Number 53                                                                                                                                       May 2003

In February we flew to Washington DC so that Gretchen could identify ascidians at the Smithsonian’s Edgewater MD site only to be marooned in a hotel by snow for several days. We finally got there and the ascidians got identified although it meant doing some (the remaining 1071!) at home in Seattle.  In March we drove to southern California where we presented a paper on the southern hemisphere Corella eumyota that we found in France last summer (see AN#52 Work in Progress).  On the way back we visited Bill Smith at UC Santa Barbara to see his very impressive facilities for rearing mutant Cionas.  Soon after returning from the California trip we flew to Prince Edward Island, Canada, to participate in a workshop relating to the overgrowth of mussel lines by millions of invasive Styela clava.  Since this part of Canada is mostly soft bottom, the only firm substrates are pilings, docks, and the mussels and apparatus used in their culture.  Since S. clava is cultured for food in Korea, we hope that perhaps North American markets can be developed for S. clava.  Ciona intestinalis is also a big problem particularly on oyster cages.  Kevin Heasman from New Zealand gave a talk on his extensive experience with invasive ascidians and mussel culture in South Africa and New Zealand.  Sea ice 2 feet thick was a new experience for us.  The S. clava survive just fine all winter below the ice.  In May we presented a hands-on display of the Sea Squirts of the San Juans for the open house at the Friday Harbor Laboratories which was the formal kick-off of next year’s centennial celebration.  We also gave a short talk on the history of the laboratories.  Also in May we joined the Smithsonian invasions group to survey Tampa Bay, Florida, for invasive ascidians, and gave an ascidian identification workshop at Bodega marine lab in northern California.  Mid June to late July we will work at the Friday Harbor Labs, August we’ll participate in a nonindigenous species survey along the NE U.S. coast from Maine to New York, and in September we join the Smithsonian group again for a week in the Ketchikan area.
   There are 116 New Publications citations at the end of this issue—definitely there is something of interest to everyone working on ascidians. If you have not yet sent us your new publications, we would greatly appreciate receiving a hard copy if available, or pdf.

*Ascidian News is not part of the scientific literature and should not be cited as such.

                                                                  NEWS AND VIEWS

1.  Patrick Lemaire (lemaire@lgpd.univ-mrs.fr), Marseille, France is organizing an International Urochordate Meeting for Oct 11-14, 2003, at Carry-le Rouet, France, near Marseille.  This meeting will focus on evolutionary, developmental and functional genomics aspects of work carried out in our favorite animals.  An advisory committee (Mike Levine, Hiroki Nishida, Yasushi Okamura, Christian Sardet, Nori Satoh, and Billie Swalla) will select speakers on the basis of the submittted abstracts. For registration and abstract forms see: http://www.lgpd.univ-mrs.fr/more_info/lemaire/Front_Page.htm
There you can find the preliminary programme, registration form, abstract submission form and the meeeting venue website.  The deadline for the abstract submission is May 31, and the abstracts can only be accepted after your registration (see information on registration form and abstract submission form).  For more information write to: urochor@ibdm.univ-mrs.fr
The following sessions are planned:
         Oct 12 Session 1: Urochordate evolution
                    Session 2: Early embryonic patterning
         Oct 13 Session 3: Cell biological approaches
                     Session 4: Neural development
         Oct 14 Session 5: Of genomes and large scale approaches
                     Session 6: Round tables on community-oriented tools

2.  From Anna Di Gregorio  (and2015@med.cornell.edu):  After completing her postdoctoral work in the laboratory of Michael Levine at UC Berkeley, Anna Di Gregorio has recently moved to the East Coast and has set up her ascidian lab in the Department of Cell and Developmental Biology, at the Weill Medical College of Cornell University in Manhattan (NY). The main focus of the new lab is to study the evolutionary origins of notochord and other chordate tissues using as a main model system Ciona intestinalis, and in the future, other marine invertebrates, representative of sister phyla.
Collaborations, visits and suggestions are welcome!
http://www.med.cornell.edu/research/htmls/anna_di_gregorio.html

