Organization of the Hox gene cluster of the silkworm, <Emphasis Type="Italic">Bombyx mori</Emphasis>: a split of the Hox cluster in a non-<Emphasis Type="Italic">Drosophila</Emphasis> insect

A bacterial artificial chromosome (BAC) contig was constructed by chromosome walking, starting from the Hox genes of the silkworm, Bombyx mori. Bombyx orthologues of the labial (lab) and zerknült (zen) genes were newly identified. The size of the BAC contig containing the Hox gene cluster—except the lab and Hox 2 genes—was estimated to be more than 2 Mb. The Bombyx Hox cluster was mapped to linkage group (LG) 6. The lab gene was mapped on the same LG, but far apart from the cluster. Fluorescence in situ hybridization analysis confirmed that the major Hox gene cluster and lab were at different locations on the same chromosome in B. mori.Edited by M. Akam     

Current problems with the zootype and the early evolution of Hox genes

"Hox cluster type" genes have sparked intriguing attempts to unite all metazoan animals by a shared pattern of expression and genomic organization of a specific set of regulatory genes. The basic idea, the zootype concept, claims the conservation of a specific set of "Hox cluster type genes" in all metazoan animals, i.e., in the basal diploblasts as well as in the derived triploblastic animals. Depending on the data used and the type of analysis performed, different opposing views have been taken on this idea. We review here the sum of data currently available in a total evidence analysis, which includes morphological and the most recent molecular data. This analysis highlights several problems with the idea of a simple "Hox cluster type" synapomorphy between the diploblastic and triploblastic animals and suggests that the "zootype differentiation" of the Hox cluster most likely is an invention of the triploblasts. The view presented is compatible with the idea that early Hox gene evolution started with a single proto-Hox (possibly a paraHox) gene. J. Exp. Zool. (Mol. Dev. Evol.) 291:169-174, 2001.     

Two more Posterior Hox genes and Hox cluster dispersal in echinoderms

Background

Hox genes are key elements in patterning animal development. They are renowned for their, often, clustered organisation in the genome, with supposed mechanistic links between the organisation of the genes and their expression. The widespread distribution and comparable functions of Hox genes across the animals has led to them being a major study system for comparing the molecular bases for construction and divergence of animal morphologies. Echinoderms (including sea urchins, sea stars, sea cucumbers, feather stars and brittle stars) possess one of the most unusual body plans in the animal kingdom with pronounced pentameral symmetry in the adults. Consequently, much interest has focused on their development, evolution and the role of the Hox genes in these processes. In this context, the organisation of echinoderm Hox gene clusters is distinctive. Within the classificatory system of Duboule, echinoderms constitute one of the clearest examples of Disorganized (D) clusters (i.e. intact clusters but with a gene order or orientation rearranged relative to the ancestral state).

Results

Here we describe two Hox genes (Hox11/13d and e) that have been overlooked in most previous work and have not been considered in reconstructions of echinoderm Hox complements and cluster organisation. The two genes are related to Posterior Hox genes and are present in all classes of echinoderm. Importantly, they do not reside in the Hox cluster of any species for which genomic linkage data is available.

Conclusion

Incorporating the two neglected Posterior Hox genes into assessments of echinoderm Hox gene complements and organisation shows that these animals in fact have Split (S) Hox clusters rather than simply Disorganized (D) clusters within the Duboule classification scheme. This then has implications for how these genes are likely regulated, with them no longer covered by any potential long-range Hox cluster-wide, or multigenic sub-cluster, regulatory mechanisms.

     

Parsimony Analysis as a Specific Kind of Homology Estimation and the Implications for Character Weighting

“Remane-Hennigian systematists” still reject parsimony analysis for phylogenetics, because homology or apomorphy analyses are not included. In contrast, “pattern cladists” regard homology as a deductive concept after applying a parsimony test of character congruence. However, as in molecular phylogeny, selection of “good” characters is always done on the basis of ana priorihomology analysis. The distribution criterion of homology—“homologous characters have identical or hierarchical distribution”—is the basis of parsimony analysis. Because this criterion also might fail in cases of genealogical reticulation or concerted homoplasy, character congruence is not a strict test but another probabilistic criterion of homology. A synthetic approach is proposed for phenotypic analysis with application ofa prioricriteria of homology. The resultinga prioriprobabilities of homology serve as criteria for selection and weighting of characters (very low = not selected/poor/mediocre/good/Dollo characters). After application of a parsimony algorithm the final cladogram decides homology estimations.     

