Larval convergence in a colonial tunicate: the organization of the sarcotubular complex in <Emphasis Type="Italic">Ecteinascidia turbinata</Emphasis> (Perophoridae,Phlebobranchiata, Tunicata,Chordata)

Ecteinascidia turbinata is a colonial ascidian that as an adult shares characters with phlebobranch ascidians, whereas the larvae are similar to aplousobranch ascidian larvae. The sarcotubular complex consists of invaginations of the sarcolemma that contact the sarcoplasmatic reticulum via dyads or triads. If present, the invaginations of the sarcolemma in tunicates have been characterized as laminar or tubular. We comparatively investigated the sarcotubular complex of E. turbinata and seven other tunicate species using 3D-reconstruction techniques based on electron micrographs of serial sections. The mononucleate muscle cells in E. turbinata possess intermediate and close junctions and contain several layers of peripheral myofibrillae. The myofibrillae are surrounded by continuous cisternae of the sarcoplasmic reticulum that forms interconnected rings around the z-bands. The invaginations of the sarcolemma are laminar, contacting the sarcoplasamatic reticulum at the height of the z-bands via dyads and triads. We present a clear definition of character states encountered in Tunicata: laminar invaginations are characterized by a width to length ratio of smaller than 1:20, tubular invagination by a width to length ratio of larger than 1:10. Laminar invaginations are found in stolidobranch ascidians and E. turbinata. Tubular invaginations are present in aplousobranch ascidians and appendicularians. This character state distribution across taxa supports the hypothesis that E. turbinata should be included in Phlebobranchiata as suggested by adult characters and that the larval similarities with Aplousobranchiata arose by convergent evolution. An erratum to this article can be found at     

Nervous and muscle system development in <Emphasis Type="Italic">Phascolion strombus</Emphasis> (Sipuncula)

Recent interpretations of developmental gene expression patterns propose that the last common metazoan ancestor was segmented, although most animal phyla show no obvious signs of segmentation. Developmental studies of non-model system trochozoan taxa may shed light on this hypothesis by assessing possible cryptic segmentation patterns. In this paper, we present the first immunocytochemical data on the ontogeny of the nervous system and the musculature in the sipunculan Phascolion strombus. Myogenesis of the first anlagen of the body wall ring muscles occurs synchronously and not subsequently from anterior to posterior as in segmented spiralian taxa (i.e. annelids). The number of ring muscles remains constant during the initial stages of body axis elongation. In the anterior-posteriorly elongated larva, newly formed ring muscles originate along the entire body axis between existing myocytes, indicating that repeated muscle bands do not form from a posterior growth zone. During neurogenesis, the Phascolion larva expresses a non-metameric, paired, ventral nerve cord that fuses in the mid-body region in the late-stage elongated larva. Contrary to other trochozoans, Phascolion lacks any larval serotonergic structures. However, two to three FMRFamide-positive cells are found in the apical organ. In addition, late larvae show commissure-like neurones interconnecting the two ventral nerve cords, while early juveniles exhibit a third, medially placed FMRFamidergic ventral nerve. Although we did not find any indications for cryptic segmentation, certain neuro-developmental traits in Phascolion resemble the conditions found in polychaetes (including echiurans) and myzostomids and support a close relationship of Sipuncula and Annelida.     

Molecular phylogeny of horsetails (<Emphasis Type="Italic">Equisetum</Emphasis>) including chloroplast <Emphasis Type="Italic">atpB</Emphasis> sequences

Equisetum is a genus of 15 extant species that are the sole surviving representatives of the class Sphenopsida. The generally accepted taxonomy of Equisetum recognizes two subgenera: Equisetum and Hippochaete. Two recent phylogenetical studies have independently questioned the monophyly of subgenus Equisetum. Here, I use original (atpB) and published (rbcL, trnL-trnF, rps4) sequence data to investigate the phylogeny of the genus. Analyses of atpB sequences give an unusual topology, with E. bogotense branching within Hippochaete. A Bayesian analysis based on all available sequences yields a tree with increased resolution, favoring the sister relationships of E. bogotense with subgenus Hippochaete.     

The expanded cattle <Emphasis Type="Italic">KIR</Emphasis> genes are orthologous to the conserved single-copy <Emphasis Type="Italic">KIR3DX1</Emphasis> gene of primates

Cattle are the only non-primate species for which expansion of the killer cell immunoglobulin-like receptor (KIR) genes has been reported. We analyzed cattle KIR sequences to determine their relationship to the two divergent lineages of primate KIR: one comprising the KIR3DX1 gene of unknown function, the second comprising all other primate KIR genes, which encode variable major histocompatibility complex class I receptors. Phylogenetics and analysis of repetitive elements shows that cattle KIR subdivide into the same two lineages as primate KIR. Unlike the primates, the lineage of variable and likely functional cattle KIR corresponds to the KIR3DX1 lineage of primate KIR, whereas the variable lineage of primate KIR is represented in cattle by one KIR gene and a related gene fragment.     