3.  The new ascidian website developed by Arjan Gittenberger (gittenberger@yahoo.com), National Museum of Natural History, Naturalis, Leiden The Netherlands, is expanding rapidly, with many more species added in the past 6 months.  Each entry includes a color photo, location and some information. At present there are a total of 235 photographs of 146 ascidian species/morphs.  Please visit the site at  www.ascidians.com.  Arjan writes: If anyone has any suggestions (e.g. for better identifications of the species), please let me know.”  Contributions are welcomed.

4.  From David Keys (dnkeys@lbl.gov) and Dario Boffelli (dboffelli@lbl.gov) :
Ciona intestinalis samples needed for polymorphism study.
   We are writing to asking your help putting together a global collection of Ciona intestinalis genomic DNA.  As most of you know, the polymorphism rate in Ciona has turned out to be quite high. (OK, amazingly high.) The two haplotypes from the single California Ciona intestinalis used to produce the genomic sequence here at the Joint Genome Institute had a 1.2% polymorphism rate. We (Dario Boffelli at the Lawrence Berkeley National Labs and David Keys at the JGI) are setting up a small project at LBNL to characterize just how much polymorphism is out there.  We’ve got several goals. The one we suspect will be most important to the general community will be a simple assessment of how close the sequence hosted at the JGI (http://www.jgi.doe.gov/ciona/index.html) is to the sequences of animals pulled from various locations around the world. In addition we are hypothesizing that it may be possible to locate functional regions, such as coding or perhaps even cis-reg domains, by looking for regions of low polymorphism.  This is similar to an approach that Dario has been doing using human, ape and monkey sequences (Boffelli et al. (2003) Phylogenetic shadowing of primate sequences to find functional regions of the human genome. Science. 299:1391-4). Currently we are not sure whether there will be enough polymorphism for this. However, any study to assess this can easily be set up to produce data for the general polymorphism assessment.
   Our initial plan is to generate sequence for a few genomic intervals containing enhancer, promoter and exon regions from multiple Ciona intestinalis individuals from as many world wide locations as possible. We would like your help in gathering the tissue samples for this.  If possible, we would like to do the DNA preps here to keep the protocol standardized. For any scientist who sends material, we will make sequence data from those animals available to them immediately so they can assess how their experimental population relates to the JGI reference genome.
   If you can supply material for this study, please contact us for collecting and shipping info (obviously, we'll pay for shipping).  We’d also appreciate any contacts you could send our way for commercial or marine lab collectors who might not otherwise see this message.

5.  The element vanadium was named “in honor of Vanadis, the Norse goddess of beauty” (quoted from Uncle Tungsten, Memories of a Chemical Boyhood by Oliver Sacks 2001).  Vanadium salts occur in many beautiful colors, depending on what it is combined with and which valency is present.  Even a very tiny amount, just a few atoms, of this transition element, can be enough to produce brilliant colors.  “Because it had four strikingly different valencies or oxidation states, and it was easy to transform these into one another, vanadium was an ideal element to experiment with.”  Sacks as a boy would start with a test tube of pentavalent ammonium vanadate in solution, add zinc amalgam, and the solution would change from yellow to royal blue, the color of tetravalent vanadium.  If allowed to react further until the vanadium changed to trivalent, the solution turned green.  “If one waited still longer, the green would disappear and be replaced by a beautiful lilac, the color of divalent vanadium. The reverse experiment was even more beautiful, especially if one layered potassium permanganate, a deep purple layer, over the delicate lilac; this would be oxidized over a period of hours and form separate layers, one above the other, of lilac divalent vanadium on the bottom, then green trivalent, then blue tetravalent, then yellow pentavalent vanadium.”

