Hydrothermal Vent Tubeworm Riftia Research Articles

Co-expression analysis reveals distinct alliances around two carbon fixation pathways in hydrothermal vent symbionts

Most autotrophic organisms possess a single carbon fixation pathway. The chemoautotrophic symbionts of the hydrothermal vent tubeworm Riftia pachyptila, however, possess two functional pathways: the Calvin–Benson–Bassham (CBB) and the reductive tricarboxylic acid (rTCA) cycles. How these two pathways are coordinated is unknown. Here we measured net carbon fixation rates, transcriptional/metabolic responses and transcriptional co-expression patterns of Riftia pachyptila endosymbionts by incubating tubeworms collected from the East Pacific Rise at environmental pressures, temperature and geochemistry. Results showed that rTCA and CBB transcriptional patterns varied in response to different geochemical regimes and that each pathway is allied to specific metabolic processes; the rTCA is allied to hydrogenases and dissimilatory nitrate reduction, whereas the CBB is allied to sulfide oxidation and assimilatory nitrate reduction, suggesting distinctive yet complementary roles in metabolic function. Furthermore, our network analysis implicates the rTCA and a group 1e hydrogenase as key players in the physiological response to limitation of sulfide and oxygen. Net carbon fixation rates were also exemplary, and accordingly, we propose that co-activity of CBB and rTCA may be an adaptation for maintaining high carbon fixation rates, conferring a fitness advantage in dynamic vent environments.

Bacterial symbiont subpopulations have different roles in a deep-sea symbiosis.

The hydrothermal vent tubeworm Riftia pachyptila hosts a single 16S rRNA phylotype of intracellular sulfur-oxidizing symbionts, which vary considerably in cell morphology and exhibit a remarkable degree of physiological diversity and redundancy, even in the same host. To elucidate whether multiple metabolic routes are employed in the same cells or rather in distinct symbiont subpopulations, we enriched symbionts according to cell size by density gradient centrifugation. Metaproteomic analysis, microscopy, and flow cytometry strongly suggest that Riftia symbiont cells of different sizes represent metabolically dissimilar stages of a physiological differentiation process: While small symbionts actively divide and may establish cellular symbiont-host interaction, large symbionts apparently do not divide, but still replicate DNA, leading to DNA endoreduplication. Moreover, in large symbionts, carbon fixation and biomass production seem to be metabolic priorities. We propose that this division of labor between smaller and larger symbionts benefits the productivity of the symbiosis as a whole.

Polyclonal symbiont populations in hydrothermal vent tubeworms and the environment

Horizontally transmitted symbioses usually house multiple and variable symbiont genotypes that are acquired from a much more diverse environmental pool via partner choice mechanisms. However, in the deep-sea hydrothermal vent tubeworm Riftia pachyptila (Vestimentifera, Siboglinidae), it has been suggested that the Candidatus Endoriftia persephone symbiont is monoclonal. Here, we show with high-coverage metagenomics that adult R. pachyptila house a polyclonal symbiont population consisting of one dominant and several low-frequency variants. This dominance of one genotype is confirmed by multilocus gene sequencing of amplified housekeeping genes in a broad range of host individuals where three out of four loci ( atpA, uvrD and recA) revealed no genomic differences, while one locus ( gyrB) was more diverse in adults than in juveniles. We also analysed a metagenome of free-living Endoriftia and found that the free-living population showed greater sequence variability than the host-associated population. Most juveniles and adults shared a specific dominant genotype, while other genotypes can dominate in few individuals. We suggest that although generally permissive, partner choice is selective enough to restrict uptake of some genotypes present in the environment.

