Octopine Dehydrogenase Research Articles

Expression of pyruvate kinase, malate and octopine dehydrogenase genes in the gills of the Mediterranean mussel Mytilus galloprovincialis (Lamarck, 1819) under conditions of hypoxia and reoxygenation

The effect of hypoxia on the expression level of pyruvate kinase (PKM), octopine dehydrogenase (OcDH) and malate dehydrogenase (MDH) genes in the gill tissue of the Mediterranean mussel Mytilus galloprovincialis (L., 1819) was studied experimentally. The control group of mollusks was kept at 9–10°C and the oxygen level in the water was 8.5 mgO2 l–1. Experimental – at 9–10°C and 2.2 mgO2 l–1. The exposure was 24 and 72 hours. Some of the individuals were subsequently subjected to the reoxygenation procedure. Under conditions of hypoxia, the expression level of OcDH and MDH genes increased by 3 and 2 times, respectively (p 0.05). At the same time, OcDH expression showed sensitivity to the oxygen content in the medium. Under the conditions of reoxygenation, the process was completely suppressed. The expression of the PKM gene, on the contrary, did not depend on hypoxic effects and remained at the level of control values.

Reproductive effort affects cellular response in the mantle of Nodipecten subnodosus scallops exposed to acute hyperthermia

In marine ecosystems, temperature regulates the energy metabolism of animals. In the last decades, the temperature increase was related to mass mortality events of marine ectotherms, particularly during high-energy investment for reproduction. In scallops, the mantle has been poorly investigated while this tissue covers more than 40 % of the body mass, contributing to the perception of surrounding environmental stimuli. Our aim was to assess the cellular and molecular responses linked to energy metabolism in the mantle of adult N. subnodosus facing acute hyperthermia during reproductive effort. Scallops collected in spring (late gametogenesis) and summer (ripe gonads) were exposed to a control temperature (22 °C) or acute hyperthermia (30 °C) for 24 h. In spring, increased arginine kinase (AK) activity together with increased pyruvate kinase/citrate synthase ratio (PK/CS) suggested an enhanced carbohydrate, pyruvate, and arginine metabolism to maintain the adenylate energy charge (AEC) in the mantle of scallops coping with acute thermal increase. In summer, animals decreased their AEC (5 %) and arginine phosphate pool (40 %) and increased their anaerobic metabolism as shown by enhanced activities of lactate-dehydrogenase (LDH) and octopine dehydrogenase (ODH), respectively. The abundance of twenty proteins involved in energy metabolism (isocitrate dehydrogenase, ATP synthase subunit β), protein protection (cognates heat shock protein 70), and cytoskeleton (actins and tubulins) were affected only by season. These results underlie the role of the mantle of N. subnodosus in the seasonal responses of this tissue to thermal fluctuations during reproductive effort with possible implications for the physiological performance of scallops under heat waves in wild or harvest conditions.

Behavioral Characteristics and Related Physiological and Ecological Indexes of Cultured Scallops (Mizuhopecten yessoensis) in Response to Predation by the Crab Charybdis japonica

To investigate the effects of predation by the paddle crab Charybdis japonica on the culture and survival of scallops (Mizuhopecten yessoensis) during bottom culture, we investigated the behavioral characteristics of three sizes (small, medium, and large) of scallops in response to exposure to crabs. We found that scallops escaped from crab predation by continuous shell closure or movement. Shell closure force increased with scallop size, and scallops of the same size that were stimulated by the presence of crabs closed their shell more frequently than control scallops. We also measured the activities of superoxide dismutase, catalase, arginine kinase, and octopine dehydrogenase in the gill, adductor muscle, and mantle of scallops before and after exposure to predation. Tissues that showed significant differences between control and test specimens were selected for deep sequencing of the transcriptome to identify and validate the key genes that were sensitive to predation. We found that when M. yessoensis is stimulated by the presence of predators, its behavioral characteristics and related physiological and ecological indexes undergo significant changes. The results are relevant for developing specifications for M. yessoensis seedling casting during bottom culture.

