Dinoflagellate Symbiodinium Sp Research Articles

Isolation and characterization of a tandem-repeated cysteine protease from the symbiotic dinoflagellate Symbiodinium sp. KB8.

A cysteine protease belonging to peptidase C1A superfamily from the eukaryotic, symbiotic dinoflagellate, Symbiodinium sp. strain KB8, was characterized. The protease was purified to near homogeneity (566-fold) by (NH4)2SO4 fractionation, ultrafiltration, and column chromatography using a fluorescent peptide, butyloxycarbonyl-Val-Leu-Lys-4-methylcoumaryl-7-amide (Boc-VLK-MCA), as a substrate for assay purposes. The enzyme was termed VLKP (VLK protease), and its activity was strongly inhibited by cysteine protease inhibitors and activated by reducing agents. Based on the results for the amino acid sequence determined by liquid chromatography–coupled tandem mass spectrometry, a cDNA encoding VLKP was synthesized. VLKP was classified into the peptidase C1A superfamily of cysteine proteases (C1AP). The predicted amino acid sequence of VLKP indicated a tandem array of highly conserved precursors of C1AP with a molecular mass of approximately 71 kDa. The results of gel-filtration chromatography and SDS-PAGE suggested that VLKP exists as a monomer of 31–32 kDa, indicating that the tandem array is likely divided into two mass-equivalent halves that undergo equivalent posttranslational modifications. The VLKP precursor contains an inhibitor prodomain that might become activated after acidic autoprocessing at approximately pH 4. Both purified and recombinant VLKPs had a similar substrate specificity and kinetic parameters for common C1AP substrates. Most C1APs reside in acidic organelles such as the vacuole and lysosomes, and indeed VLKP was most active at pH 4.5. Since VLKP exhibited maximum activity during the late logarithmic growth phase, these attributes suggest that, VLKP is involved in the metabolism of proteins in acidic organelles.

Symbiodinolactone A, a new 12-membered macrolide from symbiotic marine dinoflagellate Symbiodinium sp.

A new 12-membered macrolide, symbiodinolactone A, was isolated from the culture broth of symbiotic marine dinoflagellate Symbiodinium sp. The gross structure of symbiodinolactone A was elucidated by spectroscopic analyses, and the relative configuration was elucidated by the J-based configuration analysis and density-functional theory calculations. Symbiodinolactone A is the new 12-membered macrolide possessing an E double bond between C-4 and C-5, a branched methyl group at C-7, and a 1,2,3-trihydroxybutyl group at C-11. Symbiodinolactone A is the first usual size macrolide and the first non-nitrogen-containing macrolide from dinoflagellate Symbiodinium sp. Symbiodinolactone A might be generated by the unexplained dinoflagellate polyketide biosynthetic machinery.

Rubisco Expression in the Dinoflagellate Symbiodinium sp. Is Influenced by Both Photoperiod and Endosymbiotic Lifestyle

Although the importance of anthozoan-dinoflagellate (genus Symbiodinium) endosymbioses in the establishment of coral reef ecosystems is evident, little is known about the molecular regulation of photosynthesis in the intra-gastrodermal symbiont communities, particularly with respect to the rate-limiting Calvin cycle enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco). In this study, we analyzed rubisco mRNA (rbcL) and protein (RBCL) concentrations over the diel cycle in both cultured and endosymbiotic Symbiodinium samples. In the former, rbcL expression increased upon illumination and decreased during the dark, a pattern that was upheld under continual dark incubation. A different trend in rbcL expression was observed in endosymbiotic Symbiodinium residing within sea anemone (Aiptasia pulchella) tissues, in which illumination gradually led to decreased rbcL mRNA expression. Unexpectedly, RBCL protein expression did not vary over time within anemone tissues, and in neither cultured nor endosymbiotic samples was a correlation between gene and protein expression documented. It appears, then, that photoperiod, lifestyle, and posttranscriptional regulation are all important drivers of RBCL expression in this ecologically important dinoflagellate.