Comparison of proton-specific ATPase activities in plume and root tissues of two co-occurring hydrocarbon seep tubeworm species Lamellibrachia luymesi and Seepiophila jonesi

Lamellibrachia luymesi and Seepiophila jonesi are co-occurring species of vestimentiferan tubeworms found at hydrocarbon seepage sites on the upper Louisiana slope of the Gulf of Mexico. Like all vestimentiferans, they rely on internal sulfide-oxidizing symbiotic bacteria for nutrition. These symbionts produce hydrogen ions as a byproduct of sulfide oxidation, which the host tubeworm needs to eliminate to prevent acidosis. The hydrothermal vent tubeworm Riftia pachyptila uses a high activity of P- and V-type H+-ATPases located in its plume epithelium to excrete protons. Unlike R. pachyptila, the seep species grow a posterior root, which they can use in addition to their plumes as a nutrient exchange surface. In this study we measured the ATPase activities of plume and root tissues collected from L. luymesi and S. jonesi, and used a combination of inhibitors to determine the relative activities of P- and V-type H+-ATPases. We found that the total H+-ATPase activity of their plumes was approximately 14 μmol h−1 g−1 wet weight, and that of their roots was between 5 and 7 μmol h−1 g−1 wet weight. These activities were more than ten times lower than those measured in R. pachyptila. We suggest that seep tubeworms might use passive channels to eliminate protons across their roots, in addition to ATP-dependant proton pumps located in their plumes and roots. In addition, we found strong differences between the types of ATPase activities in the plumes of L. luymesi and S. jonesi. While the H+-ATPase activity of L. luymesi plumes is dominated by P-type ATPases, S. jonesi has an unusually high activity of V-type H+-ATPases. We suggest that S. jonesi relies on its high V-type H+-ATPase activity to drive carbon dioxide uptake across its plume surface. L. luymesi, on the other hand, might rely partially on bicarbonate uptake across its root.

Toward a mechanistic understanding of larval dispersal: insights from genomic fingerprinting of the deep‐sea hydrothermal vent tubeworm Riftia pachyptila

AbstractLarval dispersal is critical for the maintenance of species populations in patchy and ephemeral hydrothermal vent habitats. On fast‐spreading ridges, such as the East Pacific Rise, rates of habitat turnover are comparable to estimated lifespans of many of the inhabiting species. Traditionally, dispersal questions have been addressed with two very different approaches, larval studies and population genetics. Population genetic studies of vent‐endemic species have been informative for determining whether patterns of dispersal are suggestive of stepping stone or island models and estimating rates of gene flow (effective migrants per generation) over broad geographic ranges. However, these studies leave fundamental questions unanswered about the specific mechanisms by which larvae disperse and species maintain their populations and biogeographic ranges. With the goal of examining genetic structure and elucidating alternative larval dispersal mechanisms, we employed a genomic DNA fingerprinting technique, amplified fragment length polymorphisms (AFLPs). To assess the potential utility of AFLPs, and genetic structure of the hydrothermal vent tubeworm Riftia pachyptila, genomic ‘fingerprints’ were recovered from 29 individuals from five vent fields spanning a distance of up to c. 5000 km along the East Pacific Rise. In contrast to previous population genetic studies that found little to no genetic structure using allozymes and mitochondrial DNA, genetic analyses of 630 polymorphic AFLP loci identified distinct subclades within R. pachyptila populations. Significant levels of differentiation were observed among populations from all vent regions as well as within each region. Discrete assemblages of tubeworms separated by as little as c. 400 m within a given vent region were genetically distinguishable and cohorts (based on size‐frequency distribution) within an aggregation were found to be most closely related. These results suggest that mechanisms of larval dispersal act to retain cohort fidelity in R. pachyptila.

Metabolite uptake, stoichiometry and chemoautotrophic function of the hydrothermal vent tubeworm Riftia pachyptila: responses to environmental variations in substrate concentrations and temperature