Expression of Pyruvate Kinase, Malate and Octopine Dehydrogenase Genes in the Gills of the Mediterranean Mussel Mytilus galloprovincialis (Lamarck, 1819) under Conditions of Hypoxia and Reoxygenation

Expression of Pyruvate Kinase, Malate and Octopine Dehydrogenase Genes in the Gills of the Mediterranean Mussel Mytilus galloprovincialis (Lamarck, 1819) under Conditions of Hypoxia and Reoxygenation

Behavioral characteristics and transcriptome analysis of Mizuhopecten yessoensis in response to Neptunea arthritica cumingii predation during laboratory bottom-sowing culture

In this study, we investigated the effects of predation by the whelk Neptunea arthritica cumingii on the culture and survival of scallops (Mizuhopecten yessoensis) during bottom culture. The behavioral characteristics of three sizes (small, medium, and large) of scallops were assessed in response to exposure to N. cumingii. We also measured the activities of superoxide dismutase, catalase, arginine kinase, and octopine dehydrogenase in the gill, adductor muscle, and mantle tissues of scallops before and after exposure to predation. We found that scallops escaped from N. cumingii predation by continuous shell closure or movement. The shell closure force increased with the scallop size, and scallops of the same size that were stimulated by the presence of N. cumingii closed their shells more frequently than control scallops. The shell closure ability of scallops of all sizes decreased as the duration of the experiment continued. The enzyme activities in the three scallop tissues changed significantly after continuous stimulation, and the trend was particularly obvious for the enzyme activities in the adductor muscles of medium-sized scallops. Transcriptome analysis of the adductor muscles from medium-sized scallops detected 405 differentially expressed genes (172 upregulated and 233 downregulated), where the expression levels of RYK, APC, BAMBI, WNT16, CACYBP, and RUVBL1 changed after stimulation by N. cumingii. The sequencing results were verified by quantitative real-time PCR. We hypothesize these genes may be related to muscle movement and regulation in M. yessoensis after stimulation.

Protein metabolism and immune response levels in cuttlefish (Sepia pharaonis) and the effects of inking stress on development

Inking is a defensive behavior that protects cephalopods from predator attack, a process that is accomplished by generating jet propulsion from powerful contractions of the mantle muscle. However, most studies have focused on the “defensive effects” of the ink, with the result that there is limited information concerning the effects of inking on the animals themselves. On the basis of our previous research, we integrated information from different tissues after continuous ink release to develop a better understanding of the physiological stress resulting from inking in the cuttlefish (Sepia pharaonis). The results showed that throughout the recovery period (60 days) of the inking treatment, the observed declines in growth performance due to decreased protein synthesis and increased energy metabolism levels were supported by a decrease in mantle muscle glycogen concentration (P < 0.01), increased hexokinase (HK) and pyruvate kinase (PK) specifical activities (P < 0.001), up-regulated expression of arginine kinase (AK) and octopine dehydrogenase (ODH) (P < 0.001), and up-regulated immune superoxide dismutase (SOD) and heat shock protein 70 (HSP70) in the digestive gland (P < 0.001). Muscle principal component analysis (PCA) revealed that amino acids (lysine, tyrosinase, arginine, leucine, isoleucine, and aspartate) were involved in protein turnover after the ink release, and essential fatty acids such as oleic acid (18:1n-9), linoleic acid (18:2n-6), arachidonic acid (ARA, 20:4n-6), and docosahexaenoic acid (DHA, 22:6n-3) were disturbed by inking stress. Moreover, the inducible HSP70 in the digestive gland was sensitively up-regulated even after the 60-day recovery, and thus the transcription level of induced HSP70 in cuttlefish may be used as an indicator of physiological stress. In conclusion, a high growth rate requires a high level of protein synthesis, and it appears that considerable ink release has a significant impact on cuttlefish (S. pharaonis) physiological stress. The transcription level of HSP70 may be used as a stress indicator

Seasonal metabolic and oxidative stress responses of commercially important invertebrate species—correlation with their habitat