                                                                 WORK IN PROGRESS

1.  Elisabetta Tosti, Cell Biology lab, Stazione Zoologica, Villa Comunale, 80121 Napoli, Italy  tosti@szn.it : Bioactive aldehydes from diatoms block the fertilization current in ascidian oocytes by E. Tosti, G. Romano, I. Buttino, A. Cuomo, A. Ianora, A. Miralto, in press, Molecular Reproduction and Development.
   Abstract: We studied the effects of bioactive aldehydes from diatoms, unicellular algae at the base of the marine food web, on fertilization and early developmental processes of the ascidian Ciona intestinalis.  Using electrophysiological techniques, we showed  that the fertilization current which is generated in oocytes upon interaction with the spermatozoon was inhibited when oocytes were pre-incubated in 2-trans-4-trans-decadienal  (DD). This event was accompanied by the inhibition of the voltage-gated calcium current activity of the plasma membrane, whereas DD did not inhibit the subsequent contraction of the cortex.   DD did not act as a channel inhibitor since it did not affect the steady state conductance of the plasma membrane or gap junctional communication within blastomeres of the embryo. On the other hand, DD did affect actin reorganisation differently from that of other actin blockers.  Possibly this effect on actin reorganisation was responsible for the subsequent teratogenic action on larval development. A reversibility of DD effect indicates that it  may specifically target certain fertilization mechanisms.  Diatom aldehydes have been shown to interfere with reproductive mechanisms in copepods, polychaetes and echinoderms.  Our data provide evidence that such compounds may also adversely affect ascidian fertilization mechanisms.
  In this paper we show that diatom reactive aldehydes such as DD may have a dual effect on reproductive processes, influencing primary fertilization events such as gating of fertilization channels and secondary processes such as actin reorganisation which is responsible for the segregation of cell lines. These findings add to a growing body of evidence on the antiproliferative effects of diatom-derived aldehydes. Our results also report, for the first time, on the action of a fertilization channel blocker in marine invertebrates.

2.  George Mackie and C.L. Singla, Univ. of Victoria, Victoria, BC, Canada. mackie@uvic.ca:
The cupular strand of Corella inflata (Ascidiacea): a sense organ with GnRH-immunoreactive primary sensory neurons. In preparation.
   The cupular strand of Corella resembles the cupular organs of Ciona but unlike the latter, which are isolated, dome-shaped  structures located in the inner mantle epithelium, each containing 15-20 sensory cells,  the cupular strand is a single long structure containing some 1500 sensory cells and located not in the mantle but in the branchial sac, where it runs along the dorsal midline of the dorsal fold.  Preparations have been examined by optical and electron microscopy, and also by immunofluorescence microscopy using anti-tubulin and anti-gonadotropin-releasing hormone (GnRH) antisera. The sensory cells and their axons were immunoreactive with anti-tunicate I GnRH antiserum, but not with antisera against catfish and chicken GnRH.  All three GnRH antisera however labelled the dorsal strand plexus, a nerve net that runs in the dorsal blood sinus with extensions around the brain and in the vicinity of the gonads and gonoducts. It is suggested that the sensory neurons contain a different form of GnRH from the one(s) contained in the dorsal strand plexus and that it may function as a neuromodulator affecting ciliary beating in the branchial sac. The dorsal strand plexus on the other hand is probably the source of a form (or forms) of GnRH that regulate reproductive processes. It was possible to follow the sensory axons from the macula of the cupular strand to points where they disappeared into branches of the visceral nerve, which enters a nerve root at  the back of the brain.
   The function of the cupular strand has not been investigated physiologically but it seems likely that it is a hydrodynamic sensor registering changes in water flow velocity through the atrial cavity.  In discussing these findings we compare the cupular strand of Corella with putative hydrodynamic sensors described in other ascidians, and we consider the distribution and possible functions of GnRH cells in Corella in the context of existing information on the locations of such cells and their significance in ascidians generally.  [This paper continues similar research on a related species reported in a recently published article: Mackie GO & Singla CL 2003. The capsular organ of Chelyosoma productum (Ascidiacea: Corellidae):a new tunicate hydrodynamic sense organ. Brain Behav. Evol. 61:45-58.]

