Seven UV-absorbing, mycosporine-like amino acids (MAA) have been identified in organs of Thelenota ananas from the Great Barrier Reef. Concentrations of MAA are greatest in the external layer of the dorsal body wall and very low to undetectable in most internal organs, except for gut tissues. Four of the compounds are present in coral sand from the habitat of T. ananas, at concentrations 1000-fold lower than in tissues. Relative to habitat sand, sand in the gut is enriched about threefold in its content of these compounds, which appear to be removed from the gut contents prior to their defecation. Gut contents and feces contain asterina–330, a compound absent from habitat sediment but which is the single most concentrated MAA in the tissues. Mycosporine-glycine is the next most concentrated MAA in epidermis, and is absent from habitat sand, gut contents and feces. The organ distribution of MAA supports the adaptive interpretation that MAA provide protection from solar UV radiation in exposed epidermal tissues. The high concentrations of MAA in the gonads of other holothurian species are consistent with this interpretation, since broadcast gametes would also require protection from UV. Metazoans are unlikely to synthesize MAA de novo; it is uncertain whether MAA are obtained directly from ingested materials, but the presence in the tissues of certain MAA absent from the diet suggests secondary metabolic modification of dietary compounds. Enrichment of gut contents with MAA relative to habitat sand may indicate selective feeding on algal-rich particles having a high MAA content, or synthesis of MAA by microbes in the gut. Supported by AIMS, and by National Geographic Society grant 3883-88 and NSF DCB–8509487 (Regulatory Biology) to JMS.

This study aimed to determine the antioxidant and anticancer activity of methanol extract of Alikokop leaves (Dischidia nythesiana). The extraction method used is maceration extraction using methanol solvent. The antioxidant activity test was carried out using the DPPH (1,1-diphenyl-2-picrylhydrazyl) free radical deterrent method and the anticancer activity assay was carried out using the MTT (Microculture Tetrazolium) method using P-388 cancer cells. From this study, the antioxidant test results showed that the methanol extract of Alikokop leaves had antioxidant activity with moderate concentration, with IC50 of 67.136 μg / mL, and the results of anticancer tests showed that methanol extract of Alikokop leaves had anticancer activity on leukemia cells P388 with concentration inhibition values medium, with IC50 of 24.88 μg / mL. The methanol extract of Alikokop leaves has the potential as an antioxidant and anticancer leukemia.

Mycosporine-like amino acids (MAAs) are UVR-absorbing metabolites typically produced by cyanobacteria and marine algae, but their properties are not limited to direct sun screening protection. Herein, we examine the antioxidant activities of porphyra-334 and shinorine and demonstrate that these MAAs are prospective activators of the cytoprotective Keap1-Nrf2 pathway. The ability of porphyra-334 and shinorine to bind with Keap1 was determined using fluorescence polarization (FP) and thermal shift assays to detect Keap1 receptor antagonism. Concomitantly, the ability of porphyra-334 and shinorine to dissociate Nrf2 from Keap1 was confirmed also by measurement of increased mRNA expression of Nrf2 targeted genes encoding oxidative stress defense proteins in primary skin fibroblasts prior and post UVR exposure. Surprisingly, enhanced transcriptional regulation was only promoted by MAAs in cells after exposure to UVR-induced oxidative stress. Furthermore, the in-vitro antioxidant activities of porphyra-334 and shinorine determined by the DPPH free-radical quenching assay were low in comparison to ascorbic acid. However, their antioxidant capacity determined by the ORAC assay to quench free radicals via hydrogen atom transfer is substantial. Hence, the dual nature of MAAs to provide antioxidant protection may offer a prospective chemotherapeutic strategy to prevent or retard the progression of multiple degenerative disorders of ageing.

Oxidative stress is a contributing factor in the progression of numerous pathological conditions including neurodegeneration, cancer and ageing. The Keap1-Nrf2 pathway is a master regulator of oxidative stress: reactive oxygen species are sensed by Keap1 to release Nrf2 for transcriptional activation of protective genes controlled by the nuclear antioxidant response element. We have identified homologs of Nrf2 in early eukaryotes and used virtual screens to predict natural products able to activate Nrf2 by competitive inhibition of Keap1-Nrf2 binding. Mycosporine-like amino acids (MAAs) are water-soluble metabolites produced by taxonomically diverse organisms, particularly marine algae and seaweeds. These compounds absorb UV radiation – thus acting as “primary sunscreens” – and reported also to protect against oxidative damage. We have tested the MAAs, porphyra-334, shinorine, and palythine for in-vitro antioxidant activity using the DPPH free-radical quenching assay and report also their ability to activate the Keap1-Nrf2 pathway using fluorescence polarization and thermal shift assays to detect Keap1 receptor antagonism. Our results demonstrate that shinorine and porphyra-334 are competitive inhibitors of Keap1-Nrf2 binding having potential to protect against UVR via sunscreen absorption and transcriptional activation of endogenous defenses against UV-induced oxidative damage.