Habitat conditions play a crucial role in food acquisition and the energetic and physiological performance of marine organisms during seasonal changes. However, across a wide distribution, populations of the same species might face physiological constraints driven by the oceanographic and climatological characteristics of their habitats. In the context of climate change, biochemical and metabolic data may be important in identifying marine areas in which inhabiting organisms are more or less physiologically constrained. As a first step in identifying such marine areas on the Greek coast, we studied the seasonal metabolic and antioxidant responses of blue crab Callinectes sapidus, cuttlefish Sepia officinalis, sea cucumber Holothuria tubulosa and clam Venus verrucosa, collected from 3 different Greek gulfs: Thermaikos, Pagasitikos and Vistonikos. Seasonal analysis of metabolic responses was based on the activity of the following intermediary metabolic enzymes: lactate dehydrogenase (L-LDH), octopine dehydrogenase (ODH), β-hydroxyacyl-CoA dehydrogenase (HOAD) and citrate synthase (CS). Antioxidant responses were based on the activities of the following enzymes: superoxide dismutase (SOD), glutathione reductase (GR) and catalase, and on lipid peroxidation estimation. The blue crab and clam exhibited the most significant seasonal changes compared to the sea cucumber and cuttlefish, which exhibited no changes. The intermediary metabolism enzymatic activity of blue crab and clam decreased and antioxidant enzymatic activity increased during summer. Higher activity levels for all examined species were observed in Pagasitikos and lower levels in Vistonikos Gulf, indicating corresponding oxidative stress. These results are correlated with the spatial oceanographic conditions of the habitats and are discussed based on future projections of global warming in the Mediterranean Sea.

Effects of microplastics and mercury in the freshwater bivalve Corbicula fluminea (Müller, 1774): Filtration rate, biochemical biomarkers and mercury bioconcentration

The main objectives of this study were to investigate the effects of a mixture of microplastics and mercury on Corbicula fluminea, the post-exposure recovery, and the potential of microplastics to influence the bioconcentration of mercury by this species. Bivalves were collected in the field and acclimated to laboratory conditions for 14 days. Then, a 14-day bioassay was carried out. Bivalves were exposed for 8 days to clean medium (control), microplastics (0.13 mg/L), mercury (30 µg/L) and to a mixture (same concentrations) of both substances. The post-exposure recovery was investigated through 6 additional days in clean medium. After 8 and 14 days, the following endpoints were analysed: the post-exposure filtration rate (FR); the activity of cholinesterase enzymes (ChE), NADP-dependent isocitrate dehydrogenase (IDH), octopine dehydrogenase, catalase, glutathione reductase, glutathione peroxidase and glutathione S-transferases (GST), and the levels of lipid peroxidation (LPO). After 8 days of exposure to mercury, the bioconcentration factors (BCF) were 55 in bivalves exposed to the metal alone and 25 in bivalves exposed to the mixture. Thus, microplastics reduced the bioconcentration of mercury by C. fluminea. Bivalves exposed to microplastics, mercury or to the mixture had significantly (p ≤ 0.05) decreased FR and increased LPO levels, indicating fitness reduction and lipid oxidative damage. In addition, bivalves exposed to microplastics alone had significant (p ≤ 0.05) reduction of adductor muscle ChE activity, indicating neurotoxicity. Moreover, bivalves exposed to mercury alone had significantly (p ≤ 0.05) inhibited IDH activity, suggesting alterations in cellular energy production. Antagonism between microplastics and mercury in FR, ChE activity, GST activity and LPO levels was found. Six days of post-exposure recovery in clean medium was not enough to totally reverse the toxic effects induced by the substances nor to eliminate completely the mercury from the bivalve’s body. These findings have implications to animal, ecosystem and human health.

Genetic structure of Octopus minor around Chinese waters as indicated by nuclear DNA variations (Mollusca, Cephalopoda).

Octopus minor is an economically important resource commonly found in Chinese coastal waters. The nuclear gene (RD and ODH) approach of investigation has not reported in this species. Rhodopsin (RD) and octopine dehydrogenase (ODH) genes were used to elaborate the genetic structure collected from eight localities ranging from the northern to the southern coast of China. In total, 118 individuals for the RD gene and 108 for the ODH were sequenced. Overall (RD and ODH) genes resulted in high (0.741±0.032; 0.805±0.038) haplotype and low nucleotide (0.01261±0.00165; 0.00747±0.00086) diversity. Molecular variance displayed higher values among the populations and lower values within the population where the fixation index FST denoted 0.880 and 0.584 in RD and ODH genes respectively. The Dongshan population clustered separately in a phylogenetic tree as in the haplotype networking assessment. The current data suggests that the Dongshan population needs separate management.