3.  From Patrick Frank, Dept. of Chemistry, Stanford Univ., Stanford, CA 94305-5080 (Frank@ssrl.slac.stanford.edu):  We recently completed a study on vanadium in blood cells from Ascidia ceratodes as found in Bodega Bay, California, and included a comparison with analogous results for A. ceratodes in Monterey Bay, CA (P. Frank, R.M.K. Carlson, E.J. Carlson and K.O. Hodgson 2003 "The vanadium environment in blood cells of Ascidia ceratodes is divergent at all organismal levels: an XAS and EPR spectroscopic study" J. Inorg. Biochem. 94: 59-71). Essentially, both the blood cell vanadium content and the intracellular acidity vary virtually from cell-to-cell within a given animal, from animal-to-animal, and, on average, between the populations of Monterey Bay and Bodega Bay, CA.  The signet-ring blood cell intravacuolar acidity in the Bodega A. ceratodes is as low as pH 0, and on average is noticably more acidic than the Monterey population. This pH difference produces observable differences in the distributions of dissolved vanadium complexes. We've also noted that Bodega individuals average much larger than the Monterey. Frozen blood cells also vary from uniformly canary-yellow (Bodega) to yellow-green (Monterey). One wonders whether there may be an incipient speciation event in progress there.

4.  Ryo Koyanagi (rkoyanag@zool.unizh.ch) and Thomas G. Honegger (thho@zool.unizh.ch) , Zool. Inst., Univ. of Zürich, CH-8057 Zürich, Switzerland. Molecular cloning and sequence analysis of an ascidian egg b-N-acetylhexosaminidase with potential role in fertilization.  In press, Develop. Growth Differ.
   Beta-N-acetylhexosaminidase, which is found almost ubiquitously in sperm of invertebrates and vertebrates, supposedly mediates a carbohydrate-based transient sperm-egg coat binding. In ascidians and mammals beta-hexosaminidase released at fertilization from eggs has been proposed to modify sperm-receptor glycoproteins of the egg envelope, thus setting up a block to polyspermy. Previously, we showed that in potential sperm-receptor glycoproteins of the ascidian Phallusia mammillata N-acetylglucosamine is the prevailing glycoside residue and that the egg harbors three active molecular forms of b-hexosaminidase. In the present study, we have isolated and characterized P. mammillata beta-hexosaminidase cDNA from an ovarian cDNA library. The deduced amino acid sequence showed high similarities with other known b-hexosaminidases, on the other hand P. mammillata beta-hexosaminidase has an unique potential N-glycosylation site. A phylogenetic analysis suggests that the P. mammillata beta-hexosaminidase developed independently after having branched off from the common ancestor gene of the chordate enzyme before two isoforms of the mammalian enzyme appeared. In situ hybridization revealed stage-specific expression of b-hexosaminidase mRNA during oogenesis in the oocyte and the accessory test and follicle cells. This suggests that the three egg beta-hexosaminidase forms are specific for the oocyte, the test and follicle cells respectively.

