Seven Hox gene sequences from the asterinid starfish Patiriella exigua (Echinodermata: Asteroidea)

Partial homeobox gene fragments were amplified from Patiriella exigua genomic DNA by PCR using degenerate primers. The primers spanned a region of 82 nucleotides within the homeobox, encompassing amino acids 21–47 of the homeodomain. The deduced amino acid sequences of the seven Hox gene sequences plus one Xlox-type gene sequence from P. exigua are presented and compared with similar sequences from other organisms. This work is a preliminary step in the analysis of the evolution of development in the Patiriella species group and the roles of Hox gene expression therein.     

Morphometric classification of neurosecretory granules in the neurohypophysis of the hagfish,Eptatretus burgeri

Summary Neurosecretory axons in the neurohypophysis of the hagfish, Eptatretus burgeri, were statistically classified into six types according to the size of secretory granules. These types are comparable with those in higher vertebrates. The concentration of each axon type is different in three regions: anterior dorsal wall, posterior dorsal wall, and ventral wall. The regional differences of the hagfish neurohypophysis are discussed in relation to the regional differentiation of the tetrapod neurohypophysis into the median eminence and the pars nervosa.We wish to express our appreciation to Professor Hideshi Kobayashi for the guidance of this study. It was aided by grants from the Ford Foundation and the Ministry of Education to Professor H. Kobayashi, and also by grants from the Population Council, New York and the Ministry of Education to Professor S. Ishii.     

Iodine distribution in the thyroid follicles of the hagfish,Eptatretus burgeri and lamprey,Lampetra japonica: Electron-probe X-ray microanalysis

Summary In the thyroid follicles of species of cyclostomes, a hagfish and a lamprey, the distribution of stable iodine was examined by electron-probe X-ray microanalysis. A high concentration of stable iodine, heterogeneously distributed, was observed in the follicular cells of hagfish thyroid follicles. In the lamprey a low concentration of iodine was seen in the follicular lumina. The relative values for stable iodine determined in this way corresponded to values obtained by a chemical analytical method.     

Hox genes in the echiuroid Urechis unicinctus

An echiuroid species, Urechis unicinctus, was surveyed for Hox genes using polymerase chain reaction with homeobox-specific degenerate primers. We identified nine distinct homeodomain-containing gene fragments. These nine fragments were classified by comparative analysis. This analysis revealed that this echiuroid possessed at least three Hox genes from the anterior group, five from the central group, and one from the posterior group.     

Evidence for Hox Gene Duplication in Rainbow Trout (Oncorhynchus mykiss): A Tetraploid Model Species

We examined the genomic organization of Hox genes in rainbow trout (Oncorhynchus mykiss), a tetraploid teleost derivative species, in order to test models of presumptive genomic duplications during vertebrate evolution. Thirteen putative clusters were localized in the current rainbow trout genetic map; however, analysis of the sequence data suggests the presence of at least 14 Hox clusters. Many duplicated genes appear to have been retained in the genome and share a high percentage of amino acid similarity with one another. We characterized two Hox genes located within the HoxCb cluster that may have been lost independently in other teleost species studied to date. Finally, we identified conserved syntenic blocks between salmonids and human, and provide data supporting two new linkage group homeologies (i.e., RT-3/16, RT-12/29) and three previously described homeologies (RT-2/9, RT-17/22, and RT-27/31) in rainbow trout. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. The sequence data for this study have been submitted to GenBank under the following accession numbers: AY567792, AY567793, AY567794, AY567795, AY567796, AY567797, AY567798, AY567799, AY567800, AY567801, AY567802, AY567803, AY567804, AY567805, AY567806, AY567807, AY567808, AY567809, AY567810, AY567812, AY567813, AY567814, AY567815, AY567816, and AY567817. [Reviewing Editor : Dr. Axel Meyer]     

Isolation of Hox and Parahox genes in the hemichordate Ptychodera flava and the evolution of deuterostome Hox genes

