The ParaHox gene Gsx patterns the apical organ and central nervous system but not the foregut in scaphopod and cephalopod mollusks
© Wollesen et al. 2015
Received: 19 June 2015
Accepted: 17 December 2015
Published: 29 December 2015
It has been hypothesized that the ParaHox gene Gsx patterned the foregut of the last common bilaterian ancestor. This notion was corroborated by Gsx expression in three out of four lophotrochozoan species, several ecdysozoans, and some deuterostomes. Remarkably, Gsx is also expressed in the bilaterian anterior-most central nervous system (CNS) and the gastropod and annelid apical organ. To infer whether these findings are consistent with other mollusks or even lophotrochozoans, we investigated Gsx expression in developmental stages of representatives of two other molluscan classes, the scaphopod Antalis entalis and the cephalopod Idiosepius notoides.
Gsx is not expressed in the developing digestive tract of Antalis entalis and Idiosepius notoides. Instead, it is expressed in cells of the apical organ in the scaphopod trochophore and in two cells adjacent to this organ. Late-stage trochophores express Aen-Gsx in cells of the developing cerebral and pedal ganglia and in cells close to the pavilion, mantle, and foot. In postmetamorphic specimens, Aen-Gsx is expressed in the cerebral and pedal ganglia, the foot, and the nascent captacula. In early squid embryos, Ino-Gsx is expressed in the cerebral, palliovisceral, and optic ganglia. In late-stage embryos, Ino-Gsx is additionally expressed close to the eyes and in the supraesophageal and posterior subesophageal masses and optic lobes. Developmental stages close to hatching express Ino-Gsx only close to the eyes.
Our results suggest that Gsx expression in the foregut might not be a plesiomorphic trait of the Lophotrochozoa as insinuated previously. Since neither ecdysozoans nor deuterostomes express Gsx in their gut, a role in gut formation in the last common bilaterian ancestor appears unlikely. Gsx is consistently expressed in the bilaterian anterior-most CNS and the apical organ of lophotrochozoan larvae, suggesting a recruitment of Gsx into the formation of this organ in the Lophotrochozoa. The cephalopod posterior subesophageal mass and optic ganglia and the scaphopod pedal ganglia also express Gsx. In summary, Gsx expression only appears to be conserved in the anterior-most brain region during evolution. Accordingly, Gsx appears to have been recruited into the formation of other expression domains, e.g., the apical organ or the foregut, in some lophotrochozoans.
KeywordsBrain Cephalopoda Evolution Development Hox Homeobox genes Invertebrate Lophotrochozoa Mollusca Ontogeny Scaphopoda Lophotrochozoa
The Hox and ParaHox gene clusters are considered to be derived from a hypothetical ProtoHox cluster by duplication . Both belong to the homeobox gene family and exhibit highly conserved amino acid sequences in phylogenetically distantly related animals [1, 2]. In the majority of bilaterians investigated, it has been shown that Hox genes are expressed in tempo-spatial collinearity during development, in particular in neuroectodermal domains [2, 3]. Cephalopod and gastropod mollusks were among the first examples among bilaterians that apparently do not exhibit such a collinear mode of Hox gene expression [4, 5]. Tempo-spatial collinear expression of the three ParaHox genes has also been proposed for the last common bilaterian ancestor . It has been hypothesized that Gsx was expressed in the foregut, Xlox in the midgut, and Cdx in the hindgut in the last common bilaterian ancestor [1, 6]. While Xlox expression in the midgut and Cdx expression in the hindgut was found in various bilaterians, no Gsx expression has been reported in the foregut of any deuterostome representative to date [1, 6]. This was explained by the fact that the blastopore does not develop into the prospective mouth in deuterostomes. Deuterostomes instead evolved a new mouth and hence Gsx might have lost its role in patterning the anterior-most region of the digestive tract. Interestingly, the deuterostome hemichordate Ptychodera flava, for example, does express Gsx around the blastopore, however, apparently not in the digestive tract of subsequent developmental stages . Holland anticipated that protostome invertebrates may show Gsx expression in the foregut since their blastopore usually does become the future mouth [1, 6].
Gsx gene expression domains in metazoan developmental stages as revealed by in situ hybridization experiments
Name of Gsx ortholog
Gsx expression domains
Ectodermal cells along the oral/aboral body axis (rare in oral region)
Posterior endoderm, i.e., prospective oral end
Developing mesenteries (ectoderm),
Late planula larva
Columnar ectodermal cells in tentacle buds
Planula and embryos
Endodermal cells in oral and aboral region
Anterior and posterior endoderm
Bilateral pair of 4-5 cells in dorso-median episphere (anlagen of cerebral ganglia?)