Hox gene expression in larval development of the polychaetes Nereis virens and Platynereis dumerilii (Annelida, Lophotrochozoa)

The bilaterian animals are divided into three great branches: the Deuterostomia, Ecdysozoa, and Lophotrochozoa. The evolution of developmental mechanisms is less studied in the Lophotrochozoa than in the other two clades. We have studied the expression of Hox genes during larval development of two lophotrochozoans, the polychaete annelids Nereis virens and Platynereis dumerilii. As reported previously, the Hox cluster of N. virens consists of at least 11 genes (de Rosa R, Grenier JK, Andreeva T, Cook CE, Adoutte A, Akam M, Carroll SB, Balavoine G, Nature, 399:772–776, 1999; Andreeva TF, Cook C, Korchagina NM, Akam M, Dondua AK, Ontogenez 32:225–233, 2001); we have also cloned nine Hox genes of P. dumerilii. Hox genes are mainly expressed in the descendants of the 2d blastomere, which form the integument of segments, ventral neural ganglia, pre-pygidial growth zone, and the pygidial lobe. Patterns of expression are similar for orthologous genes of both nereids. In Nereis, Hox2, and Hox3 are activated before the blastopore closure, while Hox1 and Hox4 are activated just after this. Hox5 and Post2 are first active during the metatrochophore stage, and Hox7, Lox4, and Lox2 at the late nectochaete stage only. During larval stages, Hox genes are expressed in staggered domains in the developing segments and pygidial lobe. The pattern of expression of Hox cluster genes suggests their involvement in the vectorial regionalization of the larval body along the antero-posterior axis. Hox gene expression in nereids conforms to the canonical patterns postulated for the two other evolutionary branches of the Bilateria, the Ecdysozoa and the Deuterostomia, thus supporting the evolutionary conservatism of the function of Hox genes in development. Milana Kulakova, Nadezhda Bakalenko and Elena Novikova contributed equally to this work.     

Ancient origins of axial patterning genes: Hox genes and ParaHox genes in the Cnidaria

Among the bilaterally symmetrical, triploblastic animals (the Bilateria), a conserved set of developmental regulatory genes are known to function in patterning the anterior–posterior (AP) axis. This set includes the well-studied Hox cluster genes, and the recently described genes of the ParaHox cluster, which is believed to be the evolutionary sister of the Hox cluster ( Brooke et al. 1998 ). The conserved role of these axial patterning genes in animals as diverse as frogs and flies is believed to reflect an underlying homology (i.e., all bilaterians derive from a common ancestor which possessed an AP axis and the developmental mechanisms responsible for patterning the axis). However, the origin and early evolution of Hox genes and ParaHox genes remain obscure. Repeated attempts have been made to reconstruct the early evolution of Hox genes by analyzing data from the triphoblastic animals, the Bilateria ( Schubert et al. 1993 ; Zhang and Nei 1996 ). A more precise dating of Hox origins has been elusive due to a lack of sufficient information from outgroup taxa such as the phylum Cnidaria (corals, hydras, jellyfishes, and sea anemones). In combination with outgroup taxa, another potential source of information about Hox origins is outgroup genes (e.g., the genes of the ParaHox cluster). In this article, we present cDNA sequences of two Hox-like genes ( anthox2 and anthox6 ) from the sea anemone, Nematostella vectensis. Phylogenetic analysis indicates that anthox2 (=Cnox2) is homologous to the GSX class of ParaHox genes, and anthox6 is homologous to the anterior class of Hox genes. Therefore, the origin of Hox genes and ParaHox genes occurred prior to the evolutionary split between the Cnidaria and the Bilateria and predated the evolution of the anterior–posterior axis of bilaterian animals. Our analysis also suggests that the central Hox class was invented in the bilaterian lineage, subsequent to their split from the Cnidaria.     