The hydrothermal vent tubeworm Riftia pachyptila is a dominant member of many hydrothermal vent communities along the East Pacific rise and is one of the fastest growing metazoans known. Riftia flourish in diffuse hydrothermal fluid flows, an environment with high spatial and temporal heterogeneity in physical and chemical conditions. To date, physiological and biochemical studies of Riftia have focused on Riftia's adaptations to its chemoautotrophic bacterial symbionts. However the relation between in situ physico-chemical heterogeneity and Riftia host and symbiont metabolism, in particular symbiont chemoautotrophic function, remain poorly understood. Accordingly, we conducted experiments using shipboard high-pressure respirometers to ascertain the effect of varying substrate concentrations and temperature on Riftia metabolite uptake and symbiont carbon fixation. Our results show that substrate concentrations can strongly govern Riftia oxygen and sulfide uptake rates, as well as net carbon uptake (which is a proxy for chemoautotrophic primary production). However, after sufficient exposure to sulfide and oxygen, Riftia were capable of sustaining symbiont autotrophic function for several hours in seawater devoid of sulfide or oxygen, enabling the association to support symbiont metabolism through brief periods of substrate deficiency. Overall, temperature had the largest influence on Riftia metabolite uptake and symbiont autotrophic metabolism. In sum, while Riftia requires sufficient availability of substrates to support symbiont chemoautotrophic function, it is extremely well poised to buffer the temporal and spatial heterogeneity in environmental substrate concentrations, alleviating the influence of environmental heterogeneity on symbiont chemoautotrophic function.

Sulfide binding is mediated by zinc ions discovered in the crystal structure of a hydrothermal vent tubeworm hemoglobin

Key to the remarkable ability of vestimentiferan tubeworms to thrive in the harsh conditions of hydrothermal vents are hemoglobins that permit the sequestration and delivery of hydrogen sulfide and oxygen to chemoautotrophic bacteria. Here, we demonstrate that zinc ions, not free cysteine residues, bind sulfide in vestimentiferan hemoglobins. The crystal structure of the C1 hemoglobin from the hydrothermal vent tubeworm Riftia pachyptila has been determined to 3.15 A and revealed the unexpected presence of 12 tightly bound Zn(2+) ions near the threefold axes of this D(3) symmetric hollow sphere. Chelation experiments on R. pachyptila whole-coelomic fluid and purified hemoglobins reveal a role for Zn(2+) ions in sulfide binding. Free cysteine residues, previously proposed as sulfide-binding sites in vestimentiferan hemoglobins, are found buried in surprisingly hydrophobic pockets below the surface of the R. pachyptila C1 molecule, suggesting that access of these residues to environmental sulfide is restricted. Attempts to reduce the sulfide-binding capacities of R. pachyptila hemoglobins by addition of a thiol inhibitor were also unsuccessful. These findings challenge the currently accepted paradigm of annelid hemoglobin evolution and adaptation to reducing environments.

Experimental application of vascular and coelomic catheterization to identify vascular transport mechanisms for inorganic carbon in the vent tubeworm, Riftia pachyptila

Maintaining deep sea animals in in situ conditions has always been technically difficult because of the high-pressure requirements. Even more difficult are any attempts in manipulating or sampling these organisms while keeping them alive in high-pressure aquaria. We present a technique to withdraw blood samples by vascular catheterization which allows withdrawal of samples of during maintenance of specimens under high-pressure conditions. We have developed this technique to answer a long debated question, how carbon dioxide is transported from the ambient sea water to the bacterial symbionts inside the trophosome of the hydrothermal vent tubeworm Riftia pachyptila. Our results indicate that the carbon supply to the symbionts is mainly through inorganic CO 2 while its incorporation into malate and succinate may serve storage functions at periods of low CO 2 availability in the environment.

Characterization of carbonic anhydrases from Riftia pachyptila, a symbiotic invertebrate from deep-sea hydrothermal vents.