Pair of each three sensory cells in apical organ
Cells around stomodeum
Two apical tuft cells and sensory cup cells of apical organ
Cells around mouth opening
Ventral portion of nascent digestive gland
Post-torsional competent veliger
Ventral portion of digestive gland
Cells around mouth opening
Cells at ventral border of the yolk-filled cells
Cells in cerebral ganglia anlagen
Cells in foregut close to radula anlage
Posterior radula sac
2 cells each in the lateral episphere on both sides
1 cell each lateral to the anus on both sides
1 pair of cells in the apical organ and another pair lateral to latter
1 cell each lateral to the anus on both sides
1 cell each in posterolateral mantle on both sides
Several cells in the region of the cerebral and pedal ganglia and ventral foot
Metamorphic competent trochophore
Several cells in the region of the cerebral and pedal ganglia, the ventral foot, and the captacula
Several cells in the region of the cerebral and pedal ganglia, the ventral foot, and the captacula
Cerebral, optic, and palliovisceral ganglia
Cerebral, optic, and palliovisceral ganglia
Inferior frontal lobes, precommissural lobes, anterior and posterior basal lobes, inferior buccal lobes,
Inferior frontal lobes, precommissural lobes, anterior and posterior basal lobes, inferior buccal lobes, peduncle lobes, and optic lobes
Region around eyes
Few cells in apical hemisphere in apical organ and cerebral ganglia
Cells of ventral plate during differentiation of trunk CNS
Two bilateral clusters of cells close to stomodeum
Cells in midgut and posterior foregut
Bilateral cell clusters in dorso-median episphere
Multiple bilaterally expression domains in dorsolateral episphere that persist during later larval development
Large cells at dorsal part of head at position of adult eyes
No expression in older juveniles
Embryo (stages 5-8)
Small domain of anterior CNS
ind (intermediate neuroblasts defective)
Intermediate column cells of developing CNS
In intermediate ectodermal domain of antennal segment
Dorsal ectodermal region of the ocular region
Intermediate column cells of developing CNS
Gastrula and subsequent larval stage
Two bilateral neuroectodermal domains
Provided as maternal message with no zygotic activation in subsequent developmental stages
Cells around blastopore of gastrula (disappear in tornaria larva)
CNS (neural tube, hindbrain, mesencephalon, diencephalon)
CNS (forebrain, midbrain, hindbrain)
In hindbrain rhombomeres
In mesencephalon, diencephalon, and intermediate spinal cord
Neuroectoderm (spinal cord, dorsal rhombencephalon, optic tectum, dorsal diencephalon, hypothalamus, rostral telencephalon)
Gsh-1 and Gsh-2
Anterior neural plate/CNS
Gsx is also involved in the development of the CNS in bilaterians, and it is expressed in distinct cells of the apical organ in the gastropod mollusk G. varia and the annelid P. dumerilii (Table 1; [12, 13]). In addition, Gsx expression was also found in the radula sac, a molluscan evolutionary novelty . Recent phylogenomic analyses on mollusks have revived a classical hypothesis placing the Aculifera, i.e., the worm-shaped and spicule-bearing aplacophorans and the eight-shelled polyplacophorans, as a sister group to the Conchifera [30–32]. The Conchifera is an anatomically diverse clade comprising scaphopods, gastropods, bivalves, monoplacophorans, and cephalopods. Until now, conchiferan interrelationships are unsettled, and attempts to infer the evolution of their body plans are scarce (c.f. [31, 32]; but see [33, 34]).
Ontogeny of the scaphopod Antalis entalis and the cephalopod Idiosepius notoides
In this study, we describe hitherto unknown Gsx orthologs and their expression domains in the scaphopod Antalis entalis and the cephalopod squid Idiosepius notoides (Fig. 1). Our results question the widely assumed role of Gsx in patterning the foregut of the last common bilaterian ancestor and highlight similarities as well as differences among mollusks, lophotrochozoans, and bilaterians.
Collection and culture of animals
Adults of the scaphopod Antalis entalis were collected from approximately 30 m depth by the staff of the research vessel Neomys off the coast of Roscoff (France). Individuals were immediately transferred into dishes filled with seawater (see also ). Spawning occurred spontaneously or was induced by heat shocks, i.e., individuals were exposed to alternating water temperatures. Unfertilized eggs were rinsed several times and fertilized with sperm. Early- and mid-stage trochophore larvae were cultured in Millipore-filtered seawater (MFSW) with 50 mg streptomycin sulfate and 60 mg penicillin G per liter MFSW. Early cleavage stages, metamorphic competent larvae, and settled individuals were cultured in MFSW without antibiotics. Water was changed every other day. Metamorphosis occurred spontaneously or was induced by adding shell-gravel from the collection site.
Adults of the pygmy squid Idiosepius notoides were dip-netted in the sea grass beds of Moreton Bay, Queensland, Australia. Embryos were cultured and staged as described previously . Development from freshly laid fertilized eggs (stage 1) to hatchlings (stage 30) takes 9–10 days at 25 °C.
RNA extraction and fixation of animals
For Antalis entalis, a total of several hundred individuals of mixed developmental stages including early cleavage stages, trochophore larvae, metamorphic competent individuals, and early juveniles were collected and stored at −20 to −80 °C in RNAlater (Lifetechnologies, Vienna, Austria). RNA was extracted with a RNA extraction kit (Qiagen, Roermond, Netherlands) and stored at −80 °C.