Hox Genes from the Tapeworm Taenia asiatica (Platyhelminthes: Cestoda)

Hox genes are important in forming the anterior-posterior body axis pattern in the early developmental stage of animals. The conserved nature of the genomic organization of Hox genes is well known in diverse metazoans. To understand the Hox gene architecture in human-infecting Taenia tapeworms, we conducted a genomic survey of the Hox gene using degenerative polymerase chain reaction primers in Taenia asiatica. Six Hox gene orthologs from 276 clones were identified. Comparative analysis revealed that T. asiatica has six Hox orthologs, including two lab/Hox1, two Hox3, one Dfd/Hox4, and one Lox2/Lox4. The results suggest that Taenia Hox genes may have undergone independent gene duplication in two Hox paralogs. The failure to detect Post1/2 orthologs in T. asiatica may suggest that sequence divergence or the secondary loss of the posterior genes has occurred in the lineage leading to the cestode and trematode.     

Genomic organization of Hox and ParaHox clusters in the echinoderm,Acanthaster planci

The organization of echinoderm Hox clusters is of interest due to the role that Hox genes play in deuterostome development and body plan organization, and the unique gene order of the Hox complex in the sea urchin Strongylocentrotus purpuratus, which has been linked to the unique development of the axial region. Here, it has been reported that the Hox and ParaHox clusters of Acanthaster planci, a corallivorous starfish found in the Pacific and Indian oceans, generally resembles the chordate and hemichordate clusters. The A. planci Hox cluster shared with sea urchins the loss of one of the medial Hox genes, even‐skipped (Evx) at the anterior of the cluster, as well as organization of the posterior Hox genes. genesis 52:952–958, 2014. © 2014 Wiley Periodicals, Inc.     

Are Hox Genes Ancestrally Involved in Axial Patterning? Evidence from the Hydrozoan Clytia hemisphaerica (Cnidaria)

Background

The early evolution and diversification of Hox-related genes in eumetazoans has been the subject of conflicting hypotheses concerning the evolutionary conservation of their role in axial patterning and the pre-bilaterian origin of the Hox and ParaHox clusters. The diversification of Hox/ParaHox genes clearly predates the origin of bilaterians. However, the existence of a “Hox code” predating the cnidarian-bilaterian ancestor and supporting the deep homology of axes is more controversial. This assumption was mainly based on the interpretation of Hox expression data from the sea anemone, but growing evidence from other cnidarian taxa puts into question this hypothesis.

Methodology/Principal Findings

Hox, ParaHox and Hox-related genes have been investigated here by phylogenetic analysis and in situ hybridisation in Clytia hemisphaerica, an hydrozoan species with medusa and polyp stages alternating in the life cycle. Our phylogenetic analyses do not support an origin of ParaHox and Hox genes by duplication of an ancestral ProtoHox cluster, and reveal a diversification of the cnidarian HOX9-14 genes into three groups called A, B, C. Among the 7 examined genes, only those belonging to the HOX9-14 and the CDX groups exhibit a restricted expression along the oral-aboral axis during development and in the planula larva, while the others are expressed in very specialised areas at the medusa stage.

Conclusions/Significance

Cross species comparison reveals a strong variability of gene expression along the oral-aboral axis and during the life cycle among cnidarian lineages. The most parsimonious interpretation is that the Hox code, collinearity and conservative role along the antero-posterior axis are bilaterian innovations.     

Hox Gene Loss during Dynamic Evolution of the Nematode Cluster

HOX GENES ARE IMPORTANT: their central role in anterior-posterior patterning provides a framework for molecular comparison of animal body plan evolution. The nematode Caenorhabditis elegans stands out as having a greatly reduced Hox gene complement. To address this, orthologs of C. elegans Hox genes were identified in six species from across the Nematoda, and they show that rapid homeodomain sequence evolution is a general feature of nematode Hox genes. Some nematodes express additional Hox genes belonging to orthology groups that are absent from C. elegans but present in other bilaterian animals. Analysis of the genomic environment of a newly identified Brugia malayi Hox6-8 ortholog (Bm-ant-1) revealed that it lay downstream of the Bm-egl-5 Hox gene and that their homeodomain exons are alternately cis spliced to the same 5' exon. This organization may represent an intermediate state in Hox gene loss via redundancy. The Hox clusters of nematodes are the product of a dynamic mix of gene loss and rapid sequence evolution, with the most derived state observed in the model C. elegans.     