5.  Gretchen Lambert: New records of ascidians from the NE Pacific: a new species of Trididemnum, range extension and redescription of Aplidiopsis pannosum (Ritter, 1899) including its larva, and several nonindigenous species.  In press, Zoosystema (December 2003).
  Abstract:  A new species of aplousobranch ascidian, Trididemnum alexi, is described from the San Juan Archipelago of Washington state, USA. Colonies are smooth dark reddish brown; the largest is 8 cm in maximum width and up to 2 cm thick in some regions because of the large and complex hypozooidal cloacal canals.  A thick (200-400 µm) superficial bladder cell layer is present.  The spicules are 20-30 µm in diameter, irregularly stellate with short pointed rays (8-10 in optical equatorial plane) and scattered thinly through the tunic but absent from the bladder cell layer.  Zooids have 3 rows of stigmata, with 12-13 stigmata per side in the first two rows in most zooids (occasionally 11 or 14) and usually 1-2 fewer in the third row.  The tubular atrial siphon opens dorsal to the middle row of stigmata. The single testis is covered by a sperm duct with 8-9 coils. Larvae form in the basal portion of the colony, are 0.8-1 mm in trunk length with 3 adhesive papillae and usually 7 pairs of lateral ampullae with curved tips in the fully formed tadpoles.  A redescription of Aplidiopsis pannosum (Ritter, 1899) includes the morphological analysis of several larval stages.  This species, though widespread in the north Pacific, was not previously known to occur south of Alaska on the west coast of North America. Range extensions are also included for several nonindigenous ascidians: Ciona savignyi, Botrylloides violaceus, Styela clava and Molgula manhattensis.

6.  Paolo Burighel, L. Manni, G. Zaniolo, N.J. Lane and colleagues (burighel@civ.bio.unipd.it), Univ. of Padua, Italy.
   We have demonstrated the presence of a new, possibly mechanoreceptor organ, "the coronal organ" in the oral siphons of botryllids (Botryllus schlosseri and Botrylloides leachi) (in press in J. Comparative Neurology vol. 461).  This organ is composed of a line of sensory cells, accompanied by supporting cells that runs along the margin of the velum and the tentacles of the siphon.  The sensory cells are "hair cells" resembling those of the vertebrate lateral line, or, in general, the acoustico-lateralis system, because they bear a single cilium, located centrally or eccentrically to a hair bundle of numerous stereovilli.  In contrast to other sensory cells of ascidians, the coronal hair cells are secondary sensory cells, since they lack axonal processes directed towards the cerebral ganglion.  Moreover, at their base, they form synapses with nerve fibres, most of which exhibit acetylcholinesterase activity.  The absence of axonal extensions was confirmed by experiments with lipophilic dyes.  Different kinds of synapses were recognised: usually, each hair cell forms a few afferent synapses with dendrites of neurons located in the ganglion; efferent synapses, both axo-somatic (between an axon coming from the ganglion and the hair cell) and axo-dendritic (between an axon coming from the ganglion and an afferent fibre) were occasionally found.  In the paper we compare the secondary sensory cells of ascidians with the sensory cells and placodes of vertebrates and propose that the coronal organ is homologous to the vertebrate acoustico-lateralis system.

7.  Carlo Brena (carlobrena72@hotmail.com), Univ. of Padua, Italy.
   During my PhD thesis I studied with light and electron microscopy (applying to it also histochemistry and immunohistochemistry) the alimentary tract of several species of appendicularian belonging to the three families of these tunicates. The results (The alimentary system of Appendicularians, simplifications, specializations, adaptation [in Italian]), show that the three families, especially from the point of view of the gut, are strikingly different from one another, with a strong separation, in particular, between Oikopleurids on one side and Fritillarids and Kowalevskids on the other side. From both the general morphology and the cytological differentiation, there are many more differences between these two groups than among all the other tunicates. In particular, they show ultrastructural specializations, so far apparently unique, e.g. some apical junctions between cells in Kowalevskia. The three families, in particular fritillarids and kowalevskids, show the body with strong simplification which might account for their extremely short life cycles and high biomass and dispersal in all oceans. In particular, fritillarids have some gut cells extremely specialized, probably involved in osmotic regulation. The results should arouse interest on these so far poorly known animals, from the ultrastructural, ecological/physiological and evolutionary points of view. As for this last point, a picture and some references on the exceptionality of these tunicates has been introduced in the newly released book The Development of Animal Form: Ontogeny Morphology and Evolution, by Alessandro Minelli, Cambridge Press.
   Most of my thesis material has been or will soon be published; in addition to the articles already published on Oikopleura dioica digestive system (TEM: Burighel et al. 2001) and histochemistry (Cima et al. 2002) [see past issues of AN for complete citations], three articles are in press:
Brena, C., F. Cima & P. Burighel 2003. The highly specialised gut of Fritillariidae (Appendicularia, Tunicata).  Marine Biology, in press.
Brena, C., F. Cima & P. Burighel 2003. The exceptional “blind” gut of Appendicularia sicula (Appendicularia, Tunicata).  Zoologischer Anzeiger in press.
Brena, C., F. Cima & P. Burighel 2003.  The alimentary tract of Kowalevskiidae (Appendicularia, Tunicata) and evolutionary implications. Journal of Morphology in press.