For Idiosepius notoides, the egg jelly and chorion were removed from approximately 300 specimens covering freshly laid zygotes (stage 1) to hatchlings (stage 30). RNA was extracted using TriReagent according to the manufacturer’s instructions (Astral Scientific Pty. Ltd., Caringbah, Australia, see also ). Individuals of all the above-described developmental stages were fixed for in situ hybridization experiments as previously described .
RNAseq and transcriptome assembly
Total RNA from pooled developmental stages of Antalis entalis was sequenced by Illumina technology (Eurofins, Ebersberg, Germany). Paired-end reads of an average read length of 100 bp were obtained and subsequently filtered (rRNA removal). Adapter and low-quality sequences were trimmed, normalized, and assembled de novo into contigs with the assembler Trinity .
RNA from developmental stages of Idiosepius notoides was sequenced by 454 and Illumina technology (both Eurofins) as described previously . After filtering, the adapter and low-quality reads were trimmed, normalized, and assembled de novo by Eurofins (454 transcriptome) or using Trinity (Illumina transcriptome).
Alignment and phylogenetic analysis
Molecular isolation of RNA transcripts
First-strand cDNA synthesis of the RNA pooled from different developmental stages of Antalis entalis and Idiosepius notoides, respectively, was carried out by reverse transcription using the First-strand cDNA Synthesis Kit for rt-PCR (Roche Diagnostics GmbH, Mannheim, Germany). Identified Gsx orthologs of A. entalis and I. notoides were used to design gene-specific primers, and PCR products were size-fractioned by gel electrophoresis. Gel bands of the expected length were excised and cleaned up using a QIAquick Gel Extraction Kit (QIAgen, Hilden, Germany). By insertion into pGEM-T Easy Vectors (Promega, Mannheim, Germany), cleaned-up products were cloned. Plasmid minipreps were grown overnight, cleaned up with the QIAprep Spin MiniprepKit (QIAgen), and sent for sequencing. The sequenced minipreps matched both transcripts identified as Aen-Gsx and Ino-Gsx in the phylogenetic analysis (Figs. 4, 5).
Probe synthesis and whole-mount in situ hybridization
Riboprobe templates were amplified via standard PCR from miniprepped plasmids using M13 forward and reverse primers. In vitro transcription reactions were performed with these templates, digoxigenin-UTP (DIG RNA Labeling Kit, Roche Diagnostics) and SP6/T7 polymerase (Roche Diagnostics GmbH) for the syntheses of antisense riboprobes according to the manufacturer’s instructions. For whole-mount in situ hybridization experiments, specimens were rehydrated into PBT (PBS + 0.1 % Tween-20) and treated with Proteinase-K (25 µg/ml for Idiosepius notoides and 45 µg/ml for Antalis entalis) in PBT at 37 °C for 10 min. Specimens were prehybridized in hybridization buffer for 4 h at 50 °C (A. entalis) or 65 °C (I. notoides), and hybridization with a probe concentration of 0.5 μg/ml (I. notoides) to 1 μg/ml (A. entalis) was carried out overnight at 50 °C (A. entalis) or 65 °C (I. notoides). For A. entalis as well as I. notoides, a minimum of 20 individuals per stage were investigated, and negative controls were carried out with sense probes for all genes and developmental stages. The majority of whole-mount preparations were cleared in a solution of benzyl benzoate/benzyl alcohol (2:1), mounted on objective slides, and analyzed. Preparations were documented with an Olympus BX53 Microscope (Olympus, Hamburg, Germany). In addition, scaphopod developmental stages were scanned with a Leica confocal SP5 II microscope (Leica Microsystems, Wetzlar, Germany) using bright-field, autofluorescence, and reflection mode scans . If necessary, images were processed with Adobe Photoshop 9.0.2 software (San Jose, CA, USA) to adjust contrast and brightness.
After in situ hybridization experiments, developmental stages of Antalis entalis were post-fixed in 100 % EtOH and embedded in agar low viscosity resin (Agar Scientific, Essex, United Kingdom). Specimens were semithin sectioned with a diamond knife (Histo Jumbo Diatome) at a thickness of 0.5 µm with an ultramicrotome (Leica EM UC6, Wetzlar, Germany). Sections were mounted on objective slides, stained with Eosin using standard histological protocols, and covered with cover slips. Alternatively, after in situ hybridization, specimens were embedded in O.C.T. medium (VWR, Vienna, Austria) and cut into 15–30 µm cryosections with a cryotome (Leica CM 3050S). Sections were stained with Dapi (Sigma-Aldrich, St. Louis, MO, USA) and Cellmask Green plasma membrane stain (ThermoFisher, Waltham, MA, USA) in order to stain cell nuclei and cell membranes. Sections were mounted in Fluoromount G (Southern Biotech, Birmingham, Alabama, USA) and covered with cover slips. Semithin as well as cryotome sections was documented with an Olympus BX53 Microscope (Olympus).
Statement of ethical approval
Developmental stages and adults of the pygmy squid Idiosepius notoides were collected, anesthetized, and fixed according to internationally recognized standards (University of Queensland Animal Welfare Permit No. 158/09 “The cultivation of Idiosepius (pygmy squid) for studies in developmental biology” to BMD).