The symbiotic hydrothermal vent tubeworm Riftia pachyptila needs to supply its internal bacterial symbionts with carbon dioxide, their inorganic carbon source. Our aim in this study was to characterize the carbonic anhydrase (CA) involved in CO(2) transport and conversion at various steps in the plume and the symbiotic tissue, the trophosome. A complete 1209 kb cDNA has been sequenced from the trophosome and identified as a putative alpha-CA based on BLAST analysis and the similarities of total deduced amino-acid sequence with those from the GenBank database. In the plume, the putative CA sequence obtained from cDNA library screening was 90% identical to the trophosome CA, except in the first 77 nucleotides downstream from the initiation site identified on trophosome CA. A phylogenetic analysis showed that the annelidan Riftia CA (CARp) emerges clustered with invertebrate CAs, the arthropodan Drosophila CA and the cnidarian Anthopleura CA. This invertebrate cluster appeared as a sister group of the cluster comprising mitochondrial and cytosolic isoforms in vertebrates: CAV, CAI II and III, and CAVII. However, amino acid sequence alignment showed that Riftia CA was closer to cytosolic CA than to mitochondrial CA. Combined biochemical approaches revealed two cytosolic CAs with different molecular weights and pI's in the plume and the trophosome, and the occurrence of a membrane-bound CA isoform in addition to the cytosolic one in the trophosome. The physiologic roles of cytosolic CA in both tissues and supplementary membrane-bound CA isoform in the trophosome in the optimization of CO(2) transport and conversion are discussed.

Effects of metabolite uptake on proton-equivalent elimination by two species of deep-sea vestimentiferan tubeworm, Riftia pachyptila and Lamellibrachia cf luymesi: proton elimination is a necessary adaptation to sulfide-oxidizing chemoautotrophic symbionts.

Intracellular symbiosis requires that the host satisfy the symbiont's metabolic requirements, including the elimination of waste products. The hydrothermal vent tubeworm Riftia pachyptila and the hydrocarbon seep worm Lamellibrachia cf luymesi are symbiotic with chemolithoautotrophic bacteria that produce sulfate and protons as end-products. In this report, we examine the relationship between symbiont metabolism and host proton equivalent elimination in R. pachyptila and L. cf luymesi, and the effects of sulfide exposure on proton-equivalent elimination by Urechis caupo, an echiuran worm that lacks intracellular symbionts (for brevity, we will hereafter refer to proton-equivalent elimination as 'proton elimination'). Proton elimination by R. pachyptila and L. cf luymesi constitutes the worms' largest mass-specific metabolite flux, and R. pachyptila proton elimination is, to our knowledge, the most rapid reported for any metazoan. Proton elimination rates by R. pachyptila and L. cf luymesi correlated primarily with the rate of sulfide oxidation. Prolonged exposure to low environmental oxygen concentrations completely inhibited the majority of proton elimination by R. pachyptila, demonstrating that proton elimination does not result primarily from anaerobic metabolism. Large and rapid increases in environmental inorganic carbon concentrations led to short-lived proton elimination by R. pachyptila, as a result of the equilibration between internal and external inorganic carbon pools. U. caupo consistently exhibited proton elimination rates 5-20 times lower than those of L. cf luymesi and R. pachyptila upon exposure to sulfide. Treatment with specific ATPase inhibitors completely inhibited a fraction of proton elimination and sulfide and inorganic carbon uptake by R. pachyptila, suggesting that proton elimination occurs in large part via K(+)/H(+)-ATPases and Na(+)/H(+)-ATPases. In the light of these results, we suggest that protons are the primary waste product of the symbioses of R. pachyptila and L. cf luymesi, and that proton elimination is driven by symbiont metabolism, and may be the largest energetic cost incurred by the worms.

Evolution of the sulfide-binding function within the globin multigenic family of the deep-sea hydrothermal vent tubeworm Riftia pachyptila.