Mitochondrial <Emphasis Type="Italic">cox</Emphasis>1 and plastid <Emphasis Type="Italic">rbc</Emphasis>L genes of <Emphasis Type="Italic">Gracilaria vermiculophylla</Emphasis> (Gracilariaceae,Rhodophyta)

The mitochondrial cytochrome c oxidase subunit I gene sequence was recently developed for DNA barcoding of red algal species. We determined the 1245 base pairs of the gene from 27 taxa of an agar-producing species, Gracilaria vermiculophylla, and putative relatives and compared the results with rbcL data from the same species. A total of 392 positions (31.5%) were variable, 282 positions (22.6%) were parsimoniously informative, and average sequence divergence was 13% in an ingroup. Within G. vermiculophylla, pairwise divergence of the gene was variable up to 11 bp (0.9%). Seven recognized haplotypes of cox1 tended to be geographically related. In the aligned 1386 bp of rbcL, three haplotypes were recognized. These results suggest that cox1 is a valuable molecular marker within species and will be very useful in haplotype analyses.     

Expression of <Emphasis Type="Italic">Pax group III</Emphasis> genes in the honeybee (<Emphasis Type="Italic">Apis mellifera</Emphasis>)

Pax group III genes are involved in a number of processes during insect segmentation. In Drosophila melanogaster, three genes, paired, gooseberry and gooseberry-neuro, regulate segmental patterning of the epidermis and nervous system. Paired acts as a pair-rule gene and gooseberry as a segment polarity gene. Studies of Pax group III genes in other insects have indicated that their expression is a good marker for understanding the underlying molecular mechanisms of segmentation. We have cloned three Pax group III genes from the honeybee (Apis mellifera) and examined their relationships to other insect Pax group III genes and their expression patterns during honeybee segmentation. The expression pattern of the honeybee homologue of paired is similar to that of paired in Drosophila, but its expression is modulated by anterior–posterior temporal patterning similar to the expression of Pax group III proteins in Tribolium. The expression of the other two Pax group III genes in the honeybee indicates that they also act in segmentation and nervous system development, as do these genes in other insects.     

Functional conservation of the human <Emphasis Type="Italic">EXT1</Emphasis> tumor suppressor gene and its <Emphasis Type="Italic">Drosophila</Emphasis> homolog <Emphasis Type="Italic">tout</Emphasis><Emphasis Type="Italic">velu</Emphasis>

Heparan sulfate proteoglycans play a vital role in signaling of various growth factors in both Drosophila and vertebrates. In Drosophila, mutations in the tout velu (ttv) gene, a homolog of the mammalian EXT1 tumor suppressor gene, leads to abrogation of glycosaminoglycan (GAG) biosynthesis. This impairs distribution and signaling activities of various morphogens such as Hedgehog (Hh), Wingless (Wg), and Decapentaplegic (Dpp). Mutations in members of the exostosin (EXT) gene family lead to hereditary multiple exostosis in humans leading to bone outgrowths and tumors. In this study, we provide genetic and biochemical evidence that the human EXT1 (hEXT1) gene is conserved through species and can functionally complement the ttv mutation in Drosophila. The hEXT1 gene was able to rescue a ttv null mutant to adulthood and restore GAG biosynthesis.     

Conserved and novel <Emphasis Type="Italic">Wnt</Emphasis> clusters in the basal eumetazoan <Emphasis Type="Italic">Nematostella vectensis</Emphasis>

Evolutionarily conserved gene clusters are interesting for two reasons: (1) they may illuminate ancient events in genome evolution and (2) they may reveal ongoing stabilizing selection; that is, the conservation of gene clusters may have functional significance. To test if the Wnt family of signaling factors exhibits conserved clustering in basal metazoans and if those clusters are of functional importance, we searched the genomic sequence of the sea anemone Nematostella vectensis for Wnt clusters and correlated the clustering we observed with published expression patterns. Our results indicate that the Wnt1–Wnt6–Wnt10 cluster observed in Drosophila melanogaster is partially conserved in the cnidarian lineage; Wnt6 and Wnt10 are separated by less than 4,500 nucleotides in Nematostella. A novel cluster comprised of Wnt5–Wnt7/Wnt7b was observed in Nematostella. Clustered Wnt genes do not exhibit Hox-like colinearity nor is the expression of linked Wnt genes more similar than the expression of nonlinked Wnt genes. Wnt6 and Wnt10 are not expressed in a spatially or temporally contiguous manner, and Wnt5 and Wnt7 are expressed in different germ layers.     