Aen-Gsx expression in developmental stages of the scaphopod Antalis entalis
The alignment of multiple amino acid sequences shows that Aen-Gsx and Ino-Gsx exhibit high sequence similarity with their bilaterian orthologs (Fig. 4). Aen-Gsx as well as Ino-Gsx clusters with their bilaterian orthologs in the phylogenetic analysis (Fig. 5).
Ino-Gsx expression in Idiosepius notoides
Gsx does not pattern the digestive tract of scaphopods and cephalopods
To date, it is commonly hypothesized that the digestive tract of the last common bilaterian ancestor expressed Gsx in a collinear fashion together with the two other ParaHox genes, Cdx and Xlox [1, 6, 12, 20, 50]. This hypothesis is seemingly corroborated by the fact that among the Lophotrochozoa, the annelids Platynereis dumerilii and Nereis virens, as well as the gastropod Gibbula varia, express Gsx in their anterior digestive tract (Table 1; [12–14]). Our results for the scaphopod Antalis entalis and the cephalopod Idiosepius notoides, however, show that this is not the case for all mollusks, and therefore, neither for all lophotrochozoans, a scenario that was already suggested by data on the annelid Capitella teleta (; Table 1). Moreover, all ecdysozoan representatives investigated lack Gsx expression in their digestive tract, and among the deuterostomes investigated, only the hemichordate Ptychodera flava expresses Gsx around the blastopore . The lack of Gsx expression in the foregut of the other deuterostomes has been explained by the fate of the blastopore that does not transform into the definite mouth in deuterostomes as it does in protostomes, but, instead, into the anus . Accordingly, the latter hypothesis would argue for Gsx expression in the deuterostome hindgut which, however, appears to be absent (; Table 1). It is important to mention that Gsx orthologs have either not been found or are indeed absent in representatives of the Acoelomorpha, which are characterized by having a single mouth/anus opening in their digestive tract and may form the sister taxon to all remaining Bilateria (the so-called Nephrozoa; [51, 52]; but see  for a controversial view). In cnidarians, Gsx is endodermally expressed in the planula larva of Nematostella vectensis, Clytia hemisphaerica, and Podocoryne carnea [9–11]. In the coral Acropora millepora, Gsx is expressed in the ectoderm of the planula larva . Comparisons of the cnidarian and nephrozoan expression domains are difficult since mouth and digestive system cannot be easily homologized. Hence, the data currently available argue for a last common nephrozoan and probably also bilaterian ancestor without Gsx expression in the digestive tract and for a recruitment of Gsx into foregut patterning in selected lineages. Accordingly, the gastropod G. varia and the polychaete annelids N. virens and P. dumerilii have acquired Gsx expression in the foregut secondarily during evolution (Table 1). In contrast, other genes such as Brachyury, Nkx2.1, or FoxA appear to be evolutionary highly conserved in the digestive system within the Lophotrochozoa [54–59].
Gsx is expressed in the anterior-most portion of the molluscan CNS
In contrast to the digestive tract, Gsx is consistently expressed in the anterior CNS of bilaterians and hence an ancestral role in CNS development was proposed (Table 1; ). Shared Gsx expression domains among mollusks are the cerebral ganglia that subsequently develop into the supraesophageal mass in cephalopods (present study; ). In scaphopod and gastropod larvae, the apical organ is located in the anterior-most region. In the scaphopod Antalis entalis, Gsx is expressed in two flask-shaped cells of this organ and in two cells that are located laterally to it but do not constitute a part of the apical organ (Fig. 4d). With two apical tuft cells and further putative sensory cells, the larva of the gastropod Gibbula varia possesses more Gsx-expressing cells in the apical organ than the one of A. entalis (present study; ). The flask-shaped Gsx-expressing cells of A. entalis do not appear to be homologous to any of the Gsx-expressing cell types of G. varia judging by their morphology. However, detailed ultrastructural studies and molecular fingerprints on the various cell types occurring in lophotrochozoan apical organs are necessary to further assess homologies in this organ on the cellular level. Among all metazoans with an apical organ (Cnidaria, Ambulacraria, and Lophotrochozoa), only both above-mentioned mollusks and the annelid Platynereis dumerilii possess Gsx-expressing cells in the apical organ, suggesting that Gsx has been recruited into the patterning of this sensory organ in lophotrochozoans only (Table 1; present study; [12, 13]).
Gsx expression has also been reported for the polychaete annelids Nereis virens and Capitella teleta [14, 15]. As far as known, both species lack an apical organ as do cephalopods as direct developers (present study; [3, 15]). The vertical lobe as the anterior-most portion of the cephalopod CNS does not express Gsx (Figs. 9d, 10). This resembles the expression patterns of other homeobox genes such as Otx or the POU genes which are consistently expressed in the gastropod cerebral ganglia and large parts of the cephalopod cerebral ganglia/supraesophageal mass but not in the vertical lobe [44, 60, 61]. The vertical lobe is considered an evolutionary innovation of coleoid cephalopods, i.e., all cephalopods except the nautiluses as basal cephalopod offshoots . As an evolutionary younger brain region confined to coleoid cephalopods, the vertical lobe also differentiates relatively late during ontogeny compared to other brain regions . Hence, the vertical lobe probably evolved after Otx expression domains had already been established in the supraesophageal mass of coleoid cephalopods.