構造解明を指向したポリオール天然物シンビオジノライドの合成研究

Symbiodinolide is a polyol marine natural product isolated from dinoflagellate Symbiodinium sp. This natural product exhibits voltage-dependent N-type Ca2+ channel-opening activity at 7 nM and COX-1 inhibition activity at 2 µM. The planar structure of symbiodinolide was elucidated by the detailed 2D NMR analysis. However, the huge and complex molecular structure of this molecule featuring a molecular weight of 2,860 and the presence of 61 chiral centers has hampered the complete stereochemical determination. Herein, we report our synthetic study of symbiodinolide toward the structural elucidation.

Combined Amplicon Pyrosequencing Assays Reveal Presence of the Apicomplexan “type-N” (cf. Gemmocystis cylindrus) and Chromera velia on the Great Barrier Reef, Australia

BackgroundThe coral is predominantly composed of the metabolically dependent coral host and the photosynthetic dinoflagellate Symbiodinium sp. The system as a whole interacts with symbiotic eukaryotes, bacteria and viruses. Gemmocystiscylindrus (cf. “type-N” symbiont) belonging to the obligatory parasitic phylum Apicomplexa (Alveolata) is ubiquitous in the Caribbean coral, but its presence in the Great Barrier Reef coral has yet to be documented. Approaches allowing identification of the healthy community from the pathogenic or saprobic organisms are needed for sustainable coral reef monitoring.Methods & Principal FindingsWe investigated the diversity of eukaryotes associated with a common reef-building corals from the southern Great Barrier Reef. We used three tag encoded 454 amplicon pyrosequencing assays targeting eukaryote small-subunit rRNA gene to demonstrate the presence of the apicomplexan type-N and a photosynthetic sister species to Apicomplexa - Chromera velia . Amplicon pyrosequencing revealed presence of the small-subunit rRNA genes of known eukaryotic pathogens ( Cryptosporidium and Cryptococcus ). We therefore conducted bacterial tag encoded amplicon pyrosequencing assay for small-subunit rRNA gene to support effluent exposure of the coral. Bacteria of faecal origin (Enterobacteriales) formed 41% of total sequences in contrast to 0-2% of the coral-associated bacterial communities with and without C . velia , respectively.SignificanceThis is the first time apicomplexan type-N has been detected in the Great Barrier Reef. Eukaryote tag encoded amplicon pyrosequencing assays demonstrate presence of apicomplexan type-N and C. Velia in total coral DNA. The data highlight the need for combined approaches for eukaryotic diversity studies coupled with bacterial community assessment to achieve a more realistic goals of defining the holobiont community and assessing coral disease. With increasing evidence of Apicomplexa in coral reef environments, it is important not only to understand the evolution of these organisms but also identify their potential as pathogens.

Symbiospirols: Novel Long Carbon-Chain Compounds Isolated from Symbiotic Marine Dinoflagellate Symbiodinium sp.

Symbiospirols A, B, and C, long carbon-chain compounds with a molecular weight of 1207 were isolated from the cultured symbiotic dinoflagellate Symbiodinium sp. Their planar structures and partial relative stereochemistries were elucidated based on 1D and 2D NMR spectra and a degradation reaction. Symbiospirols consisted of a C67-linear chain with a beta,beta'-dihydroxyl ketone moiety, eight hydroxyl groups, one tetrahydropyran ring, and two 1,6-dioxaspiro[4,4]nonane rings. Symbiospirol A had an inhibitory effect against l-phosphatidylserine-induced PKC activation.

Symbiodinolide, a Novel Polyol Macrolide that Activates N‐Type Ca2+ Channel, from the Symbiotic Marine Dinoflagellate Symbiodinium sp.

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Synthesis of (±)-Symbioimine

Symbioimine is isolated from the marine dinoflagellate Symbiodinium sp. A 12-step synthesis is reported in which the tricyclic core is constructed using an intramolecular Diels-Alder reaction.

Symbiodinolide, a novel polyol macrolide that activates N-type Ca 2+ channel, from the symbiotic marine dinoflagellate Symbiodinium sp.

