A compendium of Caenhorabditis elegans regulatory transcription factors : a resource for mapping transcription regulatory networks

Background: Transcription regulatory networks are composed of interactions between transcription factors and their target genes. Whereas unicellular networks have been studied extensively, metazoan transcription regulatory networks remain largely unexplored. Caenorhabditis elegans provides a powerful model to study such metazoan networks because its genome is completely sequenced and many functional genomic tools are available. While C. elegans gene predictions have undergone continuous refinement, this is not true for the annotation of functional transcription factors. The comprehensive identification of transcription factors is essential for the systematic mapping of transcription regulatory networks because it enables the creation of physical transcription factor resources that can be used in assays to map interactions between transcription factors and their target genes. Results: By computational searches and extensive manual curation, we have identified a compendium of 934 transcription factor genes (referred to as wTF2.0). We find that manual curation drastically reduces the number of both false positive and false negative transcription factor predictions. We discuss how transcription factor splice variants and dimer formation may affect the total number of functional transcription factors. In contrast to mouse transcription factor genes, we find that C. elegans transcription factor genes do not undergo significantly more splicing than other genes. This difference may contribute to differences in organism complexity. We identify candidate redundant worm transcription factor genes and orthologous worm and human transcription factor pairs. Finally, we discuss how wTF2.0 can be used together with physical transcription factor clone resources to facilitate the systematic mapping of C. elegans transcription regulatory networks. Conclusion: wTF2.0 provides a starting point to decipher the transcription regulatory networks that control metazoan development and function. Published: 30 December 2005 Genome Biology 2005, 6:R110 (doi:10.1186/gb-2005-6-13-r110) Received: 26 September 2005 Revised: 7 November 2005 Accepted: 28 November 2005 The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2005/6/13/R110 Genome Biology 2005, 6:R110 R110.2 Genome Biology 2005, Volume 6, Issue 13, Article R110 Reece-Hoyes et al. http://genomebiology.com/2005/6/13/R110 Background Metazoan genomes contain thousands of predicted proteincoding genes. During development, pathology, and in response to environmental changes, each of these genes is expressed in different cells, at different times and at different levels. Spatial and temporal gene expression is controlled transcriptionally through the action of regulatory transcription factors (TFs) [1,2]. Transcription of each gene can be upor down-regulated by TFs that bind to cis-regulatory DNA elements. These elements include upstream elements located in the proximal promoter, and enhancers or silencers that can be located at a greater distance from the transcription start site. Frequently, the expression level of a gene is the result of a balance between transcription activation and repression governed by multiple cis-regulatory elements and, hence, multiple TFs. The combinatorial nature of gene transcription provides an exquisite level of flexibility to regulate genome expression. The understanding of differential gene expression at a genome-wide, or systems, level has been greatly facilitated by the mapping and analysis of transcription regulatory networks (Figure 1) [3-5]. Such networks are composed of two types of components, or nodes: the gene targets that are subject to transcriptional control and the TF proteins that execute transcriptional control. Whereas transcription regulatory networks have been extensively studied in relatively simple unicellular systems, they remain largely unexplored in complex, metazoan systems. The nematode Caenorhabditis elegans is a powerful model to decipher metazoan transcription regulatory networks. The C. elegans genome has been completely sequenced and is predicted to contain 19,735 protein-coding genes (WormBase WS140)[6]. Several functional genomic resources enable the systematic dissection of differential gene expression at a systems level and in a high-throughput manner. For instance, microarrays are available to investigate temporal and, to a certain extent, spatial gene expression levels [7-9]. In addition, C. elegans 'ORFeome' [10] and 'Promoterome' [11] resources provide open reading frame (ORF) and promoter clones, respectively. These clones can be used for a wide variety of experiments that aim to dissect transcription regulatory networks (see below). With seven years of progressive refinement of the genome annotation since the publication of the genome sequence [1214], comprehensive predictions of protein-coding genes are available. However, there is no up-to-date compendium of predicted C. elegans TFs. Several lists of putative C. elegans TFs have been generated previously (Table 1; we refer to this combined set as wTF1.0 (worm transcription factors version 1.0)), but none of these are comprehensive or readily accessible. The earliest lists were created by scanning sequences of C. elegans proteins, as predicted from the genome annotation, for well-defined DNA binding domains: Hobert and Ruvkun [15] focused on homeodomain, paired domain, T-box, basic helix-loop-helix (bHLH), basic region leucine zipper (bZIP), Fork Head, erythroblast transformation specific (ETS) and nuclear hormone receptor (NHR) proteins; and Clarke and Berg [16] focused on various zinc-finger proteins. For comparative genomic studies primarily focusing on Drosophila melanogaster [17] or Arabidopsis thaliana [18], a list of predicted C. elegans proteins containing various DNA binding domains was compiled. While several (sub)-families of C. elegans TFs have been studied in greater detail (for example, bHLH [19], CUT homeodomain [20], and DM zinc-finger [21]), the most recent list of "all C. elegans TFs" was compiled five years ago when Riechmann and colleagues [18] scanned WormPep 20 (19,101 proteins). During the past few years, the creation of improved computational tools [13] and the completion of the C. briggsae genome sequence [14] have enabled a great improvement in the annotation of the C. elegans genome. Here, we used a combination of bioinformatics and extensive manual curation to generate wTF2.0, a comprehensive compendium of predicted C. elegans TF genes. We discuss how wTF2.0 can be used together with physical ORFeome and Promoterome clone resources to decipher transcription regulatory networks that control metazoan differential gene expression at a systems level. Results and discussion TF predictions: Gene Ontology term-based searches To identify a comprehensive compendium of predicted worm TFs, we first interrogated WormBase version 140 (WS140)[6] for proteins that possess domains annotated with one of the following Gene Ontology (GO) terms: 'regulation of transcription, DNA-dependent', 'transcription factor activity', and 'DNA binding'. (WS140 is the most recent reference release of WormBase that is permanently accessible.) We identified a total of 930 proteins (Figure 2, Additional data file 1). Of these, 232 were identified by all three GO terms, 368 by two GO terms and 330 by only one GO term (Figure 3). We observed that this collection of proteins not only contains predicted regulatory TFs, but also proteins that function in other nuclear processes (for example, DNA replication and repair). Moreover, it contains numerous false positive predictions (for example, small GTP-binding proteins). We removed both types of false positives (Figure 2, Additional data file 2). Next, we examined each of the remaining proteins for the presence of a predicted DNA binding domain either by visual inspection of the protein sequence (AT-hooks and C2H2 zinc-fingers), or using InterPro v10.0 (2005)[22], SMART [23] and Pfam [24] databases (Additional data file 2). We found several proteins that, upon closer inspection, do not possess a DNA binding domain despite their WormBase protein domain annotation (Additional data file 2). For instance, several proteins were found that are annotated to be a NHR TF, but that only contain a predicted ligand binding domain. However, we retained proteins that do not have a clear DNA binding domain but for which experimental evidence is availGenome Biology 2005, 6:R110 http://genomebiology.com/2005/6/13/R110 Genome Biology 2005, Volume 6, Issue 13, Article R110 Reece-Hoyes et al. R110.3