Gsx is expressed in the posterior portion of the molluscan CNS
Idiosepius notoides and Antalis entalis express Gsx in posterior portions of their CNS such as the scaphopod pedal ganglia and the cephalopod palliovisceral ganglia (the latter develop into the future posterior subesophageal mass). This is in contrast to the gastropod Gibbula varia and the annelid Capitella teleta, where Gsx expression is restricted to the anterior CNS [12, 15]. The scaphopod and cephalopod condition is, however, similar to the condition found in Platynereis dumerilii and certain vertebrates insofar that both mollusks and the polychaete express Gsx in more posterior regions of their nervous system. These domains comprise the scaphopod pedal ganglia, the cephalopod palliovisceral lobe/posterior subesophageal mass, the polychaete nerve cord, and the hindbrain of vertebrates [13, 24–27]. Interestingly, Gsx is also expressed in portions of the developing visual system of few representatives of all three bilaterian superphyla. The mollusks I. notoides and Nereis virens, the arthropods Drosophila melanogaster, as well as the teleost fish Oryzias latipes, express Gsx in portions of their visual system (Table 1; present study; [14, 17, 28]). Further studies on other bilaterian representatives are needed to assess if Gsx expression in the eyes and related brain regions may be an ancestral trait among nephrozoans or bilaterians.
This study suggests that Gsx expression in the foregut is not a molluscan plesiomorphy and together with already published data argues against Gsx expression in the foregut of the last common bilaterian ancestor. It is therefore most likely that Gsx has been independently recruited into the development of the foregut in some lophotrochozoan representatives. Gsx is consistently expressed in the developing anterior nervous system of bilaterians, which is probably an apomorphy of Bilateria. In contrast to other metazoan taxa, Gsx expression was only found in the larval apical organ in lophotrochozoans, indicating that Gsx expression in the apical organ may be a lophotrochozoan synapomorphy.
Anterior basal lobe
- Aen :
- A. entalis :
Central nervous system
Dorsal basal lobe
- Gsx :
Genomic screened homeobox protein
- G. varia :
Hours after fertilization
Inferior buccal lobe
Inferior frontal lobe
Intermediate neuroblasts defective
- Ino :
- I. notoides :
Median basal lobe
Middle subesophageal mass
National Center for Biotechnology Information
- P. dumerilii :
Posterior basal lobe
Posterior subesophageal mass
TW designed the project together with AW. TW reared and fixed all developmental stages of Idiosepius notoides and Antalis entalis, extracted the RNA, and assembled the transcriptomes. TW cloned all genes, and TW and SVRM carried out the in situ hybridization experiments. CMD performed the phylogenetic analysis. TW analyzed all data and drafted the manuscript. AW contributed to data interpretation and writing of the manuscript. BMD commented on a later version of the manuscript. All authors read and approved the final version of the manuscript.
The staff from the research vessel Neomys (SBR in Roscoff) is thanked for collecting adult scaphopods. Thomas Eder and Thomas Rattei (Vienna) are thanked for advice with transcriptome assembly. Julia Bauder is thanked for help with semithin sectioning. We kindly thank three anonymous reviewers for their constructive criticism. During his stay at the SBR in Roscoff (France), TW was generously supported by the Faculty of Life Science, University of Vienna, and an ASSEMBLE (Association of European Marine Biological Laboratories) Grant. This work was also supported by Grants to BMD of the Australian Research Council. Research in the lab of AW also benefitted from a Grant of the Austrian Science Fund (FWF) on comparative aspects of molluscan EvoDevo to AW (Grant number: P24276-B22).
The authors declare that they have no competing interests.