Because of their importance for proper development of the bilaterian embryo, Hox genes have taken center stage for investigations into the evolution of bilaterian metazoans. Taxonomic surveys of major protostome taxa have shown that Hox genes are also excellent phylogenetic markers, as specific Hox genes are restricted to one of the two great protostome clades, the Lophotrochozoa or the Ecdysozoa, and thus support the phylogenetic relationships as originally deduced by 18S rDNA studies. Deuterostomes are the third major group of bilaterians and consist of three major phyla, the echinoderms, the hemichordates, and the chordates. Most morphological studies have supported Hemichordata+Chordata, whereas molecular studies support Echinodermata+Hemichordata, a clade known as Ambulacraria. To test these competing hypotheses, complete or near complete cDNAs of eight Hox genes and four Parahox genes were isolated from the enteropneust hemichordate Ptychodera flava. Only one copy of each Hox gene was isolated suggesting that the Hox genes of P. flava are arranged in a single cluster. Of particular importance is the isolation of three posterior or Abd-B Hox genes; these genes are only shared with echinoderms, and thus support the monophyly of Ambulacraria.     

FMRFamide-like immunoreactivity in the brain of the Pacific hagfish,Eptatretus stouti (Myxinoidea)

Summary The distribution of FMRFamide-like immunoreactivity was investigated in the brain of a myxinoid, the Pacific hagfish,Eptatretus stouti, by means of immunocytochemistry. In the forebrain, labelled cell bodies occurred in the infundibular nucleus of the hypothalamus and some closely adjacent nuclei. Labelled fibers formed a diffuse network in the forebrain, but there was no evidence for the presence of intracerebral ganglionic cells of the terminal nerve or a central projection of the terminal nerve. In the hindbrain, a group of labelled cells was found in the trigeminal sensory nucleus. A distinet terminal arborization occurred in the ventrally adjacent nucleus A of Kusunoki and around the nuclei of the branchial motor column. These findings suggest that FMRFamide may play a role in the central control of branchiomotor activity.     

Distribution of 5-HT (serotonin) immunoreactivity in the central nervous system of the inshore hagfish,Eptatretus burgeri

Summary Immunohistochemical techniques were used to study the distribution of serotonin in the central nervous system of the hagfish,Eptatretus burgeri, in order to produce a detailed map of serotonin-containing structures. In the hypothalamus, many serotonin-containing neurons contacted the cerebrospinal fluid. Most of the serotonin-containing cell bodies were located in the raphe region, where they were compactly distributed at the level of the nucleus motorius tegmenti pars anterior but more diffusely distributed at the level of the nucleus motorius tegmenti pars posterior. Serotonin-containing cell bodies and varicose fibers were widely distributed throughout the brain and upper spinal cord segments, but the distribution density was not even. On the basis of its abundance, serotonin can be judged to have an important function in the control of the hagfish central nervous system. From a phylogenetic point of view, serotonin-containing neurons in the raphe region appear to be a common property of all classes of vertebrates studied except the lampreys, whereas serotonin-containing cerebrospinal fluid-contacting neurons may be considered to be a primitive condition in all nonmammalian vertebrates.     

Keratin-like components of gland thread cells modulate the properties of mucus from hagfish (Eptatretus stouti)

Summary The hagfishes (cyclostomes) are known to secrete copious amounts of mucus mainly by the holocrine mode from the slime glands. Stressed animals release two types of cells (gland thread cells, GTCs; gland mucous cells, GMCs) which rupture on contact with water and rapidly form a mass of viscous mucus. Herein we report some key sequential events of this process and document a novel role for cytoskeletal polymers. After electrostimulation of Pacific hagfish (Eptatretus stouti), the exudate was collected in a stabilization buffer and GTCs segregated from GMC vesicles. Water was added progressively to mixtures of known quantities of these entities. The changing mucous composition and properties were monitored by light- and electron microscopy, viscometry and immunogold assay. Sequentially, the threads uncoil from GTCs, aggregate with the vesicles, the vesicles rupture and release mucin-like substances, at least some of which adhere to the thread. It was found that the intermediate filament (IF)-rich threads markedly facilitate hydration and modulate the viscoelastic and cohesive properties of the resultant mucus. It was speculated that the thread abets localization of mucus in an aqueous environment and promotes adhesion of mucus to surfaces such as the fish integument. As judged by immunostaining in situ, GTCs, as well as several cell-types in the epidermis, contain keratin-like components. The role of biopolymers on the properties of teleost and mammalian mucus is discussed.     