Toxicity of mercury and post-exposure recovery in Corbicula fluminea: Neurotoxicity, oxidative stress and oxygen consumption

The toxicity of mercury to the invasive species Corbicula fluminea and the post-exposure recovery were investigated in relation to previous developmental exposure to distinct environmental conditions. Bivalves were collected in the estuaries of Minho River (M-est) and Lima River (L-est) that have several abiotic differences, including in environmental contamination, with the former being generally less contaminated. After 14 days of acclimation to laboratory conditions, two 14-day bioassays were performed simultaneously: one with bivalves from the M-est and the other with bivalves from the L-est. In each bioassay, the treatments were: dechlorinated tap water (clean medium) for 8 days, clean medium for 14 days, 31 µg/L of mercury for 8 days, 31 µg/L of mercury for 14 days and 31 µg/L of mercury for 8 days followed by 6 days in clean medium (post-exposure recovery). The effect criteria were the oxygen consumption rate (OCR), the activity of the enzymes cholinesterases (ChE), NADP-dependent isocitrate dehydrogenase (IDH), octopine dehydrogenase (ODH), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx) and glutathione S-transferases (GST) and lipid peroxidation (LPO) levels. Exposure to mercury for 8 days caused significant (p ≤ 0.05) inhibition of GR activity in M-est bivalves, whereas no significant adverse effects were observed in L-est animals. Moreover, evidences of delayed toxicity caused by 8-day exposure to mercury in OCR, IDH activity and LPO levels were found in M-est individuals but not in those of the L-est. Exposure to mercury for 14 days caused significant (p ≤ 0.05) depression of the OCR and of IDH activity in animals from both estuaries, indicating reduced individual fitness and hypoxia conditions. Moreover, oxidative stress and lipid peroxidation damage were observed in bivalves from the M-est exposed to mercury for 14 days but not in L-est animals. Differences in M-est and L-est environmental conditions to which animals were exposed in the wild likely contributed to the differences of sensitivity to mercury between M-est and L-est bivalves. The results of this study highlight the importance of investigating delayed toxicity, post-exposure recovery, and of taking into consideration the background contamination and other abiotic conditions of the original habitats when assessing the effects of environmental contaminants on animals from wild populations.

Time Course of Metabolic Capacities in Paralarvae of the Common Octopus, Octopus vulgaris, in the First Stages of Life. Searching Biomarkers of Nutritional Imbalance.

The culture of the common octopus (Octopus vulgaris) is promising since the species has a relatively short lifecycle, rapid growth, and high food conversion ratios. However, recent attempts at successful paralarvae culture have failed due to slow growth and high mortality rates. Establishing an optimal nutritional regime for the paralarvae seems to be the impeding step in successful culture methods. Gaining a thorough knowledge of food regulation and assimilation is essential for paralarvae survival and longevity under culture conditions. The aim of this study, then, was to elucidate the characteristic metabolic organization of octopus paralarvae throughout an ontogenic period of 12 days post-hatching, as well as assess the effect of diet enrichment with live prey containing abundant marine phospholipids. Our results showed that throughout the ontogenic period studied, an increase in anaerobic metabolism took place largely due to an increased dependence of paralarvae on exogenous food. Our studies showed that this activity was supported by octopine dehydrogenase activity, with a less significant contribution of lactate dehydrogenase activity. Regarding aerobic metabolism, the use of amino acids was maintained for the duration of the experiment. Our studies also showed a significant increase in the rate of oxidation of fatty acids from 6 days after-hatching. A low, although sustained, capacity for de novo synthesis of glucose from amino acids and glycerol was also observed. Regardless of the composition of the food, glycerol kinase activity significantly increased a few days prior to a massive mortality event. This could be related to a metabolic imbalance in the redox state responsible for the high mortality. Thus, glycerol kinase might be used as an effective nutritional and welfare biomarker. The studies in this report also revealed the important finding that feeding larvae with phospholipid-enriched Artemia improved animal viability and welfare, significantly increasing the rate of survival and growth of paralarvae.

Changes to coral health and metabolic activity under oxygen deprivation.