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- Holland PWH. Evolution of homeobox genes. WIREs Dev Biol. 2013;2:31–45.View ArticleGoogle Scholar
- Duboule D, Dollé P. The structural and functional organization of the murine Hox gene family resembles that of Drosophila homeotic genes. EMBO J. 1989;8:1497–505.PubMedPubMed CentralGoogle Scholar
- Kulakova MA, Bakalenko N, Novikova E, Cook CE, Eliseeva E, Steinmetz PRH, Kostyuchenko RP, Dondua A, Arendt D, Akam M, Andreeva T. Hox gene expression in the larval development of the polychaetes Nereis virens and Platynereis dumerilii (Annelida, Lophotrochozoa). Dev Genes Evol. 2007;217:39–54.PubMedView ArticleGoogle Scholar
- Lee PN, Callaerts P, de Couet HG, Martindale MQ. Cephalopod Hox genes and the origin of morphological novelties. Nature. 2003;424:1061–5.PubMedView ArticleGoogle Scholar
- Samadi L, Steiner G. Involvement of Hox genes in shell morphogenesis in the encapsulated development of a top shell gastropod (Gibbula varia L.). Dev Genes Evol. 2009;219:523–30.PubMedView ArticleGoogle Scholar
- Holland PW. Beyond the Hox: how widespread is homeobox gene clustering? J Anat. 2001;199:13–24.PubMedPubMed CentralView ArticleGoogle Scholar
- Ikuta T, Chen YC, Annunziata R, Ting HC, Tung CH, Koyanagi R, Tagawa K, Humphreys T, Fujiyama A, Saiga H, Satoh N, Yu IK, Arnone MI, Su YH. Identification of an intact ParaHox cluster with temporal collinearity but altered spatial collinearity in the hemichordate Ptychodera flava. BMC Evol Biol. 2013;13:129.PubMedPubMed CentralView ArticleGoogle Scholar
- Hayward DC, Catmull J, Reece-Hoyes JS, Berghammer H, Dodd H, Hann SJ, Miller DJ, Ball EE. Gene structure and larval expression of cnox-2Am from the coral Acropora millepora. Dev Genes Evol. 2001;211:10–9.PubMedView ArticleGoogle Scholar
- Finnerty JR, Paulson D, Burton P, Pang K, Martindale MQ. Early evolution of a homeobox gene: the parahox gene Gsx in the Cnidaria and the Bilateria. Evol Dev. 2003;5:331–45.PubMedView ArticleGoogle Scholar
- Quiquand M, Yanze N, Schmich J, Schmid V, Galliot B, Piraino S. More constraint on ParaHox than Hox gene families in early metazoan evolution. Dev Biol. 2009;328:173–87.PubMedView ArticleGoogle Scholar
- Yanze N, Spring J, Schmidli C, Schmid V. Conservation of Hox/ParaHox-related genes in the early development of a cnidarian. Develop Biol. 2001;236:89–98.PubMedView ArticleGoogle Scholar
- Samadi L, Steiner G. Conservation of ParaHox genes’s function in patterning of the digestive tract of the marine gastropod Gibbula varia. BMC Dev Biol. 2010;10:74.PubMedPubMed CentralView ArticleGoogle Scholar
- Hui JHL, Raible F, Korchagina N, Dray N, Samain S, Magdelenat G, Jubin C, Segurens B, Balavoine G, Arendt D, Ferrier DEK. Features of the ancestral bilaterian inferred from Platynereis dumerilii ParaHox genes. BMC Biol. 2009;7:43.PubMedPubMed CentralView ArticleGoogle Scholar
- Kulakova MA, Cook CE, Andreeva TF. ParaHox gene expression in larval and postlarval development of the polychaete Nereis virens (Annelida, Lophotrochozoa). BMC Dev Biol. 2008;8:61.PubMedPubMed CentralView ArticleGoogle Scholar
- Fröbius AC, Seaver EC. Capitella sp. I homeobrain-like, the first lophotrochozoan member of a novel paired-like homeobox gene family. Gene Expr Patt. 2006; 6:895–91.
- Weiss JB, Von Ohlen T, Mellerick DM, Dressler G, Doe CQ, Scott MP. Dorsoventral patterning in the Drosophila central nervous system: the intermediate neuroblasts defective homeobox gene specifies intermediate column identity. Genes Dev. 1998;12:3591–602.PubMedPubMed CentralView ArticleGoogle Scholar
- Urbach R, Technau GM. Segment polarity and DV patterning gene expression reveals segmental organization of the Drosophila brain. Development. 2003;130:3607–20.PubMedView ArticleGoogle Scholar
- Wheeler SR, Carrico ML, Wilson BA, Skeath JB. The Tribolium columnar genes reveal conservation and plasticity in neural precursor patterning along the embryonic dorsal-ventral axis. Genomes Dev Control. 2005;279:491–500.Google Scholar
- Arnone MI, Rizzo F, Annunciata R, Cameron RA, Peterson KJ, Martínez P. Genetic organization and embryonic expression of the ParaHox genes in the sea urchin S. purpuratus: insights into the relationship between clustering and collinearity. Dev Biol. 2006;300:63–73.PubMedView ArticleGoogle Scholar
- Annunziata R, Martinez P, Arnone MI. Intact cluster and chordate-like expression of ParaHox genes in a sea star. BMC Biol. 2013;11:68.PubMedPubMed CentralView ArticleGoogle Scholar
- Brooke NM, Garcia-Fernàndez J, Holland PWH. The ParaHox gene cluster is an evolutionary sister of the Hox gene cluster. Nature. 1998;392:920–2.PubMedView ArticleGoogle Scholar
- Osborne PW, Benoit G, Laudet V, Schubert M, Ferrier DEK. Differential regulation of ParaHox genes by retinoic acid in the invertebrate chordate amphioxus (Branchiostoma floridae). Dev Biol. 2009;327:252–62.PubMedView ArticleGoogle Scholar
- Hudson C, Lemaire P. Induction of anterior neural fates in the ascidian Ciona intestinalis. Mech Dev. 2001;100:189–203.PubMedView ArticleGoogle Scholar
- Hsieh-Li HM, Witte DP, Szucsik JC, Weinstein M, Li H, Potter SS. Gsh-2, a murine homeobox gene expressed in the developing brain. Mech Dev. 1995; 50:177–86.