A 62-membered novel polyol macrolide with a molecular weight of 2859 mu, symbiodinolide, was isolated from the symbiotic dinoflagellate Symbiodinium sp. Symbiodinolide exhibited a potent voltage-dependent N-type Ca 2+ channel-opening activity at 7 nM and immediately ruptured the tissue surface of the acoel flatworm Amphiscolops sp. at 2.5 μM. The planar structure of symbiodinolide was elucidated by spectroscopic analysis and chemical degradations including hydrolysis and ethenolysis using the second-generation Grubbs' catalyst. Symbiodinolide was found to be a structural congener of zooxanthellatoxins. The relative stereochemistries of C26–C32, C44–C51, and C64–C66 parts, and the absolute stereochemistries of C69–73, C83–C103, and C3′–C18′ parts in symbiodinolide were established.

Symbioimine and Neosymbioimine, Amphoteric Iminium Metabolites from the Symbiotic Marine Dinoflagellate Symbiodinium sp.

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Symbioimine and neosymbioimine, amphoteric iminium metabolites from the symbiotic marine dinoflagellate Symbiodinium sp.

Two amphoteric iminium metabolites, symbioimine ( 1) and neosymbioimine ( 2), were isolated from a cultivated symbiotic marine dinoflagellate Symbiodinium sp. Compounds 1 and 2 have a characteristic 6,6,6-tricyclic iminium ring structure and an aryl sulfate moiety. The plausible biogenetic pathway of 1 and 2 can be explained by an intramolecular Diels–Alder reaction followed by imine cyclization. Symbioimine ( 1) inhibited the differentiation of RAW264 cells into osteoclasts (EC 50 = 44 μM), and significantly inhibited cyclooxygenase-2 activity at 10 μM. Thus, symbioimine is a potent anti-resorptive and anti-inflammatory drug.

Symbioimine exhibiting inhibitory effect of osteoclast differentiation, from the symbiotic marine dinoflagellate Symbiodinium sp.

An amphoteric iminium metabolite, symbioimine (1), was isolated from a cultivated symbiotic marine dinoflagellate Symbiodinium sp. Its structure was deduced by spectroscopic analysis and X-ray crystallographic analysis. Symbioimine (1) has a characteristic 6,6,6-tricyclic iminium ring structure and an aryl sulfate moiety. The plausible biogenetic pathway of 1 can be explained by an intramolecular Diels-Alder reaction followed by imine cyclization. Symbioimine (1) inhibited the differentiation of RAW264 cells into osteoclasts, whereas its cell viability was not affected. Thus, symbioimine (1) is an antiresorptive drug candidate for the prevention and treatment of osteoporosis in postmenopausal women.

Pseudopterosin Biosynthesis in Symbiodinium sp., the Dinoflagellate Symbiont of Pseudopterogorgia elisabethae

Investigations are reported that identify the biosynthetic source and origins of the pseudopterosins, pharmacologically important diterpene glycosides, in the gorgonian coral Pseudopterogorgia elisabethae. We report here the isolation of physiologically significant levels of endogenous pseudopterosins A, B, C, and D from purified symbionts identified as the dinoflagellate Symbiodinium sp. Biosynthetic studies in photosynthesizing symbiont isolates utilizing 14C-labeled inorganic carbon and the tritiated intermediate geranylgeranyl diphosphate yielded radiochemically pure pseudopterosins A through D and the first committed intermediate, elisabethatriene. The 14CO 2 uptake into the pseudopterosin pathway occurred at low levels compared to the 14CO 2 uptake into total lipids, suggesting a large reserve pool of the compounds. These results reveal for the first time that pseudopterosin biosynthesis is occurring within the algal symbiont and suggests the physiological implications of this biosynthesis.