The <Emphasis Type="Italic">Hox8</Emphasis> of the hemichordate <Emphasis Type="Italic">Balanoglossus misakiensis</Emphasis>

Deuterostomes comprise a monophyletic group of animals that include chordates, xenoturbellids, and the Ambulacraria, which consists of echinoderms and hemichordates. The ancestral chordate probably had 14 Hox genes aligned linearly along the chromosome, with the posterior six genes showing an independent duplication compared to protostomes. In contrast, ambulacrarians are characterized by a duplication of the posterior Hox genes, resulting in three genes known as Hox11/13a, Hox11/13b, and Hox11/13c. Here, we isolated 12 Hox genes from the hemichordate Balanoglossus misakiensis and found an extra Hox gene that has not been reported in hemichordates. The extra B. misakiensis gene was suggested to be Hox8 from paralog-characteristic residues in its hexapepetide motif and homeodomain and a comparison with Strongylocentrotus purpuratus Hox genes. Our data suggest that the ancestor of echinoderms and hemichordates may have had a full complement of 12 Hox genes.     

The cytochrome P450 genesis locus: the origin and evolution of animal cytochrome P450s

The neighbourhoods of cytochrome P450 (CYP) genes in deuterostome genomes, as well as those of the cnidarians Nematostella vectensis and Acropora digitifera and the placozoan Trichoplax adhaerens were examined to find clues concerning the evolution of CYP genes in animals. CYP genes created by the 2R whole genome duplications in chordates have been identified. Both microsynteny and macrosynteny were used to identify genes that coexisted near CYP genes in the animal ancestor. We show that all 11 CYP clans began in a common gene environment. The evidence implies the existence of a single locus, which we term the ‘cytochrome P450 genesis locus’, where one progenitor CYP gene duplicated to create a tandem set of genes that were precursors of the 11 animal CYP clans: CYP Clans 2, 3, 4, 7, 19, 20, 26, 46, 51, 74 and mitochondrial. These early CYP genes existed side by side before the origin of cnidarians, possibly with a few additional genes interspersed. The Hox gene cluster, WNT genes, an NK gene cluster and at least one ARF gene were close neighbours to this original CYP locus. According to this evolutionary scenario, the CYP74 clan originated from animals and not from land plants nor from a common ancestor of plants and animals. The CYP7 and CYP19 families that are chordate-specific belong to CYP clans that seem to have originated in the CYP genesis locus as well, even though this requires many gene losses to explain their current distribution. The approach to uncovering the CYP genesis locus overcomes confounding effects because of gene conversion, sequence divergence, gene birth and death, and opens the way to understanding the biodiversity of CYP genes, families and subfamilies, which in animals has been obscured by more than 600 Myr of evolution.     

Are Cirripedia hopeful monsters? Cytogenetic approach and evidence for a Hox gene cluster in the cirripede crustacean Sacculina carcini

The “hopeful monster” has haunted evolutionary thinking since Richard Goldschmidt coined the phrase in 1933. The phrase is directly related to genetic mechanisms in development and evolution. Cirripedes are peculiar crustaceans in that they all lack abdomens as adults. In a previous study aimed at describing the repertoire of Hox genes of the Cirripedia, we failed to isolate the abdominal-A gene in three species representative of all three cirripede orders. To address the question of whether the cirripede ancestor could have been a “hopeful monster” arising from a rearrangement of the Hox complex, we have performed a cytogenetic analysis of the Hox complex of the cirripede Sacculina carcini. We present here molecular and cytogenetic evidence for the grouping of the Hox genes on a single chromosome. This is the first direct evidence reported for the grouping of Hox genes on the same chromosome in a non-insect arthropod species.