Evolution of hedgehog and hedgehog-related genes, their origin from Hog proteins in ancestral eukaryotes and discovery of a novel Hint motif

Additional file 1:

Multiple sequence alignment of Hog domains used for the protein sequence logos. Multiple sequence alignment in this and subsequent figures was carried out using first MUSCLE and imported subsequently into Clustal_X. Color coding was modified from default Clustal_X color coding by marking all cysteine residues in yellow, small hydrophobic residues in light blue and large hydrophobic residues in cyan blue. The conserved motifs, as well as the C-terminal SRR or ARR region are indicated in the alignment. The two conserved cysteine residues found in the Hog domain of nematode Hh-related proteins are indicated with red arrows.

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Additional file 2:

Full image of the protein sequence logo of aligned Hog domains shown in Figure 1. The color scheme is similar to the one used in the multiple sequence alignments (N,Q,S,T: green; C: yellow; P: pink; G: orange; K,R: red; A,I,L,M,V: blue; F,W,Y: cyan blue; H, purple, D,E: magenta; gaps: white). The extend of the Hint domain and the SRR region are indicated above the logo with a red line. Red boxes underneath the logo indicate the different motifs A, B, F, J, K, L.

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Additional file 3:

Protein sequence logo of nematode Hog domains. Protein sequence logo generated from nematode Hog domains shown in the multiple sequence aligment of Additional file 1. This logo is in register with the Hh Hog domain logo of Additional file 2. The two conserved cysteine residues specific to nematode Hh-related proteins are indicated with red arrows.

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Additional file 4:

Multiple sequence alignment of Wart domains. Wart domains were aligned and visualized in Clustal_X as described in Figure 2.

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Additional file 5:

Phylogenetic tree analysis of Hog domains using Neighbor joining. Neighbor joining tree without protist sequences. The Hog domain of the fungal gene GmGIN1 was used as outgroup.

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Additional file 6:

Phylogenetic tree analysis of Hog domains using Maximum likelihood. Maximum likelihood tree of the same sequences as in Additional file 5 with GmGIN1 as outgroup.

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Additional file 7:

Neighbor joining tree of Hog sequences which were truncated at the N-terminus. Hog sequences were truncated at the N-terminus to have the same size as the Pt wrt sequence fragment. This analysis shows that Pt wrt clusters with the wrt genes (arrow). GmGIN1 was used as outgroup. Note: Apart from Figure 5 and 6, and Additional files 5-7 further phylogenetic analysis were carried out that are not shown here. For example, the intein from vacuolar ATPase from C. tropicalis was used as outgroup [22] and gave comparable results to the tree analyses shown here.

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Additional file 8:

Multiple sequence alignment of Ground and Ground-like domains. Alignment of nematode Ground and Ground-like domains.

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Additional file 9:

Sequences used in the analysis. List of sequences, accession numbers, notes, predicted signal peptide cleavage sites and protein sequences used in this analysis.

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Additional file 10:

Multiple sequence alignment of "Hedge" domain containing proteins and Hedgehog proteins. Note that Os hhlike and NV 200640 do not line up in the Hog domain region.

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Additional file 11:

Multiple sequence alignment of Quahog proteins. Alignment of nematode Quahog proteins.

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Additional file 12:

Structure of the Nematostella vectensis genomic assembly around Nv 239508. Current assembly of the genomic region around Nv 239508. Color arrows indicate duplicated regions. N gap indicateds two regions with unknown sequence. The green area shows the ESTs found mapping to this region. The CAGN20453 correpsonds to Nv 239508. The yellow area shows regions of sequence similarity, i.e. hydrolase domain, Hog domain, and Reverse transcriptase. CAGN20453 is not sequenced fully, but the 3' read has been mapped to the right side, since the 3' untranslated region matches better to the 2. repeat of the duplication due to some indel differences. However, as will be noted, the final resulting transcript (shown at bottom) would be rather unusual, as it would splice over another gene, i.e. the hydrolase, which is also supported by an EST. Hence, the genomic organization and gene structure in this region could be subject to change, especially given the unsequenced areas.

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Additional file 13:

Predicted protein structure of Nv 200640. Protein motif prediction of the SMART server was used to analyse the ORF Nv 200640, and the different types of conserved motifs found are indicated.

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Additional file 14:

Multiple sequence alignment of non-metazoan Hog proteins. Alignment of non-metazoan Hog proteins including also the ones which are only based on EST fragments.

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Additional file 15:

Multiple sequence alignment of the Hint region of VWA-Vint proteins with Hog domains of Hh proteins.

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Additional file 16:

Multiple sequence alignment of bacterial proteins with a novel type of Hint-like region. Full length multiple sequence alignment of bacterial proteins. Motifs A and B are marked, as well as a small region with some similarity to the beginning of motif F in BIL-As. Note: the conserved upstream regions may be secreted, since some of the sequences have bacterial signal peptides for secretion. Species and the accession number for the sequences are: bSa_STIAU_1829: Stigmatella aurantiaca DW4/3-1 (ZP_01466308); bMx_MXAN_6253: Myxococcus xanthus DK 1622 (YP_634382); bRM_MED297_11140: Reinekea sp. MED297 (ZP_01113290); bPl_plu1731: Photorhabdus luminescens subsp. laumondii TTO1 (NP_929012); bSp_Draf4685: Serratia proteamaculans 568 (ZP_01534811); bYi_YintA_01002283: Yersinia intermedia ATCC 29909 (ZP_00833384).

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Additional file 17:

Multiple sequence alignment of the Hint region of the bacterial proteins with a novel Hint-like domain with Hog domains. Multiple sequence alignment of the bacterial Hint domains from Additional file 16 with Hog domains from animals. Note the roughly similar length.

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