The giant extracellular hexagonal bilayer hemoglobin (HBL-Hb) of the deep-sea hydrothermal vent tube worm Riftia pachyptila is able to transport simultaneously O(2) and H(2)S in the blood from the gills to a specific organ: the trophosome that harbors sulfide-oxidizing endosymbionts. This vascular HBL-Hb is made of 144 globins from which four globin types (A1, A2, B1, and B2) coevolve. The H(2)S is bound at a specific location (not on the heme site) onto two of these globin types. In order to understand how such a function emerged and evolved in vestimentiferans and other related annelids, six partial cDNAs corresponding to the six globins known to compose the multigenic family of R. pachyptila have been identified and sequenced. These partial sequences (ca. 120 amino acids, i.e., 80% of the entire protein) were used to reconstruct molecular phylogenies in order to trace duplication events that have led to the family organization of these globins and to locate the position of the free cysteine residues known to bind H(2)S. From these sequences, only two free cysteine residues have been found to occur, at positions Cys + 1 (i.e., 1 a.a. from the well-conserved distal histidine) and Cys + 11 (i.e., 11 a.a. from the same histidine) in globins B2 and A2, respectively. These two positions are well conserved in annelids, vestimentiferans, and pogonophorans, which live in sulfidic environments. The structural comparison of the hydrophobic environment that surrounds these cysteine residues (the sulfide-binding domain) using hydrophobic cluster analysis plots, together with the cysteine positions in paralogous strains, suggests that the sulfide-binding function might have emerged before the annelid radiation in order to detoxify this toxic compound. Moreover, globin evolutionary rates are highly different between paralogous strains. This suggests that either the two globin subfamilies involved in the sulfide-binding function (A2 and B2) have evolved under strong directional selective constraints (negative selection) and that the two other globins (A1 and B1) have accumulated more substitutions through positive selection or have evolved neutrally after a relaxation of selection pressures. A likely scenario on the evolution of this multigenic family is proposed and discussed from this data set.

Biometry of the branchial plume in the hydrothermal vent tubewormRiftia pachyptila(Vestimentifera; Annelida)

The branchial plume of the hydrothermal vent tubeworm Riftia pachyptila is the main organ by which this mouth- and gut-less tubeworm directly exchanges metabolites with its environment. We estimated the total branchial surface area per unit wet mass, termed the specific branchial surface area (SBSA), from planimetric measurements. Changes in the SBSA during the growth of the worm were inferred from 16 individuals ranging from 1 to 112 g wet mass. Riftia pachyptila has a mean SBSA of 22 cm2·g–1, the second highest among all aquatic animals, representing 9 times the surface area of the rest of the body. Three significantly different classes of SBSA could be distinguished, corresponding to small, medium-sized, and large individuals. The SBSA values for small and medium-sized R. pachyptila are twice that for large individuals. Negative growth allometry between the length of the branchial plume and that of the trunk may be correlated with this variation in SBSA, the plume growing faster than the trunk in the small and medium-sized groups. In large individuals the trunk length exceeds the plume length, inducing an increase in body mass that lowers the SBSA. However, a lower SBSA does not imply reduced metabolite diffusion through the plume of large tubeworms, since their longer free filaments bear more developed pinnules, which are probably the preferred pathway of metabolite diffusion, owing to a minimal transepithelial distance of 2 µm.

Activity and inhibitor sensitivity of ATPases in the hydrothermal vent tubeworm Riftia pachyptila : a comparative approach

Phosphorylated ATPases may be involved in the effective pH regulation seen in the hydrothermal vent tubeworm Riftia pachyptila. R. pachyptila appears not only to have a large concentration of ATPases, but the main function of these ATPases seems to have shifted from other types of transport, such as Na+ and K+ movement, to the facilitation of H+ elimination. Plume and trophosome ATPase activity for R. pachyptila measured 646.2 ± 29.5 and 481.4 ± 32.0 μmol Pi (inorganic phosphate) g−1 wet wt h−1, respectively. Plume tissue ATPase activity (both mass-specific and protein-specific) in R. pachyptila was higher (between 7% and 55%) than the activity measured in any tissue for 7 other shallow- and deep-living species, in this study. This supports the hypothesis that R. pachyptila regulates acid/base balance via high concentrations of H+-ATPases, including Na+/H+ and K+/H+ exchangers and possibly electrogenic H+-ATPases, as evidenced by a higher total ATPase concentration (646 μmol Pi g−1 wet wt h−1), lesser Na+/K+-ATPase activity (13% of the total, as compared to 20−40% found in other animals), and higher H+-ATPase activity (226–264 μmol Pi g−1wet wt h−1). Overall, R. pachyptila appears to demonstrate elevated ATPase activity, with a greater fraction of the enzymes devoted to proton elimination, in order to effectively control its extracellular pH in the face of processes acting to acidify the internal environment.