                                                            THESIS ABSTRACTS

1.  Aplousobranch ascidians (Tunicata: Ascidiacea) from southern Africa.  Shirley Parker-Nance, Dept. of Zool., Univ. of Port Elizabeth, S. Africa.  Ph.D. thesis, awarded April 2003. Advisor: Prof. T. Wooldridge Univ. of Port Elizabeth and Prof. M. Davies-Coleman, Rhodes University.  shirley@justblue.co.za, http://www.JustBlue.co.za
   The ascidian (subphylum Tunicata: class Ascidiacea) fauna along the southern African coast constitutes an important component of the sessile benthic reef fauna. Little is known of the species composition, biodiversity and distribution of ascidians on southern African intertidal and subtidal reefs. Past research on the ascidian fauna of South Africa was sporadic and limited, with only about one hundred and seventy species recorded during the last one hundred and twenty years. This is the first taxonomic study undertaken by a South African resident scientist. The study focused on six genera in four families (Euherdmaniidae, Polyclinidae, Pseudodistomidae and Didemnidae) belonging to the class Ascidiacea and suborder Aplousobranchia. Five species new to science, two Polyclinum, two Pseudodistoma and one Polysyncraton species are described. New additional information on the distribution of five species previously known to science, including one recorded for the first time along the South African coast, is presented. A literature review and comparison of the taxonomic important characteristics is made of all species known globally for the six genera. These six genera comprise 12 Euherdmania, 15 Aplidiopsis, 40 Polyclinum, 31 Pseudodistoma, 6 Atriolum and 68 Polysyncraton species.

2.  Phylogeny of the Order Aplousobranchia (Tunicata, Ascidiacea). Tatiane Regina Moreno, Dept. of Zoology, Universidade Federal do Paraná, Brazil. (tatiane@bio.ufpr.br)  Ph.D. Thesis, March 2003. Advisors Dr. Rosana Moreira da Rocha & Dr. Walter A. P. Böeger.
   The phylogeny of Aplousobranchia Lahille (Tunicata, Ascidiacea) sensu Kott (1969, 1990, 1992) was for the first time reconstructed using morphological characters. The relationships between genera and families were analyzed employing the principles of phylogenetic systematics (Hennig 1966; Wiley 1981) with PAUP. Monophyly of the Aplousobranchia and of the families from this order were tested with ascidians from 14 different families.  A matrix with 40 taxa and 59 characters was analyzed using a heuristic search strategy: random stepwise addition (n=1000 replicates). Pyura Molina, 1782 and Molgula Forbes & Hanley, 1848 (Stolidobranchia) and Ascidia Linnaeus, 1767 and Perophora Wiegmann, 1835 (Phlebobranchia) were defined as outgroups. The analysis resulted in 154 equally most parsimonious trees (length 173, CI=0.5549, RI=0.8046). Characters correlated with replication process and the formation of colony systems, and branchial wall characters were more important in the phylogenetic reconstruction of Aplousobranchia than other characters traditionally used in Ascidiacea taxonomy, such as body division and position of the heart, gonads and epicardium. New characters included here for the first time such as body wall muscles, muscles associated with transversal blood vessels and arrangement of the larval papillae also have phylogenetic importance. The monophyly of Aplousobranchia sensu Lahille, including only Polycitoridae, Polyclinidae, and Didemnidae ascidians is supported. However, Aplousobranchia sensu Kott (1969, 1990, 1992) is not monophyletic because Perophora from outgroup was included in the cladogram in the ingroup. Several Aplousobranchia families are monophyletic. We proposed a classification based on the phylogenetic analysis, in which Aplousobranchia now has 17 families (two new), and some indeterminant taxa.