In mammals, the master transcription regulator of antioxidant defences is provided by the Nrf2 protein. Phylogenetic analyses of Nrf2 sequences are used here to derive a molecular clock that manifests persuasive evidence that Nrf2 orthologues emerged and then diverged, at two time points that correlate with well-established geochemical and palaeobiological chronologies during progression of the ‘Great Oxygenation Event’. We demonstrate that orthologues of Nrf2 first appeared in fungi around 1.5 Ga during the Paleoproterozoic when photosynthetic oxygen was being absorbed into the oceans. A subsequent significant divergence in Nrf2 is seen during the split between fungi and the Metazoa approximately 1.0–1.2 Ga, at a time when oceanic ventilation released free oxygen to the atmosphere, but with most being absorbed by methane oxidation and oxidative weathering of land surfaces until approximately 800 Ma. Atmospheric oxygen levels thereafter accumulated giving rise to metazoan success known as the Cambrian explosion commencing at ~541 Ma. Atmospheric O2 levels then rose in the mid Paleozoic (359–252 Ma) and Nrf2 diverged once again at the division between mammals and non-mammalian vertebrates during the Permian-Triassic boundary (~252 Ma). Understanding Nrf2 evolution as an effective antioxidant response may have repercussions for improved human health.

Shipboard experiments were each performed over a 2 day period to examine the proteomic response of the symbiotic coral Acropora microphthalma exposed to acute conditions of high temperature/low light or high light/low temperature stress. During these treatments, corals had noticeably bleached. The photosynthetic performance of residual algal endosymbionts was severely impaired but showed signs of recovery in both treatments by the end of the second day. Changes in the coral proteome were determined daily and, using recently available annotated genome sequences, the individual contributions of the coral host and algal endosymbionts could be extracted from these data. Quantitative changes in proteins relevant to redox state and calcium metabolism are presented. Notably, expression of common antioxidant proteins was not detected from the coral host but present in the algal endosymbiont proteome. Possible roles for elevated carbonic anhydrase in the coral host are considered: to restore intracellular pH diminished by loss of photosynthetic activity, to indirectly limit intracellular calcium influx linked with enhanced calmodulin expression to impede late-stage symbiont exocytosis, or to enhance inorganic carbon transport to improve the photosynthetic performance of algal symbionts that remain in hospite. Protein effectors of calcium-dependent exocytosis were present in both symbiotic partners. No caspase-family proteins associated with host cell apoptosis, with exception of the autophagy chaperone HSP70, were detected, suggesting that algal loss and photosynthetic dysfunction under these experimental conditions were not due to host-mediated phytosymbiont destruction. Instead, bleaching occurred by symbiont exocytosis and loss of light-harvesting pigments of algae that remain in hospite. These proteomic data are, therefore, consistent with our premise that coral endosymbionts can mediate their own retention or departure from the coral host, which may manifest as "symbiont shuffling" of Symbiodinium clades in response to environmental stress.

An essential requirement for the evolution of early eukaryotic life was the development of effective means to protect against metabolic oxidative stress and exposure to environmental toxicants. In present-day mammals, the master transcription factor Nrf2 regulates basal level homeostasis and inducible expression of numerous detoxifying and antioxidant genes. To examine early evolution of the Keap1–Nrf2 pathway, we present bioinformatics analyses of distant homology of mammalian Keap1 and Nrf2 proteins across the Kingdoms of Life. Software written for this analysis is made freely available on-line. Furthermore, utilizing protein modeling and virtual screening methods, we demonstrate potential for Nrf2 activation by competitive inhibition of its binding to Keap1, specifically by UV-protective fungal mycosporines and marine mycosporine-like amino acids (MAAs). We contend that coevolution of Nrf2-activating secondary metabolites by fungi and other extant microbiota may provide prospective compound leads for the design of new therapeutics to target activation of the human Keap1–Nrf2 pathway for treating degenerative diseases of ageing.

