Nucleic Acids Research, 2006, Vol. 34, No. 3 955–967 doi:10.1093/nar/gkj493 Transcriptional activators in yeast 1 Bjo¨ rn Titz, Sindhu Thomas, Seesandra V. Rajagopala, Tomoko Chiba , Takashi Ito1 and Peter Uetz* Institut fu¨ r Genetik, Forschungszentrum Karlsruhe, Box 3640, D-76021 Karlsruhe, Germany and 1Department of Computational Biology, Graduate School of Frontier Sciences, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa 277-8561, Japan Received September 15, 2005; Revised October 29, 2005; Accepted January 20, 2006 ABSTRACT Eukaryotic transcription activation domains (ADs) are not well defined on the proteome scale. We sys- tematicallly tested 6000 yeast proteins for trans- criptional activity using a yeast one-hybrid system and identified 451 transcriptional activators. We then determined their transcription activation strength using fusions to the Gal4 DNA-binding domain and a His3 reporter gene which contained a promoter with a Gal4-binding site. Among the 132 strongest activators 32 are known transcription factors while another 35 have no known function. Although zinc fingers, helix–loop–helix domains and several other domains are highly overrepresen- ted among the activators, only few contain character- ized ADs. We also found some striking correlations: the stronger the activation activity, the more acidic, glutamine-rich, proline-rich or asparagine-rich the activators were. About 29% of the activators have been found previously to specifically interact with the transcription machinery, while 10% are known to be components of transcription regulatory com- plexes. Based on their transcriptional activity, localization and interaction patterns, at least six previously uncharacterized proteins are suggested to be bona fide transcriptional regulators (namely YFL049W, YJR070C, YDR520C, YGL066W/Sgf73, YKR064W and YCR082W/Ahc2). INTRODUCTION Transcriptional regulators (TRs) that activate transcription are usually composed of a DNA-binding domain (DBD) and an activation domain (AD). The DBD targets these proteins to a speciﬁc binding site in the promoter or enhancer region of a gene and the AD mediates transcription initiation (1). Transcriptional activators in yeast were among the ﬁrst to be studied in detail. In all known cases they recruit additional proteins or whole complexes to the pertinent promoters, even- tually leading to the binding of one of the three RNA poly- merases. For example, the yeast transcription regulator Gal4 is involved in regulation of galactose metabolism and acti- vates transcription by recruitment of the basal transcription machinery (2–5). The Gal4–AD can recruit Tra1, a component of the SAGA complex, to the upstream activating sequence (UAS), where Gal4 itself is bound. SAGA, in turn, recruits the mediator complex to the UAS. The Gal4–AD can also directly recruit the mediator complex. In any case, the UAS bound mediator is required for recruitment of general transcription factors to the core promoter and assembly of the pre-initiation complex. While DBDs are extremely well characterized both func- tionally and structurally, ADs do not share easily recognizable motifs or structures (6). Accordingly, no speciﬁc pattern or motif for the identiﬁcation of an AD has been deﬁned in pat- tern/domain databases such as Prosite (7) or SMART (8). In contrast, >50 patterns for DBDs have been documented in the SMART database. Because the activation properties of transcriptional activat- ors cannot easily be recognized by sequence homology, several studies tried to identify more general sequence features resulting in a number of different AD classes (9), e.g. acidic activators (10), glutamine-rich activators (11) and proline-rich activators (12). In addition, a few rather unspeciﬁc properties like hydrophobic patches interspersed with hydrophilic residues (13) or amphipatic alpha helices (14) have been iden- tiﬁed. These analyses culminated in the ﬁnding that even small chemical compounds with a certain pattern of hydrophobic and hydrophilic residues are sufﬁcient for transcriptional activation (15). *To whom correspondence should be addressed. Tel: +49 7247 82 6103; Fax: +49 7247 82 3354; Email: peter.uetz@itg.fzk.de Correspondence may also be addressed to Takashi Ito. Tel: +81 4 7136 3989; Fax: +81 4 7136 3979; Email: ito@k.u-tokyo.ac.jp The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors The Author 2006. Published by Oxford University Press. All rights reserved. The online version of this article has been published under an open access model. 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