Conservation of Nuclear SSR Loci Reveals High Affinity of <Emphasis Type="Italic">Quercus infectoria</Emphasis> ssp. <Emphasis Type="Italic">veneris</Emphasis> A. Kern (Fagaceae) to Section <Emphasis Type="Italic">Robur</Emphasis>

Conservation of 16 nuclear microsatellite loci, originally developed for Quercus macrocarpa (section Albae), Q. petraea, Q. robur (section Robur), and Q. myrsinifolia, (subgenus Cyclobalanopsis) was tested in a Q. infectoria ssp. veneris population from Cyprus. All loci could be amplified successfully and displayed allele size and diversity patterns that match those of oak species belonging to the section Robur. At least in one case, limited amplification and high levels of homozygosity support the occurrence of “null alleles” caused by a possible mutation in the highly conserved primer areas, thus hindering PCR. The sampled population exhibited high levels of diversity despite the very limited distribution of this species in Cyprus and extended population fragmentation. Allele sizes of Q. infectoria at locus QpZAG9 partially match those of Q. alnifolia and Q. coccifera from neighboring populations. However, sequencing showed homoplasy, excluding a case of interspecific introgression with the latter, phylogenetically remote species. Q. infectoria ssp. veneris sequences at this locus were concordant to those of other species of section Robur, while sequences of Quercus alnifolia and Quercus coccifera were almost identical to Q. cerris.     

Phylogenetic analyses of <Emphasis Type="Italic">Uromyces viciae-fabae</Emphasis> and its varieties on <Emphasis Type="Italic">Vicia</Emphasis>, <Emphasis Type="Italic">Lathyrus</Emphasis>, and <Emphasis Type="Italic">Pisum</Emphasis> in Japan

A pea rust fungus, Uromyces viciae-fabae, has been classified into two varieties, var. viciae-fabae and var. orobi, based on differences in urediniospore wall thickness and putative host specificity in Japan. In principal component analyses, morphological features of urediniospores and teliospores of 94 rust specimens from Vicia, Lathyrus, and Pisum did not show definite host-specific morphological groups. In molecular analyses, 23 Uromyces specimens from Vicia, Lathyrus, and Pisum formed a single genetic clade based on D1/D2 and ITS regions. Four isolates of U. viciae-fabae from V. cracca and V. unijuga could infect and sporulate on P. sativum. These results suggest that U. viciae-fabae populations on different host plants are not biologically differentiated into groups that can be recognized as varieties.Contribution no. 184, Laboratory of Plant Parasitic Mycology, Institute of Agriculture and Forestry, University of Tsukuba, Japan     

<Emphasis Type="Italic">Drosophila</Emphasis><Emphasis Type="Italic">P</Emphasis> transposons of the urochordata <Emphasis Type="Italic">Ciona intestinalis</Emphasis>

P transposons belong to the eukaryotic DNA transposons, which are transposed by a cut and paste mechanism using a P-element-coded transposase. They have been detected in Drosophila, and reside as single copies and stable homologous sequences in many vertebrate species. We present the P elements Pcin1, Pcin2 and Pcin3 from Ciona intestinalis, a species of the most primitive chordates, and compare them with those from Ciona savignyi. They showed typical DNA transposon structures, namely terminal inverted repeats and target site duplications. The coding region of Pcin1 consisted of 13 small exons that could be translated into a P-transposon-homologous protein. C. intestinalis and C. savignyi displayed nearly the same phenotype. However, their P elements were highly divergent and the assumed P transposase from C. intestinalis was more closely related to the transposase from Drosophila melanogaster than to the transposase of C. savignyi. The present study showed that P elements with typical features of transposable DNA elements may be found already at the base of the chordate lineage. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Cleavage in<Emphasis Type="Italic"> Nemertoderma westbladi</Emphasis> (Nemertodermatida) and its phylogenetic significance

Recent phylogenetic analyses of ribosomal and protein coding nuclear genes place the marine worms within the Nemertodermatida as one of the oldest lineages among the bilaterian animals. We studied the early embryonic cleavage in Nemertoderma westbladi to provide the first account of nemertodermatid early development. Live embryos were studied with interference microscopy and fixed embryos were either sectioned or studied with confocal laser scanning microscopy. Initially the divisions in the embryo are radial, but then micromeres are shifted clockwise generating a spiral pattern. The four-cell stage is characterized by duets of macromeres and micromeres and thus resembles the duet cleavage reported from members of the Acoela. However, subsequent stages differ from the acoel duet pattern and also from quartet spiral cleavage. The optimization of the cleavage pattern on current phylogenetic hypotheses with Nemertodermatida and Acoela as early bilaterian branches is discussed.     