Coenzyme Q (CoQ; ubiquinone) and plastoquinone (PQ) are metabolic electron carriers that, in their reduced state, are powerful antioxidants for cellular protection against oxidative damage. Although damage resulting from generation of reactive oxygen species (ROS) is strongly implicated in the initiation of symbiotic dysfunction that leads to coral bleaching, very little is known about the redox state of these two prenylquinone pools during the stress response. Here we describe a quantitative liquid chromatography–mass spectrometry (LC–MS) method that permits simultaneous measurement of the physiological redox state of both CoQ and PQ in whole corals. The application of this method indicates that the CoQ and PQ pools in the coral–Symbiodinium symbiosis are maintained predominantly in their reduced (antioxidant) forms, and it is the coral CoQ redox state that is most affected by acute thermal stress.

The parent core structure of mycosporine-like amino acids (MAAs) is 4-deoxygadusol, which, in cyanobacteria, is derived from conversion of the pentose phosphate pathway intermediate sedoheptulose 7-phosphate by the enzymes 2-epi-5-epivaliolone synthase (EVS) and O-methyltransferase (OMT). Yet, deletion of the EVS gene from Anabaena variabilis ATCC 29413 was shown to have little effect on MAA production, thus suggesting that its biosynthesis is not exclusive to the pentose phosphate pathway. Herein, we report how, using pathway-specific inhibitors, we demonstrated unequivocally that MAA biosynthesis occurs also via the shikimate pathway. In addition, complete in-frame gene deletion of the OMT gene from A. variabilis ATCC 29413 reveals that, although biochemically distinct, the pentose phosphate and shikimate pathways are inextricably linked to MAA biosynthesis in this cyanobacterium. Furthermore, proteomic data reveal that the shikimate pathway is the predominate route for UV-induced MAA biosynthesis.

Surprisingly little is known of the toxic arsenal of cnidarian nematocysts compared to other venomous animals. Here we investigate the toxins of nematocysts isolated from the jellyfish Olindias sambaquiensis. A total of 29 unique ms/ms events were annotated as potential toxins homologous to the toxic proteins from diverse animal phyla, including cone-snails, snakes, spiders, scorpions, wasp, bee, parasitic worm and other Cnidaria. Biological activities of these potential toxins include cytolysins, neurotoxins, phospholipases and toxic peptidases. The presence of several toxic enzymes is intriguing, such as sphingomyelin phosphodiesterase B (SMase B) that has only been described in certain spider venoms, and a prepro-haystatin P-IIId snake venom metalloproteinase (SVMP) that activates coagulation factor X, which is very rare even in snake venoms. Our annotation reveals sequence orthologs to many representatives of the most important superfamilies of peptide venoms suggesting that their origins in higher organisms arise from deep eumetazoan innovations. Accordingly, cnidarian venoms may possess unique biological properties that might generate new leads in the discovery of novel pharmacologically active drugs.

Background Contemporary coral reef research has firmly established that a genomic approach is urgently needed to better understand the effects of anthropogenic environmental stress and global climate change on coral holobiont interactions. Here we present KEGG orthology-based annotation of the complete genome sequence of the scleractinian coral Acropora digitifera and provide the first comprehensive view of the genome of a reef-building coral by applying advanced bioinformatics. Description Sequences from the KEGG database of protein function were used to construct hidden Markov models. These models were used to search the predicted proteome of A. digitifera to establish complete genomic annotation. The annotated dataset is published in ZoophyteBase, an open access format with different options for searching the data. A particularly useful feature is the ability to use a Google-like search engine that links query words to protein attributes. We present features of the annotation that underpin the molecular structure of key processes of coral physiology that include (1) regulatory proteins of symbiosis, (2) planula and early developmental proteins, (3) neural messengers, receptors and sensory proteins, (4) calcification and Ca 2+ -signalling proteins, (5) plant-derived proteins, (6) proteins of nitrogen metabolism, (7) DNA repair proteins, (8) stress response proteins, (9) antioxidant and redox-protective proteins, (10) proteins of cellular apoptosis, (11) microbial symbioses and pathogenicity proteins, (12) proteins of viral pathogenicity, (13) toxins and venom, (14) proteins of the chemical defensome and (15) coral epigenetics. Conclusions We advocate that providing annotation in an open-access searchable database available to the public domain will give an unprecedented foundation to interrogate the fundamental molecular structure and interactions of coral symbiosis and allow critical questions to be addressed at the genomic level based on combined aspects of evolutionary, developmental, metabolic, and environmental perspectives.

