The EH1 motif in metazoan transcription factors

Abstract

Background: The Engrailed Homology 1 (EH1) motif is a small region, believed to have evolved convergently in homeobox and forkhead containing proteins, that interacts with the Drosophila protein groucho (C. elegans unc-37, Human Transducin-like Enhancers of Split). The small size of the motif makes its reliable identification by computational means difficult. I have systematically searched the predicted proteomes of Drosophila, C. elegans and human for further instances of the motif.

Results: Using motif identification methods and database searching techniques, I delimit which homeobox and forkhead domain containing proteins also have likely EH1 motifs. I show that despite low database search scores, there is a significant association of the motif with transcription factor function. I further show that likely EH1 motifs are found in combination with T-Box, Zinc Finger and Doublesex domains as well as discussing other plausible candidate associations. I identify strong candidate EH1 motifs in basal metazoan phyla.

Conclusion: Candidate EH1 motifs exist in combination with a variety of transcription factor domains, suggesting that these proteins have repressor functions. The distribution of the EH1 motif is suggestive of convergent evolution, although in many cases, the motif has been conserved throughout bilaterian orthologs. Groucho mediated repression was established prior to the evolution of bilateria.

Figure 1

Alignments of putative EH1 motifs in a) Homeobox and b) Paired box domain containing proteins, subdivided by domain partners and orientation, with representative non-bilaterian sequences included. Alignments were derived from meme searches, as described in text. Conserved aromatic residues (FHYW) are coloured white on a red background ('a' in the consensus). Conserved aliphatic residues (ILV), black on a yellow background. ('I' in the consensus) Conserved big residues (EFHIKLMQRWY) blue on a light yellow background ('b' in the consensus). Conservation is calculated over the full alignment of sequences in figures 1 and 2. The figure was produced using the Chroma program [41]. Gene names are standard HUGO Gene Nommenclature Committee, flybase or wormbase symbols where available, otherwise accessions for their respective databases. When available Uniprot protein accessions are also given [42], along with the starting residue of the motif.

Figure 2

Alignments of putative EH1 motifs in a) Forkhead b) T-box c) ETS d) Doublesex and e) Zinc finger containing proteins. Alignment 'a' was derived from a meme search, as described in text. Sub-alignments b-e were derived from HMMER searches with the EH1^hox HMM. Other details as for Figure 1.

Figure 3

a) Distribution of HMMER bit scores for the database search of EH1^hox HMM against the combined proteomes of human, D. melanogaster and C. elegans. Counts from scores from transcription factors (see methods) have been coloured red – i.e. the proportion of a bar coloured red is equal to the proportion of transcription factors. Scores from proteins containing a homeobox domain (interpro accession IPR001356), from which the EH1^hox HMM was derived, have been excluded, b) as for 'a', but rescaled to show region of biological relevance. High scoring hits are greatly enriched in specific transcription factor families. For scores ≥ 5.0 bits, there are 68 transcription factors and 142 non-transcription factors; for scores <5.0 bits, 3075 transcription factors and 51513 non-transcription factors giving a chi-square test p-value statistic of P < 0.0001 – the statistical significance is discussed more fully in the text.