964 Nucleic Acids Research, 2006, Vol. 34, No. 3 A proﬁles/patterns analysis of YKR064W revealed the pres- ence of a Zn[2]-Cys[6] fungal-type binuclear cluster domain in the N-terminal region. This domain binds to DNA and is also found in ArgR2p, a component of the ARGR transcription regulatory complex (ArgR1p, ArgR2p, ArgR3p, Mcm1p) (this domain already lead to the annotation as transcription factor in the YPD database) (52). A search in the eMOTIF database (53) reveals a ‘fungal transcriptional regulatory protein’ motif (the eMotif search tool is also available in SGD). Together with the strong transcriptional activation properties of YKR064W, this ﬁnding indicates a true transcriptional regulation and activator function of YKR064W relying on its DNA binding as well as its AD, probably involving the ARGR transcription regu- latory complex. YCR082w (¼Ahc2) is a protein localized in the nucleus as well as in the cytoplasm without known function. It was identiﬁed as a strong activator in this study and is known to interact with chromatin reorganization components like a histone acetyltransferase complex (Ahc1) (26) and Abf1, a DNA-binding protein involved in chromatin reorganization (54). In addition, YCR082w interacts with a transcription factor, Srb4, a subunit of the RNA polymerase II mediator complex (27). DISCUSSION Transcriptional activation is the basis of the Y2H system (26,27). However, some bait proteins activate transcription without requiring an interacting protein that bears a separate AD. In fact, this property prevents the study of many TRs by means of the two-hybrid system. Nevertheless, we realized that this observation can extend our knowledge about the prop- erties of transcriptional ADs. In the course of this study we took previously identiﬁed (but mostly unpublished) activators from several Y2H screens (26,27) and measured their activation strength. In addition, we looked for properties distinguishing this set from non-activator proteins. Although many well-characterized TRs and nuclear proteins are highly overrepresented in the set of Y2H activ- ators, we identiﬁed many which have not been associated with transcription previously. Irrespective of their physiological role, these activators must be able to interact with and recruit the transcriptional machinery. Which features or sequences mediate the recruitment of the general transcriptional machinery to the activator? Although there is no single feature several rather general prop- erties of transcription activators have been deﬁned previously. These included the protein sequence composition as in acidic activators (25) or more speciﬁc features such as deﬁned interactions between activators and the basal transcription machinery [e.g. (21)]. Acidic activators, i.e. activators with stretches of acidic amino acids, were the ﬁrst class to be identiﬁed in yeast Figure 5. Protein–protein interactions of transcription activators. (A) This protein interaction network shows the activators (red) and the proteins inter- acting with at least two activators. Proteins involved in transcription are indicated by a diamond shape and the activation strength (bGal and LTH combined) is reflected by the node size. Individual protein names can be identified in the online version of this figure. (B) Protein–protein interactions of activators and control groups with other proteins involved in transcription (P-GO term ‘transcription’). Physical, binary protein interaction data from the MIPS database (35) were used. The right three columns include indirect inter- actions involving a bridging protein (indicated by ‘+1’) between the activator and the transcription protein. (C) Percentage of proteins interacting with com- ponents of protein complexes involved in transcription. Only high-throughput derived protein complex data from the MIPS database were considered.