                                                                  MEETINGS ABSTRACTS

Molecular Biology Society of Japan 25th annual meeting, December 11-14, 2002. Yokohama, Japan.
Michibata, H., Ueki, T. and Yamaguchi, N. Molecular physiological analysis of the mechanisms of vanadium accumulation in ascidians.  [no abstract].
 

                                                                  NEW PUBLICATIONS

Adams, B. A., Tello, J. A., Erchegyi, J., Warby, C., Hong, D. J., Akinsanya, K. O., Mackie, G. O., Vale, W., Rivier, J. E. and Sherwood, N. M. 2003. Six novel gonadotropin-releasing hormones are encoded as triplets on each of two genes in the protochordate, Ciona intestinalis. Endocrinology 144: 1907-1919.

Aiello, A., Fattorusso, E., Luciano, P., Menna, M., Esposito, G., Iuvone, T. and Pala, D. 2003. Conicaquinones A and B, two novel cytotoxic terpene quinones from the Mediterranean ascidian Aplidium conicum. Europ. J. Org. Chem. March 2003: 898-900.

Aiello, A., Fattorusso, E., Mangoni, A. and Menna, M. 2003. Three new 2,3-dihydroxy fatty acid glycosphingolipids from the Mediterranean tunicate Microcosmus sulcatus. Europ. J. Org. Chem. Feb. 2003: 734-739.

Aizenberg, J., Weiner, S. and Addadi, L. 2003. Coexistence of amorphous and crystalline calcium carbonate in skeletal tissues. Conn. Tiss. Res. 44 Suppl. 1: 20-22.

Artman, G. D. and Weinreb, S. M. 2003. An approach to the total synthesis of the marine ascidian metabolite perophoramidine via a halogen-selective tandem Heck/carbonylation strategy. Org. Lett. 5: 1523-1526.

Azumi, K., Kuribayashi, F., Kanegasaki, S. and Yokosawa, H. 2002. Zymosan induces production of superoxide anions by hemocytes of the solitary ascidian Halocynthia roretzi. Comp. Biochem. Physiol. C 133: 567-574.

Boorman, C. J. and Shimeld, S. M. 2002. The evolution of left-right asymmetry in chordates. BioEssays 24: 1004-1011.

Burton, A. 2002. Sea squirt assaults sarcomas. Lancet Oncol. 3: 648.

Canestro, C., Bassham, S. and Postlethwait, J. H. 2003. Seeing chordate evolution through the Ciona genome sequence. Genome Biol. 4: 208.

Canestro, C., Godoy, L., Gonzalez-Duarte, R. and Albalat, R. 2003. Comparative expression analysis of Adh3 during arthropod, urochordate, cephalochordate, and vertebrate development challenges its predicted housekeeping role. Evol. & Dev. 5: 157-62.

Catlow, K., Deakin, J. A., Delehedde, M., Fernig, D. G., Gallagher, J. T., Pavao, M. S. G. and Lyon, M. 2003. Hepatocyte growth factor/scatter factor and its interaction with heparan sulphate and dermatan sulphate. Biochem. Soc. Trans. 31: 352-353.

Chiba, S., Awazu, S., Itoh, M., Chin-Bow, S. T., Satoh, N., Satou, Y. and Hastings, K. E. 2003. A genomewide survey of developmentally relevant genes in Ciona intestinalis IX. Genes for muscle structural proteins. Dev Genes Evol 10: 10.