- Valerius MT, Li H, Stock JL, Weinstein M, Kaur S, Singh G, Potter SS. Gsh-1: a novel murine homeobox gene expressed in the central nervous system. Dev Dyn. 1995;203:337–51.PubMedView ArticleGoogle Scholar
- Szucsik JC, Witte DP, Li H, Pixley SK, Small KM, Potter SS. Altered forebrain and hindbrain development in mice mutant for the Gsh-2 homeobox gene. Dev Biol. 1997;191:230–42.PubMedView ArticleGoogle Scholar
- Cheesman SE, Eisen SE. Gsh1 demarcates hypothalamus and intermediate spinal cord in zebrafish. Gene Expr Patt. 2004;5:107–12.View ArticleGoogle Scholar
- Deschet K, Bourrat F, Chourrout D, Joly JS. Expression domains of the medaka (Oryzias latipes) Ol-Gsh 1 gene are reminiscent of those of clustered and orphan homeobox genes. Dev Genes Evol. 1998;208:235–44.PubMedView ArticleGoogle Scholar
- Illes CJ, Winterbottom E, Isaacs HV. Cloning and expression analysis of the anterior ParaHox genes, Gsh1 and Gsh2 from Xenopus tropicalis. Dev Dyn. 2009;238:194–203.PubMedView ArticleGoogle Scholar
- Scheltema AH. Aplacophora as progenetic aculiferans and the coelomate origin of mollusks as the sister taxon of Sipuncula. Biol Bull. 1993;184:57–78.View ArticleGoogle Scholar
- Kocot KM, Cannon JT, Todt C, Citarella MR, Kohn AB, Meyer A, Santos SR, Schander C, Moroz LL, Lieb B, Halanych KM. Phylogenomics reveals deep molluscan relationships. Nature. 2011;477:452–6.PubMedPubMed CentralView ArticleGoogle Scholar
- Smith SA, Wilson NG, Goetz FE, Feehery C, Andrade SCS, Rouse GW, Giribet G, Dunn CW. Resolving the evolutionary relationships of molluscs with phylogenomic tools. Nature. 2011;480:364–9.PubMedView ArticleGoogle Scholar
- Wanninger A, Haszprunar G. Muscle development in Antalis entalis (Mollusca, Scaphopoda) and its significance for scaphopod relationships. J Morph. 2002;254:53–64.PubMedView ArticleGoogle Scholar
- Steiner G, Dreyer H. Molecular phylogeny of Scaphopoda (Mollusca) inferred from 18S rDNA sequences: support for a Scaphopoda-Cephalopoda clade. Zool Scripta. 2003;32:343–56.View ArticleGoogle Scholar
- Shigeno S, Sasaki T, Moritaki T, Kasugai T, Vecchione M, Agata K. Evolution of the cephalopod head complex by assembly of multiple molluscan body parts: evidence from Nautilus embryonic development. J Morph. 2008;269:1–17.PubMedView ArticleGoogle Scholar
- Van Dongen CAM, Geilenkirchen WLM. The development of Dentalium with special reference to the significance of the polar lobe. I, II, III. Division chronology and development of the cell pattern in Dentalium dentale (Scaphopoda). Proc Kongl Ned Akad v Wet Ser C. 1974;77:57–100.
- Van Dongen CAM, Geilenkirchen WLM. The development of Dentalium with special reference to the significance of the polar lobe. IV. Division chronology and development of the cell pattern in Dentalium dentale after removal of the polar lobe at first cleavage. Proc Kongl Ned Akad v Wet Ser C. 1975;78:358–75.
- Van Dongen CAM, Geilenkirchen WLM. The development of Dentalium with special reference to the significance of the polar lobe. V and VI. Differentiation of the cell pattern in lobeless embryos of Dentalium vulgare (da Costa) during late larval development. Proc Kongl Ned Akad v Wet Ser C. 1976;79:245–66.
- Wanninger A, Haszprunar G. The expression of an engrailed protein during embryonic shell formation of the tusk-shell, Antalis entalis (Mollusca, Scaphopoda). Evol Dev. 2001;3:312–21.PubMedView ArticleGoogle Scholar
- Wanninger A, Haszprunar G. The development of the serotonin-like immunoreactive and FMRFamidergic nervous system in Antalis entalis (Mollusca, Scaphopoda). Zoomorph. 2003;122:77–85.Google Scholar
- Yamamoto M. Normal embryonic stages of the pygmy cuttlefish, Idiosepius pygmaeus paradoxus Ortmann. Zool Sci. 1988;5:989–98.Google Scholar
- Wanninger A, Wollesen T. Mollusca. In: Wanninger A, editor. Evolutionary developmental biology of invertebrates, vol. 2 Lophotrochozoa (Lophotrochozoa). Wien: Springer; 2015. pp. 103–53.