Nitrogen metabolites and related enzymatic activities in the body fluids and tissues of the hydrothermal vent tubeworm Riftia pachyptila.

The distribution of nitrogen metabolism end-products and the associated enzyme activities, free amino acids and purine base catabolites were investigated in all the body compartments (circulating fluids and tissues) of the hydrothermal vent tubeworm Riftia pachyptila to acquire a general overview of nitrogen metabolism in this symbiotic organism. There were striking differences between the symbiont-containing trophosome tissue and other host tissues. High concentrations of ammonia, creatinine and, in particular, urate were found in all tissues, but they were present at consistently higher concentrations in the trophosome, which also contained large amounts of urea. Uric acid crystals were present at the periphery of trophosome lobules. The urea cycle appears to be fully functional in this tissue, which also uses creatine phosphate for phosphagen storage, while arginine phosphate or a combination of both phosphagens occurs in other tissues. The amino acid patterns are dominated by sulphated compounds in all tissues except the trophosome, which has high levels of aspartate and glutamate. Although no definitive conclusions could be drawn regarding the nitrogen regime of Riftia pachyptila, this in vitro study gives several indications for future research in this area.

Fate of nitrate acquired by the tubeworm Riftia pachyptila.

The hydrothermal vent tubeworm Riftia pachyptila lacks a mouth and gut and lives in association with intracellular, sulfide-oxidizing chemoautotrophic bacteria. Growth of this tubeworm requires an exogenous source of nitrogen for biosynthesis, and, as determined in previous studies, environmental ammonia and free amino acids appear to be unlikely sources of nitrogen. Nitrate, however, is present in situ (K. Johnson, J. Childress, R. Hessler, C. Sakamoto-Arnold, and C. Beehler, Deep-Sea Res. 35:1723-1744, 1988), is taken up by the host, and can be chemically reduced by the symbionts (U. Hentschel and H. Felbeck, Nature 366:338-340, 1993). Here we report that at an in situ concentration of 40 microM, nitrate is acquired by R. pachyptila at a rate of 3.54 micromol g(-1) h(-1), while elimination of nitrite and elimination of ammonia occur at much lower rates (0. 017 and 0.21 micromol g(-1) h(-1), respectively). We also observed reduction of nitrite (and accordingly nitrate) to ammonia in the trophosome tissue. When R. pachyptila tubeworms are exposed to constant in situ conditions for 60 h, there is a difference between the amount of nitrogen acquired via nitrate uptake and the amount of nitrogen lost via nitrite and ammonia elimination, which indicates that there is a nitrogen "sink." Our results demonstrate that storage of nitrate does not account for the observed stoichiometric differences in the amounts of nitrogen. Nitrate uptake was not correlated with sulfide or inorganic carbon flux, suggesting that nitrate is probably not an important oxidant in metabolism of the symbionts. Accordingly, we describe a nitrogen flux model for this association, in which the product of symbiont nitrate reduction, ammonia, is the primary source of nitrogen for the host and the symbionts and fulfills the association's nitrogen needs via incorporation of ammonia into amino acids.

Identification and characterization of a flagellin gene from the endosymbiont of the hydrothermal vent tubeworm Riftia pachyptila.