Coles, S. L. and Eldredge, G. S. 2002. Nonindigenous species introductions on coral reefs: a need for information. Pac. Sci. 56: 191-209.

Cooper, E. L. and Parrinello, N. 2001. Immunodefense in tunicates: cells and molecules. In: Sawada, H., Yokosawa, H. and Lambert, C. C. (ed.), The Biology of Ascidians. Tokyo, Springer-Verlag, pp. 383-394.

Cruz-Monserrate, Z., Vervoort, H. C., Bai, R., Newman, D. J., Howell, S. B., Los, G., Mullaney, J. T., Williams, M. D., Pettit, G. R., Fenical, W. and Hamel, E. 2003. Diazonamide a and a synthetic structural analog: disruptive effects on mitosis and cellular microtubules and analysis of their interactions with tubulin. Mol. Pharmacol. 63: 1273-1280.

Dalfo, D., Albalat, R., Molotkov, A., Duester, G. and Gonzales-Duarte, R. 2002. Retinoic acid synthesis in the prevertebrate amphioxus involves retinol oxidation. Dev. Genes Evol. 212: 388-393.

D'Ambrosio, P., Fanelli, A., Pischetola, M. and Spagnuolo, A. 2003. Ci-GATAa, a GATA-class gene from the ascidian Ciona intestinalis: Isolation and developmental expression. Dev. Dyn. 226: 145-148.

Davis, R. A., Sandoval, I. T., Concepcion, G. P., Rocha, R. M. and Ireland, C. A. 2003. Lissoclinotoxins E and F, novel cytotoxic alkaloids from a Philippine didemnid ascidian. Tetrahedron 59: 2855-2859.

De Tomaso, A. W. and Weissman, I. L. 2003. Construction and characterization of large-insert genomic libraries (BAC and fosmid) from the ascidian Botryllus schlosseri and initial physical mapping of a histocompatibility locus. Mar. Biotechnol. 5: 103-115.

Dehal, P., Satou, Y., Campbell, R. K., Chapman, J., Degnan, B., De Tomaso, A., Davidson, B., Di Gregorio, A. and al., e. 2002. The draft genome of Ciona intestinalis: insights into chordate and vertebrate origins. Science 298: 2157-2167.

Delfourne, E. and Bastide, J. 2003. Marine pyridoacridine alkaloids and synthetic analogues as antitumor agents. Med. Res. Rev. 23: 234-252.

Deschet, K., Nakatani, Y. and Smith, W. C. 2003. Generation of Ci-Brachyury-GFP stable transgenic lines in the ascidian Ciona savignyi. Genesis 35: 248-259.

Dumollard, R., Hammar, K., Porterfield, M., Smith, P. J., Cibert, C., Rouviere, C. and Sardet, C. 2003. Mitochondrial respiration and Ca2+ waves are linked during fertilization and meiosis completion. Development 130: 683-692.

Elphick, M. R., Satou, Y. and Satoh, N. 2003. The invertebrate ancestry of endocannabinoid signalling: an orthologue of vertebrate cannabinoid receptors in the urochordate Ciona intestinalis. Gene 302: 95-101.

Endo, Y., Nonaka, M., Saiga, H., Kakinuma, Y., Matsushita, A., Takahashi, M., Matsushita, M. and Fujita, T. 2003. Origin of mannose-binding lectin-associated serine protease (MASP)-1 and MASP-3 involved in the lectin complement pathway traced back to the invertebrate, amphioxus. J. Immunol. 170: 4701-4707.

Frank, P., Carlson, R. M. K., Carlson, E. J. and Hodgson, K. O. 2003. The vanadium environment in blood cells of Ascidia ceratodes is divergent at all organismal levels: an XAS and EPR spectroscopic study. J. Inorg. Chem. 94: 59-71.

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