On Hawaiian reefs, the fast-growing, invasive algae Gracilaria salicornia overgrows coral heads, restricting water flow and light, thereby smothering corals. Field data shows hypoxic conditions (dissolved oxygen (DO2) < 2 mg/L) occurring underneath algal mats at night, and concurrent bleaching and partial tissue loss of shaded corals. To analyze the impact of nighttime oxygen-deprivation on coral health, this study evaluated changes in coral metabolism through the exposure of corals to chronic hypoxic conditions and subsequent analyses of lactate, octopine, alanopine, and strombine dehydrogenase activities, critical enzymes employed through anaerobic respiration. Following treatments, lactate and octopine dehydrogenase activities were found to have no significant response in activities with treatment and time. However, corals subjected to chronic nighttime hypoxia were found to exhibit significant increases in alanopine dehydrogenase activity after three days of exposure and strombine dehydrogenase activity starting after one overnight exposure cycle. These findings provide new insights into coral metabolic shifts in extremely low-oxygen environments and point to ADH and SDH assays as tools for quantifying the impact of hypoxia on coral health.

Characterization of Flavin-Containing Opine Dehydrogenase from Bacteria.

Opines, in particular nopaline and octopine, are specific compounds found in crown gall tumor tissues induced by infections with Agrobacterium species, and are synthesized by well-studied NAD(P)H-dependent dehydrogenases (synthases), which catalyze the reductive condensation of α-ketoglutarate or pyruvate with L-arginine. The corresponding genes are transferred into plant cells via a tumor-inducing (Ti) plasmid. In addition to the reverse oxidative reaction(s), the genes noxB-noxA and ooxB-ooxA are considered to be involved in opine catabolism as (membrane-associated) oxidases; however, their properties have not yet been elucidated in detail due to the difficulties associated with purification (and preservation). We herein successfully expressed Nox/Oox-like genes from Pseudomonas putida in P. putida cells. The purified protein consisted of different α-, β-, and γ-subunits encoded by the OdhA, OdhB, and OdhC genes, which were arranged in tandem on the chromosome (OdhB-C-A), and exhibited dehydrogenase (but not oxidase) activity toward nopaline in the presence of artificial electron acceptors such as 2,6-dichloroindophenol. The enzyme contained FAD, FMN, and [2Fe-2S]-iron sulfur as prosthetic groups. On the other hand, the gene cluster from Bradyrhizobium japonicum consisted of OdhB 1 -C-A-B 2, from which two proteins, OdhAB1C and OdhAB2C, appeared through the assembly of each β-subunit together with common α- and γ-subunits. A poor phylogenetic relationship was detected between OdhB1 and OdhB2 in spite of them both functioning as octopine dehydrogenases, which provided clear evidence for the acquisition of novel functions by “subunit-exchange”. To the best of our knowledge, this is the first study to have examined flavin-containing opine dehydrogenase.

Proteomic responses to hypoxia at different temperatures in the great scallop (Pecten maximus).

Hypoxia and hyperthermia are two connected consequences of the ongoing global change and constitute major threats for coastal marine organisms. In the present study, we used a proteomic approach to characterize the changes induced by hypoxia in the great scallop, Pecten maximus, subjected to three different temperatures (10 °C, 18 °C and 25 °C). We did not observe any significant change induced by hypoxia in animals acclimated at 10 °C. At 18 °C and 25 °C, 16 and 11 protein spots were differentially accumulated between normoxia and hypoxia, respectively. Moreover, biochemical data (octopine dehydrogenase activity and arginine assays) suggest that animals grown at 25 °C switched their metabolism towards anaerobic metabolism when exposed to both normoxia and hypoxia, suggesting that this temperature is out of the scallops’ optimal thermal window. The 11 proteins identified with high confidence by mass spectrometry are involved in protein modifications and signaling (e.g., CK2, TBK1), energy metabolism (e.g., ENO3) or cytoskeleton (GSN), giving insights into the thermal-dependent response of scallops to hypoxia.