New combinations and typifications in <Emphasis Type="Italic">Bistorta</Emphasis>, <Emphasis Type="Italic">Persicaria</Emphasis>, <Emphasis Type="Italic">Polygonum,</Emphasis> and <Emphasis Type="Italic">Rumex</Emphasis> (Polygonaceae)

New combinations are proposed in anticipation of the Polygonaceae treatment in the forthcoming volume of Intermountain Flora: Polygonum kelloggii var. esotericum, P. kelloggii var. watsonii , Rumex densiflorus var. pycnanthus , R. salicifolius var. utahensis, and R. occidentalis var. tomentellus. Typifications are proposed to facilitate ongoing studies in Polygonaceae and to maintain current usage.     

<Emphasis Type="Italic">Ty</Emphasis>1<Emphasis Type="Italic">/copia</Emphasis>- and <Emphasis Type="Italic">Ty</Emphasis>3<Emphasis Type="Italic">/gypsy</Emphasis>-like DNA sequences in <Emphasis Type="Italic">Helianthus </Emphasis>species

Two repeated DNA sequences isolated from a partial genomic DNA library of Helianthus annuus, p HaS13 and p HaS211, were shown to represent portions of the int gene of a Ty3 /gypsy retroelement and of the RNase-Hgene of a Ty1 /copia retroelement, respectively. Southern blotting patterns obtained by hybridizing the two probes to BglII- or DraI-digested genomic DNA from different Helianthus species showed p HaS13 and p HaS211 were parts of dispersed repeats at least 8 and 7 kb in length, respectively, that were conserved in all species studied. Comparable hybridization patterns were obtained in all species with p HaS13. By contrast, the patterns obtained by hybridizing p HaS211 clearly differentiated annual species from perennials. The frequencies of p HaS13- and p HaS211-related sequences in different species were 4.3x10(4)-1.3x10(5) copies and 9.9x10(2)-8.1x10(3) copies per picogram of DNA, respectively. The frequency of p HaS13-related sequences varied widely within annual species, while no significant difference was observed among perennial species. Conversely, the frequency variation of p HaS211-related sequences was as large within annual species as within perennials. Sequences of both families were found to be dispersed along the length of all chromosomes in all species studied. However, Ty3 /gypsy-like sequences were localized preferentially at the centromeric regions, whereas Ty1/ copia-like sequences were less represented or absent around the centromeres and plentiful at the chromosome ends. These findings suggest that the two sequence families played a role in Helianthusgenome evolution and species divergence, evolved independently in the same genomic backgrounds and in annual or perennial species, and acquired different possible functions in the host genomes.     

The involvement of <Emphasis Type="Italic">engrailed</Emphasis> and <Emphasis Type="Italic">wingless</Emphasis> during segmentation in the onychophoran <Emphasis Type="Italic">Euperipatoides kanangrensis</Emphasis> (Peripatopsidae: Onychophora) (Reid 1996)

As the putative sister group to the arthropods, onychophorans can provide insight into ancestral developmental mechanisms in the panarthropod clade. Here, we examine the expression during segmentation of orthologues of wingless (Wnt1) and engrailed, two genes that play a key role in defining segment boundaries in Drosophila and that appear to play a role in segmentation in many other arthropods. Both are expressed in segmentally reiterated stripes in all forming segments except the first (brain) segment, which only shows an engrailed stripe. Engrailed is expressed before segments are morphologically visible and is expressed in both mesoderm and ectoderm. Segmental wingless expression is not detectable until after mesodermal somites are clearly distinct. Early engrailed expression lies in and extends to both sides of the furrow that first demarcates segments in the ectoderm, but is largely restricted to the posterior part of somites. Wingless expression lies immediately anterior to engrailed expression, as it does in many arthropods, but there is no precise cellular boundary between the two expression domains analogous to the overt parasegment boundary seen in Drosophila. Engrailed stripes extend along the posterior part of each limb bud, including the antenna, while wingless is restricted to the distal tip of the limbs and the neurectoderm basal to the limbs. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.