- Wollesen T, Cummins SF, Degnan BM, Wanninger A. FMRFamide gene and peptide expression during central nervous system development of the cephalopod Idiosepius notoides. Evol Dev. 2010;12:113–30.PubMedView ArticleGoogle Scholar
- Wollesen T, McDougall C, Degnan BM, Wanninger A. POU genes are expressed during the formation of individual ganglia of the cephalopod central nervous system. EvoDevo. 2014;5:41.PubMedPubMed CentralView ArticleGoogle Scholar
- Grabherr MG, Haas BJ, Yassour M, Levin JZ, Thompson DA, Amit I, Adiconis X, Fan L, Raychowdhury R, Zeng Q, Chen Z, Mauceli E, Hacohen N, Gnirke A, Rhind A, Rhind N, di Palma F, Birren BW, Nusbaum C, Lindblad-Toh K, Friedman N, Regev A. Full-length transcriptome assembly from RNA-seqdata without a reference genome. Nat Biotechnol. 2011;15:644–52.View ArticleGoogle Scholar
- Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R, Thompson JD, Gibson TJ, Higgins DG. Clustal W and Clustal X version 2.0. Bioinformatics. 2007;23:2947–8.PubMedView ArticleGoogle Scholar
- Larsson A. AliView: a fast and lightweight alignment viewer and editor for large data sets. Bioinformatics. 2014;30:3276–8.PubMedPubMed CentralView ArticleGoogle Scholar
- Felsenstein J. PHYLIP—Phylogeny Inference Package (Version 3.2). Cladistics. 1989;5:164–6.Google Scholar
- Jékely G, Arendt D. Confocal detection of NBT/BCIP in situ hybridization samples by reflection microscopy. Biochemica. 2007;4:12–4.Google Scholar
- Grapin-Botton A. Antero-posterior patterning of the vertebrate digestive tract: 40 years after Nicole Le Douarin´s Ph.D. thesis. Int J Dev Biol. 2005;49:335–47.
- Hejnol A, Obst M, Stamatakis A, Ott M, Rouse GW, Edgecombe GD, Martinez P, Baguna J, Bailly X, Jondelius U, Wiens M, Müller WE, Seaver E, Wheeler WC, Martindale MQ, Giribet G, Dunn CW. Assessing the root of bilaterians animals with scalable phylogenomic methods. Proc Roy Soc B Biol Sci. 2009;276:4261–70.View ArticleGoogle Scholar
- Moreno E, Permanyer J, Martinez P. The origin of patterning systems in Bilateria—insights from the Hox and ParaHox genes in Acoelomorpha. Genomics Proteomics Bioinform. 2011;9:65–76.View ArticleGoogle Scholar
- Philippe H, Brinkmann H, Copley R, Moroz LL, Nakano H, Poustka AJ, Wallberg A, Peterson KJ, Telford MJ. Acoelomorph flatworms are deuterostomes related to Xenoturbella. Nature. 2011;470:255–60.PubMedPubMed CentralView ArticleGoogle Scholar
- Arendt D, Technau U, Wittbrodt J. Evolution of the bilaterian larval foregut. Nature. 2001;409:81–5.PubMedView ArticleGoogle Scholar
- Boyle MJ, Seaver EC. Developmental expression of FoxA and GATA genes during gut formation in the polychaete annelid. Capitella sp I Evol Dev. 2008;10:89–105.PubMedView ArticleGoogle Scholar
- Boyle MJ, Seaver EC. Expression of FoxA and GATA transcription factors correlates with regionalized gut development in two lophotrochozoan marine worms: Chaetopterus (Annelida) and Themiste lageniformis (Sipuncula). EvoDevo. 2010;1:2.PubMedPubMed CentralView ArticleGoogle Scholar
- Boyle MJ, Yamaguchi E, Seaver EC. Molecular conservation of metazoan gut formation: evidence from expression of endomesoderm genes in Capitella teleta (Annelida). EvoDevo. 2014;5:39.PubMedPubMed CentralView ArticleGoogle Scholar
- Martin-Duran JM, Amaya E, Romero R. Germ layer specification and axial patterning in the embryonic development of the freshwater planarian Schmidtea polychroa. Dev Biol. 2010;340:145–58.PubMedView ArticleGoogle Scholar
- Martin-Duran JM, Romero R. Evolutionary implications of morphogenesis and molecular patterning of the blind gut in the planarian Schmidtea polychroa. Dev Biol. 2011;352:164–76.PubMedView ArticleGoogle Scholar
- O’Brien EK, Degnan BM. Pleiotropic developmental expression of HasPOU-III, a class III POU gene, in the gastropod Haliotis asinina. Mech Dev. 2002;114:129–32.PubMedView ArticleGoogle Scholar
- Buresi A, Baratte S, Da Silva C, Bonnaud L. Orthodenticle/otx ortholog expression in the anterior brain and eyes of Sepia officinalis (Mollusca, Cephalopoda). Gene Expr Patt. 2012;12:109–16.View ArticleGoogle Scholar
- Nixon M, Young JZ. The Brains and Lives of Cephalopods. New York: Oxford University Press; 2003.Google Scholar
- Shigeno S, Tsuchiya K, Segawa S. Embryonic and paralarval development of the central nervous system of the loliginid squid Sepioteuthis lessoniana. J Comp Neurol. 2001;437:449–75.PubMedView ArticleGoogle Scholar