The bacterial endosymbionts of the hydrothermal vent tubeworm Riftia pachyptila play a key role in providing their host with fixed carbon. Results of prior research suggest that the symbionts are selected from an environmental bacterial population, although a free-living form has been neither cultured from nor identified in the hydrothermal vent environment. To begin to assess the free-living potential of the symbiont, we cloned and characterized a flagellin gene from a symbiont fosmid library. The symbiont fliC gene has a high degree of homology with other bacterial flagellin genes in the amino- and carboxy-terminal regions, while the central region was found to be nonconserved. A sequence that was homologous to that of a consensus sigma28 RNA polymerase recognition site lay upstream of the proposed translational start site. The symbiont protein was expressed in Escherichia coli, and flagella were observed by electron microscopy. A 30,000-Mr protein subunit was identified in whole-cell extracts by Western blot analysis. These results provide the first direct evidence of a motile free-living stage of a chemoautotrophic symbiont and support the hypothesis that the symbiont of R. pachyptila is acquired with each new host generation.

The ionic composition of the hydrothermal vent tube worm Riftia pachyptila: evidence for the elimination of SO2-4SO and H+ and for a Cl-/HCO-3HCO shift.

Riftia pachyptila is one of the most specialized invertebrate hosts of chemoautotrophic symbionts. Crucial to the functioning of this symbiosis is how these worms cope with fluctuating ion concentrations. Internal sulfate levels in R. pachyptila appear comparable with other benthic marine invertebrates, despite the production of sulfate internally by means of the bacterial oxidation of hydrogen sulfide, suggesting that these worms are able to eliminate sulfate effectively. Internal chloride levels appear comparable; however, coelomic fluid chloride levels decrease significantly as the amount of coelomic fluid bicarbonate increases, demonstrating a 1:1 stoichiometry. We believe this shift in chloride, out of the body fluids, is needed to compensate for changes in electrochemical balance caused by the large increase (up to and greater than 60 mmol L-1) in negatively charged bicarbonate. Riftia pachyptila fits the general pattern of monovalent ion concentrations that is seen in other benthic marine invertebrates, with a high [Na+] : [K+] ratio extracellularly and low [Na+] : [K+] ratio intracellularly. Extracellular pH values of 7.38+/-0.03 and 7.37+/-0. 04 for coelomic fluid and vascular blood, respectively, as well as intracellular pH values of 7.37+/-0.04 and 7.04+/-0.05 for plume and trophosome tissue, respectively, were measured. On the basis of significant decreases in extracellular pH and, in some cases, Na+ and K+, in worms exposed to carbonyl cyanide m-chlorophenylhydrazone, sodium vanadate, and N-ethylmaleimide, we suggest that high concentrations of H+-ATPases, perhaps Na+/H+- or K+/H+-ATPases, are involved in H+ elimination in these animals.

Partially glucose-capped oligosaccharides are found on the hemoglobins of the deep-sea tube worm Riftia pachyptila.

We report here the structural determination of N-linked oligosaccharides found on extracellular hemoglobins of the hydrothermal vent tube worm Riftia pachyptila. Structures were elucidated by a combination of electrospray ionization tandem mass spectrometry, matrix-assisted laser desorption/ionization mass spectrometry, normal-phase high performance liquid chromatography, and exoglycosidase digestion. The sugar chains were found to consist mainly of high-mannose-type glycans with some structures partially capped by one or two terminal glucose residues. The present study represents the first report of the occurrence of glucose capping of N-linked carbohydrates in a secreted glycoprotein of a metazoan. Previously, glucose capping has only been described for a membrane-bound surface glycoprotein from the unicellular parasite Leishmania mexicana amazonensis.

Metabolic responses of the hydrothermal vent tube worm Riftia pachyptila to severe hypoxia