Influence of Long-Term Exposure to Background Pollution on the Response and Recovery of the Invasive Species Corbicula fluminea to Ammonia Sub-lethal Stress: a Multi-marker Approach with Field Estuarine Populations

The influence of long-term exposure to background pollution on the response and recovery of the invasive species Corbicula fluminea to ammonia stress was investigated using a multi-marker approach. Wild clams of the tidal freshwater areas of two estuaries of the NW Iberian coast with different levels of pollution, the estuaries of Minho river (reference) and of Lima river (contaminated), were collected and exposed individually to different treatments: 8 and 14 days in dechlorinated tap water (DTW), 8 and 14 days in 1 mg L−1 of ammonia (AM), and 8 days in AM followed by 6 days in DTW. After each defined time (0, 8, and 14 days), the clams were sacrificed and the activity of the enzymes glutathione S-transferase (GST), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx), cholinesterase (ChE), octopine dehydrogenase (ODH), and the lipid peroxidation (LPO) levels were used as effect criteria. At the beginning of the bioassay, the clams from the polluted estuary presented significantly higher background levels of GST, CAT, GR, GPx, and LPO than those from the reference one indicating long-term exposure to oxidative stressors. In general, C. fluminea from both estuaries presented little sensibility to ammonia with no significant differences found between exposed and control clams for most of the biomarkers. That low sensibility of C. fluminea could be seen as advantageous for its invasion ability.

Anaerobiosis and metabolic plasticity of Pinna nobilis: Biochemical and ecological features

Changes in the energetic metabolism were studied in the fan mussel Pinna nobilis L. exposed to environmental and anthropic stress. The high polymorphism of enzymes suggests an adaptation of the fan mussel to environmental variability peculiar of transitional waters with respect to the same species living in exposed coastal sea. The electrophoretic patterns showed a predominance of LDH-A4 and the presence of both mitochondrial and cytosolic MDH isozymes. Moreover, in all the analyzed tissues and organs, MDH activity was greater than the LDH one. Metabolic plasticity of the fan mussel is further highlighted by octopine dehydrogenase and superoxide dismutase electrophoretic patterns, showing the presence of many isoforms. These evidences are also confirmed by spectroscopic determinations of alanopine, tauropine, strombine and octopine dehydrogenase activity characterized by a specific trend due to environmental variability. Specific variations in anaerobic capacity of P. nobilis L. are discussed in relation to their distribution according to the marine-brackish gradient.

Molecular evidence for co-occurring cryptic lineages within the Sepioteuthis cf. lessoniana species complex in the Indian and Indo-West Pacific Oceans

The big-fin reef squid, Sepioteuthis cf. lessoniana (Lesson 1930), is an important commodity species within artisanal and near-shore fisheries in the Indian and Indo-Pacific regions. While there has been some genetic and physical evidence that supports the existence of a species complex within S. cf. lessoniana, these studies have been extremely limited in scope geographically. To clarify the extent of cryptic diversity within S. cf. lessoniana, this study examines phylogenetic relationships using mitochondrial genes (cytochrome oxidase c, 16s ribosomal RNA) and nuclear genes (rhodopsin, octopine dehydrogenase) from nearly 400 individuals sampled from throughout the Indian, Indo-Pacific, and Pacific Ocean portions of the range of this species. Phylogenetic analyses using maximum likelihood methods and Bayesian inference identified three distinct lineages with no clear geographic delineations or morphological discriminations. Phylogeographic structure analysis showed high levels of genetic connectivity in the most widespread lineage, lineage C and low levels of connectivity in lineage B. This study provides significant phylogenetic evidence for cryptic lineages within this complex and confirms that cryptic lineages of S. cf. lessoniana occur in sympatry at both small and large spatial scales. Furthermore, it suggests that two closely related co-occurring cryptic lineages have pronounced differences in population structure, implying that underlying differences in ecology and/or life history may facilitate co-occurrence. Further studies are needed to assess the range and extent of cryptic speciation throughout the distribution of this complex. This information is extremely useful as a starting point for future studies exploring the evolution of diversity within Sepioteuthis and can be used to guide fisheries management efforts.

Gene Expression and Physiological Changes of Different Populations of the Long-Lived Bivalve Arctica islandica under Low Oxygen Conditions

