Elsevier

Gene

Volume 387, Issues 1–2, 31 January 2007, Pages 21-30
Gene

Missing link in the evolution of Hox clusters

https://doi.org/10.1016/j.gene.2006.08.011Get rights and content

Abstract

Hox cluster has key roles in regulating the patterning of the antero-posterior axis in a metazoan embryo. It consists of the anterior, central and posterior genes; the central genes have been identified only in bilaterians, but not in cnidarians, and are responsible for archiving morphological complexity in bilaterian development. However, their evolutionary history has not been revealed, that is, there has been a “missing link”. Here we show the evolutionary history of Hox clusters of 18 bilaterians and 2 cnidarians by using a new method, “motif-based reconstruction”, examining the gain/loss processes of evolutionarily conserved sequences, “motifs”, outside the homeodomain. We successfully identified the missing link in the evolution of Hox clusters between the cnidarian–bilaterian ancestor and the bilaterians as the ancestor of the central genes, which we call the proto-central gene. Exploring the correspondent gene with the proto-central gene, we found that one of the acoela Hox genes has the same motif repertory as that of the proto-central gene. This interesting finding suggests that the acoela Hox cluster corresponds with the missing link in the evolution of the Hox cluster between the cnidarian–bilaterian ancestor and the bilaterians. Our findings suggested that motif gains/diversifications led to the explosive diversity of the bilaterian body plan.

Introduction

Hox genes are the master control genes regulating the patterning of the antero-posterior axis in a metazoan embryo, except in sponges (Seimiya et al., 1994, Manuel and Le Parco, 2000), and are organized into a so-called “Hox cluster”. A Hox cluster consists of the anterior, central and posterior genes; the central genes have been identified only in bilaterians, but not in cnidarians, which are a basal metazoan phylum that arose before the divergence of bilaterians. The central Hox genes are diverse, and their diversity is considered to achieve morphological complexity in the bilaterian development. Understanding the evolution of the central Hox genes should thus lead to understanding the evolution of bilaterian body plan.

The evolutionary history of Hox clusters has been investigated both by comparing the Hox cluster organizations (Finnerty and Martindale, 1999, Ferrier and Holland, 2001) and by reconstructing the molecular phylogenetic trees (Zhang and Nei, 1996, de Rosa et al., 1999), but the evolutionary process of the central genes between the cnidarian–bilaterian ancestor (CBA) and the extant bilaterians has not been revealed. In other words, the evolutionary transition form of the central Hox genes between the CBA and the bilaterians has not been identified, leaving a “missing link”. A comparison of the Hox cluster organizations revealed a small Hox cluster consisting of only three or four genes, and this small cluster was considered to have appeared in the cnidarian–bilaterian ancestor (CBA) (Finnerty and Martindale, 1999, Ferrier and Holland, 2001). On the other hand, an alternative hypothesis that the CBA (ProtoHox cluster) have only two genes (Hox1/2 and posterior-class genes) was proposed (Garcia-Fernandez, 2005). Subsequent expansion of the Hox clusters occurred before the radiation of three major bilaterian clades, which are ecdysozoans, lophotrochozoans (these two clades compose protostomes) and deuterostomes (de Rosa et al., 1999). To identify the evolutionary transition form of Hox clusters, it is necessary to examine intermediate species that connect the CBA and the bilaterians, but there are not enough intermediate species to examine.

On the other hand, the molecular phylogenetic trees of Hox genes have been reconstructed from the conserved sequences of the homeodomain (Zhang and Nei, 1996, de Rosa et al., 1999). An evolutionary scenario of the Hox clusters based on the reconstructed tree was provisionally proposed (Zhang and Nei, 1996); however, the evolutionary relationships regarding the central genes have not been resolved, because the statistical support for clusters within the central genes was generally low. The homeodomain does not contain a great deal of phylogenetic information; it consists of only a 60-aa sequence that is highly conserved, whereas sequences outside the homeodomain are highly diverse.

To reconstruct the evolutionary relationships of Hox clusters, it was necessary to rely on the conserved sequences outside the homeodomain, such as the UbdA, Lox5 and para-peptides (de Rosa et al., 1999, Balavoine, 1998, Telford, 2000, Balavoine et al., 2002), but there are inevitable difficulties that arise when one applies the conventional molecular evolutionary analyses to those sequences. Though sequences outside the homeodomain show low similarity, they do contain several evolutionarily conserved sequences, so-called “motifs” (de Rosa et al., 1999, Balavoine, 1998, Balavoine et al., 2002). Here, focusing on and examining the motif gain/loss processes by using a new method we call “motif-based reconstruction”, we have attempted to reconstruct the evolutionary history of Hox clusters and determine the transition form of the central Hox genes between the CBA and the extant bilaterians, that is, the missing link.

Section snippets

Data collection

We collected amino acid sequences that encode the Hox genes of 16 protostomes: 6 ecdysozoans (Drosophila melanogaster, Cupiennius salei, Acanthokara kaputensis, Caenorhabditis elegans, Caenorhabditis briggsae and Priapulus caudatus) and 10 lophotrochozoans (Nereis virens, leeches (Hirudo medicinalis, Helobdella robusta and Helobdella triserialis), Lineus sanguineus, flatworms (Girardia tigrina, Polycelis nigra and Dugesia japonica), Euprymna scolopes and Lingula anatina). We also collected the

Motifs and their characteristic residues

First, we comprehensively discovered motifs among Hox genes of 2 cnidarians and 18 bilaterians, which consisted of 16 protostomes (6 ecdysozoans and 10 lophotrochozoans) and 2 deuterostomes (see Materials and methods section for the data collection), by using a motif discovery program, the MEME (Multiple EM algorithm for Motif Elicitation) (Bailey and Elkan, 1994). Significant motifs (p < 0.0001) were discovered in four regions: we called these regions the “Y”, “N”, “H” and “C” regions (except

Acknowledgements

We would like to thank Dr. Toshinori Endo and Dr. Yoshihito Niimura for their critical comments and helpful discussions with us, So Nakagawa for help in managing the bibliographic references, and Yoshiyuki Kaneko for help in preparing the manuscript. This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science, and Technology of Japan.

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