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.     

Evolution of Hox Clusters in Salmonidae: A Comparative Analysis Between Atlantic Salmon (Salmo salar) and Rainbow Trout (Oncorhynchus mykiss)

We studied the genomic organization of Hox genes in Atlantic salmon (Salmo salar) and made comparisons to that in rainbow trout (Oncorhynchus mykiss), another member of the family Salmonidae. We used these two species to test the hypothesis that the Hox genes would provide evidence for a fourth round of duplication (4R) of this gene family given the recent polyploid ancestry of the salmonid fish. Thirteen putative Hox clusters were identified and 10 of these complexes were localized to the current Atlantic salmon genetic map. Syntenic regions with the rainbow trout linkage map were detected and further homologies and homeologies are suggested. We propose that the common ancestor of Atlantic salmon and rainbow trout possessed at least 14 clusters of Hox genes, and additional clusters cannot be ruled out. Salmonid Hox cluster complements seem to be more similar to those of zebrafish (Danio rerio) than medaka (Oryzias latipes) or pufferfish (Sphoeroides nephelus and Takifugu rubripes), as both Atlantic salmon and rainbow trout have retained HoxCb ortholog, which has been lost in medaka and pufferfish but not in zebrafish. However, our data suggest that phylogenetically, the homologous genes within each cluster express mosaic relationships among the teleosts tested and, thus, leave unresolved the interfamilial relationships among these taxa. Sequence data from this article have been deposited within the EMBL/GenBank Data Libraries under the following accession numbers: AY677341, AY677342, AY677343, AY677344, AY677345, AY677346, AY677347, AY677348, AY677349, AY677350, AY677351, AY677352, AY677353, AY677354, AY677355, AY677356, AY677357, AY677358, AY677359, AY677360, AY677361, AY677362, AY677363, AY677364 and AY677365. [Reviewing Editor: Dr. Axel Meyer]     

Hox gene expression patterns in Lethenteron japonicum embryos--insights into the evolution of the vertebrate Hox code

The Hox code of jawed vertebrates is characterized by the colinear and rostrocaudally nested expression of Hox genes in pharyngeal arches, hindbrain, somites, and limb/fin buds. To gain insights into the evolutionary path leading to the gnathostome Hox code, we have systematically analyzed the expression pattern of the Hox gene complement in an agnathan species, Lethenteron japonicum (Lj). We have isolated 15 LjHox genes and assigned them to paralogue groups (PG) 1-11, based on their deduced amino acid sequences. LjHox expression during development displayed gnathostome-like spatial patterns with respect to the PG numbers. Specifically, lamprey PG1-3 showed homologous expression patterns in the rostral hindbrain and pharyngeal arches to their gnathostome counterparts. Moreover, PG9-11 genes were expressed specifically in the tailbud, implying its posteriorizing activity as those in gnathostomes. We conclude that these gnathostome-like colinear spatial patterns of LjHox gene expression can be regarded as one of the features already established in the common ancestor of living vertebrates. In contrast, we did not find evidence for temporal colinearity in the onset of LjHox expression. The genomic and developmental characteristics of Hox genes from different chordate species are also compared, focusing on evolution of the complex body plan of vertebrates.     