The bivalve Arctica islandica is extremely long lived (>400 years) and can tolerate long periods of hypoxia and anoxia. European populations differ in maximum life spans (MLSP) from 40 years in the Baltic to >400 years around Iceland. Characteristic behavior of A. islandica involves phases of metabolic rate depression (MRD) during which the animals burry into the sediment for several days. During these phases the shell water oxygen concentrations reaches hypoxic to anoxic levels, which possibly support the long life span of some populations. We investigated gene regulation in A. islandica from a long-lived (MLSP 150 years) German Bight population and the short-lived Baltic Sea population, experimentally exposed to different oxygen levels. A new A. islandica transcriptome enabled the identification of genes important during hypoxia/anoxia events and, more generally, gene mining for putative stress response and (anti-) aging genes. Expression changes of a) antioxidant defense: Catalase, Glutathione peroxidase, manganese and copper-zinc Superoxide dismutase; b) oxygen sensing and general stress response: Hypoxia inducible factor alpha, Prolyl hydroxylase and Heat-shock protein 70; and c) anaerobic capacity: Malate dehydrogenase and Octopine dehydrogenase, related transcripts were investigated. Exposed to low oxygen, German Bight individuals suppressed transcription of all investigated genes, whereas Baltic Sea bivalves enhanced gene transcription under anoxic incubation (0 kPa) and, further, decreased these transcription levels again during 6 h of re-oxygenation. Hypoxic and anoxic exposure and subsequent re-oxygenation in Baltic Sea animals did not lead to increased protein oxidation or induction of apoptosis, emphasizing considerable hypoxia/re-oxygenation tolerance in this species. The data suggest that the energy saving effect of MRD may not be an attribute of Baltic Sea A. islandica chronically exposed to high environmental variability (oxygenation, temperature, salinity). Contrary, higher physiological flexibility and stress hardening may predispose these animals to perform a pronounced stress response at the expense of life span.

Nuclear and mitochondrial markers reveal evidence for genetically segregated cryptic speciation in giant Pacific octopuses from Prince William Sound, Alaska

Multiple species of large octopus are known from the north Pacific waters around Japan, however only one large species is known in the Gulf of Alaska (the giant Pacific octopus, Enteroctopus dofleini). Current taxonomy of E. dofleini is based on geographic and morphological characteristics, although with advances in genetic technology that is changing. Here, we used two mitochondrial genes (cytochrome b and cytochrome oxidase I), three nuclear genes (rhodopsin, octopine dehydrogenase, and paired-box 6), and 18 microsatellite loci for phylogeographic and phylogenetic analyses of octopuses collected from across southcentral and the eastern Aleutian Islands (Dutch Harbor), Alaska. Our results suggest the presence of a cryptic Enteroctopus species that is allied to, but distinguished from E. dofleini in Prince William Sound, Alaska. Existence of an undescribed and previously unrecognized taxon raises important questions about the taxonomy of octopus in southcentral Alaska waters.

Control of d-octopine formation in scallop adductor muscle as revealed through thermodynamic studies of octopine dehydrogenase

Octopine dehydrogenase (OcDH) from the adductor muscle of the great scallop, Pecten maximus (Linné, 1758), catalyses the NADH-dependent condensation of l-arginine and pyruvate to d-octopine, NAD(+) and water during escape swimming and subsequent recovery. During exercise, ATP is mainly provided by the transphosphorylation of phospho-l-arginine and to some extent by anaerobic glycolysis. NADH resulting from the glycolytic oxidation of 3-phosphoglyceraldehyde to 1,3-bisphosphoglycerate is reoxidized during d-octopine formation. In some scallops d-octopine starts to accumulate during prolonged, strong muscular work, whereas in other species d-octopine formation commences towards the end of swimming and continues to rise during subsequent recovery. The activity of OcDH is regulated by a mandatory, consecutive mode of substrate binding in the order NADH, l-arginine and pyruvate, as demonstrated by isothermal titration calorimetry. The first regulatory step in the forward reaction comprises the binding of NADH to OcDH with a dissociation constant K(d) of 0.014±0.006 mmol l(-1), which reflects a high affinity and tight association of the apoenzyme with the co-substrate. In the reverse direction, NAD(+) binds first with a K(d) of 0.20±0.004 mmol l(-1) followed by d-octopine. The binary OcDH-NADH complex associates with l-arginine with a K(d) of 5.5±0.05 mmol l(-1). Only this ternary complex combines with pyruvate, with an estimated K(d) of approximately 0.8 mmol l(-1) as deduced from pyruvate concentrations determined in the muscle of exhausted scallops. At tissue concentrations of pyruvate between 0.5 and 1.2 mmol l(-1) in the valve adductor muscle of fatigued P. maximus, binding of pyruvate to OcDH plays the most decisive role in initiating OcDH activity and, therefore, in controlling the onset of d-octopine formation.