Immunolocalization and distribution of form ii rubisco in the pyrenoid and chloroplast stroma of amphidinium carterae and form i rubisco in the symbiont‐derived plastids of peridinium foliaceum (dinophyceae)

Chloroplasts of peridinin‐containing dinoflagellates have recently been shown to contain Form II Rubisco, which consists of large subunits only and is coded by nuclear genes. We have used immunoelectron microscopy to determine the distribution of Form II and Form I Rubisco in dinoflagellates. In sections of Amphidinium carterae Hulburt, the pyrenoid was intensely labeled and the rest of the chloroplast moderately labeled by antisera to Form II Rubisco from the purple non‐sulfur bacterium Rhodospirillum rubrum and the symbiotic dinoflagellate Symbiodinium sp. No labeling was observed when sections were exposed to antiserum against Form I Rubisco of the haptophyte alga Isochrysis galbana. In contrast, cell sections of the dinoflagellate Peridinium foliaceum (Stein) Biecheler, whose chloroplasts belong to a diatom endosymbiont, showed no labeling with the two antisera against Form II Rubisco, but heavy pyrenoid labeling was present after treatment with antiserum against Form I Rubisco of I. galbana. The same immunolabeling results were obtained with the free‐living diatom Phaeodactylum tricornutum Bohlin. Volumetric analysis of the distribution of Form II Rubisco in the chloroplast of A. carterae showed that, in cells grown under moderate photon irradiance, 72.9% of the plastid's Rubisco was localized in the pyrenoid, whereas in cells grown under low irradiance only 37.0% of the Rubisco was found in the pyrenoid. This light‐induced concentration of Rubisco in the pyrenoid suggests that a CO2–concentrating mechanism may elevate CO2 within the pyrenoid, favoring the efficient fixation of CO2 by pyrenoid Rubisco.

Synthetic studies on the common C25 long chain acid portion of zooxanthellatoxins from a symbiotic dinoflagellate Symbiodinium sp.

The absolute stereochemistry of a portion from C11 to C18 of the common C25 long chain acid part of zooxanthellatoxins was determined by synthesizing the degradation product 2a to be 11 S, 12 R, 13 S, 14 R, 15 R, 17 R, and 18 R.

Nucleotide sequence of a cDNA clone encoding the precursor of the peridinin-chlorophyll a-binding protein from the dinoflagellate Symbiodinium sp.

mRNA from the dinoflagellate Symbiodinium sp. isolated from the staghorn coral Acropora formosa was used for the construction of cDNA libraries. A cDNA clone was identified which encoded the precursor of peridinin-chlorophyll a-binding protein (PCP), including a 52 amino acid transit peptide and the 313 amino acid mature protein. The deduced amino acid sequence clearly contains an internal duplication, implying that amongst dinoflagellates the M(r) 35,000 form of PCP has arisen by duplication and fusion of genes encoding the M(r) 15,000 form. This is the first reported sequence of a dinoflagellate light-harvesting protein. The anatomy of the mature protein and the transit peptide are discussed.

Immunological evidence for a ribulose bisphosphate carboxylase-like protein in the symbiotic dinoflagellate Symbiodinium sp.

1. Summary Whereas previously there has been no convincing evidence for ribulose bisphosphate carboxylase in dinoflagellates, a strong and highly specific reaction was observed when antibodies to the denatured large subunit of the (silver beet) protein were used to probe Western blots of whole soluble fractions of various Symbiodinium isolates. No reaction was observed using extracts from Symbiodinium isolated from a host which had been maintained under low light intensity. The results imply extensive sequence homology between the large subunit of ribulose bisphosphate carboxylase and a dinoflagellate protein of M , approximately 35 000.

Immunological evidence for a ribulose bisphosphate carboxylase-like protein in the symbiotic dinoflagellate Symbiodinium sp.

Immunological evidence for a ribulose bisphosphate carboxylase-like protein in the symbiotic dinoflagellate Symbiodinium sp.

Symbioramide, a novel Ca2+-ATPase activator from the cultured dinoflagellate Symbiodinium sp.

A novel sphingosine derivative, symbioramide, has been isolated from the laboratory-cultured dinoflagellate Symbiodinium sp. as a sarcoplasmic reticulum (SR)Ca2+-ATPase activator, and its structure elucidated to be 1 on the basis of spectral and chemical means.