PCR-based detection of a tracheal mite of the honey bee Acarapis woodi

The effects of the tracheal mite Acarapis woodi on the health of honey bees have been neglected since the prevalence of Varroa mites to Apis mellifera colonies. However, tracheal mite infestation of honey bee colonies still occurs worldwide and could impose negative impact on apiculture. The detection of A. woodi requires the dissection of honey bees followed by microscopic observation of the tracheal sacs. We thus developed PCR methods to detect A. woodi. These methods facilitate rapid and sensitive detection of A. woodi in many honey bee samples for epidemiologic surveys.     

Evolution of echinoderms may not have required modification of the ancestral deuterostome HOX gene cluster: first report of PG4 and PG5 Hox orthologues in echinoderms

Is the extreme derivation of the echinoderm body plan reflected in a derived echinoderm Hox genotype? Building on previous work, we exploited the sequence conservation of the homeobox to isolate putative orthologues of several Hox genes from two asteroid echinoderms. The 5-peptide motif (LPNTK) diagnostic of PG4 Hox genes was identified immediately downstream of one of the partial homeodomains from Patiriella exigua. This constitutes the first unequivocal report of a PG4 Hox gene orthologue from an echinoderm. Subsequent screenings identified genes of both PG4 and PG4/5 in Asterias rubens. Although in echinoids only a single gene (PG4/5) occupies these two contiguous cluster positions, we conclude that the ancestral echinoderm must have had the complete deuterostome suite of medial Hox genes, including orthologues of both PG4 and PG4/5 (= PG5). The reported absence of PG4 in the HOX cluster of echinoids is therefore a derived state, and the ancestral echinoderm probably had a HOX cluster not dissimilar to that of other deuterostomes. Modification of the ancestral deuterostome Hox genotype may not have been required for evolution of the highly derived echinoderm body plan.     

Peter Holland,homeobox genes,and the developmental basis of animal diversity

In 1867 Alexander Kowalevsky published an account of the development of the cephalochordate Amphioxus lanceolatus (now known as Branchiostoma lanceolatum) (Kowalevsky, 1867). Together with his study of the development of urochordates (Kowalevsky, 1866; 1871), this introduced a new way of thinking about the relationship between the evolution and development of animals, and established the basis for long-standing theories of the evolutionary origin of vertebrates. Some one hundred and fifty years later, cephalochordates and urochordates are again in the spotlight, as molecular biology and genome sequencing promise further revelations about the origin of vertebrates. The work of the 2006 Kowalevsky Medal winner, Peter Holland (Fig. 1), has played a central role in their reinstatement (see Mikhailov and Gilbert (2002) for more details of the history of the Kowalevsky Medal). Here, I profile Peter Holland’s contribution to the rebirth of Evolutionary Developmental Biology in general and the study of homeobox genes and vertebrate origins in particular.     

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     

Evidence for placing the false gharial (Tomistoma schlegelii) into the family Gavialidae: inferences from nuclear gene sequences

The extant crocodylians comprise 23 species divided among three families, Alligatoridae, Crocodylidae, and Gavialidae. Currently, based on morphological data sets, Tomistoma schlegelii (false gharial) is placed within the family Crocodylidae. Molecular data sets consistently support a sister-taxon relationship of T. schlegelii with Gavialis gangeticus (Indian Gharial), which is the sole species in Gavialidae. To elucidate the placement of T. schlegelii within the extant crocodylians, we have sequenced 352bp of the dentin matrix protein 1 (DMP1) nuclear gene in 30 individuals and 424bp of the nuclear gene C-mos in 74 individuals. Molecular analysis of the DMP1 data set indicates that it is highly conserved within the Crocodylia. Of special note is a seven base-pair indel (GTGCTTT) shared by T. schlegelii and G. gangeticus, that is absent in the genus Crocodylus, Osteolaemus, and Mecistops. To date, C-mos is the largest molecular data set analyzed for any crocodylian study including multiple samples from all representatives of the eight extant genera. Analysis of these molecular data sets, both as individual gene sequences and concatenated sequences, support the hypothesis that T. schlegelii should be placed within the family Gavialidae.