intact ATP binding domain. The sedimentation heterogeneity of mammalian SUG1 suggests that it may be associated with distinct protein complexes and therefore play multiple roles.Studies in transcription interference/squelching indicate that enhancement of transcription by nuclear receptors may require protein mediators (co-activators) linking ligand-activated nuclear receptors to the transcription machinery at promoters of target genes (1-3). Several putative transcription intermediary factors interacting in a ligand-dependent manner with the region containing the ligand-dependent transcription activation function-2 (AF-2) of nuclear receptors have been described recently (for review, see Ref. 4). Ligand-enhanced interaction between the AF-2 domain of nuclear receptors and either mouse (m)SUG1 (5) or its human homologue TRIP1 (6, 7) has suggested that they may act as mediators in transcription activated by nuclear receptors. Moreover, yeast (y)SUG1, has been shown to suppress the effect of a mutation in the transcriptional activator GAL4 (8,9). Functional similarity between yeast and mammalian SUG1 is supported by the ability of mSUG1 to rescue a conditional SUG1 mutation in yeast complementation experiments (5). Yeast SUG1 was also found in purified RNA polymerase II (pol II) 1 holoenzyme complexes responsive in vitro to the transcriptional activators GAL-VP16 and GCN4 (10). However, this association is controversial as SUG1 does not appear to be present in the yeast holoenzyme preparation of Rubin et al. (11).The sequence similarity between mSUG1, ySUG1, and TRIP1 is the highest in a region containing the consensus ATP binding site motif. This motif corresponds to an AAA module recently described for a large family of putative ATPases involved in a variety of cellular processes (for review, see Ref. 12). For instance, MSS1 and TBP1, which were originally identified as transcription factors, were later shown to be members of the PA700 proteasome regulatory complex of the 26 S proteasome (13-17). Similarly, Kominami et al. (18) and then Rubin et al. (11) found SUG1 in the yeast 26 S proteasome. This agrees with genetic evidence showing that yeast cells harboring a mutant allele of ySUG1 accumulate ubiquitinylated proteins normally degraded by 26 S proteasomes (19). A role for SUG1 in the regulation of the activity of the 26 S proteasome in vivo is further supported by the identical amino acid sequence of mSUG1 with the p45 subunit of the ...

Mutations in the basal transcription initiation/DNA repair factor TFIIH are responsible for three human disorders: xeroderma pigmentosum (XP), cockayne syndrome (CS) and trichothiodystrophy (TTD). The non-repair features of CS and TTD are thought to be due to a partial inactivation of the transcription function of the complex. To search for proteins whose interaction with TFIIH subunits is disturbed by mutations in patients we used the yeast two-hybrid system and report the isolation of a novel XPB interacting protein, SUG1. The interaction was validated in vivo and in vitro in the following manner. (i) SUG1 interacts with XPB but not with the other core TFIIH subunits in the two-hybrid assay. (ii) Physical interaction is observed in a baculovirus co-expression system. (iii) In fibroblasts under non-overexpression conditions a portion of SUG1 is bound to the TFIIH holocomplex as deduced from co-purification, immunopurification and nickel-chelate affinity chromatography using functional tagged TFIIH. Furthermore, overexpression of SUG1 in normal fibroblasts induced arrest of transcription and a chromatin collapse in vivo. Interestingly, the interaction was diminished with a mutant form of XPB, thus providing a potential link with the clinical features of XP-B patients. Since SUG1 is an integral component of the 26S proteasome and may be part of the mediator, our findings disclose a SUG1-dependent link between TFIIH and the cellular machinery involved in protein modelling/degradation.