MEPS Marine Ecology Progress Series Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsTheme Sections MEPS 174:151-158 (1998) - doi:10.3354/meps174151 Metabolic responses of the hydrothermal vent tube worm Riftia pachyptila to severe hypoxia Cordelia Arndt1,2,*, Doris Schiedek2, Horst Felbeck1 1University of California San Diego, Scripps Institution of Oceanography, La Jolla, California 92093-0202, USA 2Baltic Sea Research Institute at the University of Rostock, Seestrasse 15, D-18119 Rostock-Warnemuende, Germany *E-mail: coarndt@ucsd.edu ABSTRACT: The metabolic capabilities of the hydrothermal vent tube worm Riftia pachyptila to tolerate short- and long-term exposure to hypoxia were investigated. After incubating specimens under anaerobic conditions the metabolic changes in body fluids and tissues were analyzed over time. The tube worms tolerated anoxic exposure up to 60 h. Prior to hypoxia the dicarboxylic acid, malate, was found in unusually high concentrations in the blood (up to 26 mM) and tissues (up to 5 µmol g-1 fresh wt). During hypoxia, most of the malate was degraded very quickly, while large quantities of succinate accumulated (blood: about 17 mM; tissues: about 13 µmol g-1 fresh wt). Volatile, short-chain fatty acids were apparently not excreted under these conditions. The storage compound, glycogen, was mainly found in the trophosome and appears to be utilized only during extended anaerobiosis. The succinate formed during hypoxia does not account for the use of malate and glycogen, which possibly indicates the presence of yet unidentified metabolic end products. Glutamate concentration in the trophosome decreased markedly during hypoxia, presumably due to a reduction in the autotrophic function of the symbionts during hypoxia. In conclusion, R. pachyptila is physiologically well adapted to the oxygen fluctuations frequently occurring in the vent habitat. KEY WORDS: Riftia pachyptila · Anaerobic metabolism · Hypoxia · Hydrothermal vents Full text in pdf format PreviousNextExport citation RSS - Facebook - Tweet - linkedIn Cited by Published in MEPS Vol. 174. Publication date: November 26, 1998 Print ISSN:0171-8630; Online ISSN:1616-1599 Copyright © 1998 Inter-Research.

Primary structure of the common polypeptide chain b from the multi‐hemoglobin system of the hydrothermal vent tube worm Riftia pachyptila: An insight on the sulfide binding‐site

The deep-sea tube worm Riftia pachyptila Jones possesses a multi-hemoglobin system with three different extracellular Hbs: two dissolved in the vascular blood, V1 (ca. 3,500 kDa) and V2 (ca. 400 kDa), and one in the coelomic fluid, C1 (ca. 400 kDa). V1 Hb consists of four heme-containing, globin chains (b–e) and four linker chains (L1–L4). V2 and C1 Hbs are exclusively built from globin chains, six for V2 (a–f) and five for C1 (a–e). The complete amino acid sequence of the isolated monomeric globin chain b, common to all Riftia Hbs, has been determined by automated Edman degradation sequencing of the peptides derived by digestion with trypsin, chymotrypsin, thermolysin, and CNBr. This polypeptide chain is composed of 144 amino acid residues, providing a Mr of 16, 135.0 Da. Moreover, the primary sequence of chain b revealed 3 Cys residues at position 4, 75, and 134. Cys-4 and Cys-134 are located at positions where an intra-chain disulfide bridge is formed in all annelid, vestimentiferan, or pogonophoran chains, but Cys-75 is located at a unique position only found in three globin chains belonging to Lamellibrachia and Oligobrachia, a vestimentiferan and a pogonophoran. In both groups, Hbs can bind sulfide reversibly to fuel the chemosynthetic process of the symbiotic bacteria they harbor. Sulfide-binding experiments performed on purified Hb fractions (i.e., V1, V2, and C1 Hbs) suggest that free Cys residues on globin chains, and the numerous Cys found in linker chains, as determined previously by ESI-MS, may be the sulfide binding-sites. Blocking the free Cys by N-ethylmaleimide, we confirmed that free cysteines were involved in sulfide-binding but did not account for the whole sulfide-binding capacity of V1 Hb. Furthermore, a phylogenetic tree was constructed from 18 globin-like chains of annelid, vetimentiferan, and pogonophoran extracellular Hbs to clarify the systematic position of tubeworms. Riftia chain b clearly belongs to the “strain A” family with 30 to 80% identity with the other sequences analyzed. Its position in the tree confirmed a close relationship between vestimentiferan, pogonophoran, and annelid Hbs. Proteins 29:562–574, 1997. © 1997 Wiley-Liss, Inc.