The cyclase 2-epi-5-epi-valiolone synthase (EVS) is reported to be a key enzyme for biosynthesis of the mycosporine-like amino acid shinorine in the cyanobacterium Anabaena variabilis ATCC 29413. Subsequently, we demonstrated that an in-frame complete deletion of the EVS gene had little effect on in vivo production of shinorine. Complete segregation of the EVS gene deletion mutant proved difficult and was achieved only when the mutant was grown in the dark and in a medium supplemented with fructose. The segregated mutant showed a striking colour change from native blue-green to pale yellow-green, corresponding to substantial loss of the photosynthetic pigment phycocyanin, as evinced by combinations of absorbance and emission spectra. Transcriptional analysis of the mutant grown in the presence of fructose under dark or light conditions revealed downregulation of the cpcA gene that encodes the alpha subunit of phycocyanin, whereas the gene encoding nblA, a protease chaperone essential for phycobilisome degradation, was not expressed. We propose that the substrate of EVS (sedoheptulose 7-phosphate) or possibly lack of its EVS-downstream products, represses transcription of cpcA to exert a hitherto unknown control over photosynthesis in this cyanobacterium. The significance of this finding is enhanced by phylogenetic analyses revealing horizontal gene transfer of the EVS gene of cyanobacteria to fungi and dinoflagellates. It is also conceivable that the EVS gene has been transferred from dinoflagellates, as evident in the host genome of symbiotic corals. A role of EVS in regulating sedoheptulose 7-phosphate concentrations in the photophysiology of coral symbiosis is yet to be determined.

The photosynthetic bleaching response of Acropora microphthalma, Acropora formosa, Stylophora pistillata and Pocillopora damicornis were compared under empirical conditions of environmental stress. Coral specimens were collected at Davies Reef located in the central region of the GBR from a depth of 10-15m and were exposed to natural and shaded light under temperature-controlled conditions for 48 h at a depth of 40 cm held within a shipboard, light-exposed tank. After 2 days of stress (high light/low temperature, high light/high temperature, and low light/high temperature), the acroporids were severely bleached whereas the Stylophora and Pocillopora specimens had retained some pigmentation. Light-adapted PSII Yield values declined to 0.1-0.2 units at midday in all coral species but had increased by partial recovery during the afternoon periods of exposure. From these 2-day experiments, the rate of PSII Yield recovery (Yr), calculated as the increment of light-adapted PSII Yield that had recovered from midday to dusk, were compared. Results of Yr determinations showed that A. formosa is a thermally sensitive species having reduced Yr values under both light conditions. A. microphthalma and S. pistillata were more thermally tolerant but light sensitive with greater Yr values under low light than under high light exposure. Pocillopora was least sensitive to bleaching and showed attributes of light and thermal tolerance with up to six times greater Yr values than the other coral specimens examined under identical conditions. The bleaching response of these corals under different stress conditions will be discussed in context with the photosynthetic response of their symbionts measured in hospite.

Route of the sun block: according to empirical evidence, sun-screening mycosporine-like amino acids (MAAs) in Eukarya originate from the shikimic acid pathway, whereas in cyanobacteria, biosynthesis of the MAA shinorine reportedly occurs through the pentose phosphate pathway. However, gene deletion shows that the cyanobacterium Anabaena variabilis ATCC 29143 does not biosynthesise shinorine exclusively by this route.

This study examines the response of Symbiodinium sp. endosymbionts from the coral Stylophora pistillata to moderate levels of thermal "bleaching" stress, with and without trace metal limitation. Using quantitative high throughput proteomics, we identified 8098 MS/MS events relating to individual peptides from the endosymbiont-enriched fraction, including 109 peptides meeting stringent criteria for quantification, of which only 26 showed significant change in our experimental treatments; 12 of 26 increased expression in response to thermal stress with little difference affected by iron limitation. Surprisingly, there were no significant increases in antioxidant or heat stress proteins; those induced to higher expression were generally involved in protein biosynthesis. An outstanding exception was a massive 114-fold increase of a viral replication protein indicating that thermal stress may substantially increase viral load and thereby contribute to the etiology of coral bleaching and disease. In the absence of a sequenced genome for Symbiodinium or other photosymbiotic dinoflagellate, this proteome reveals a plethora of proteins potentially involved in microbial-host interactions. This includes photosystem proteins, DNA repair enzymes, antioxidant enzymes, metabolic redox enzymes, heat shock proteins, globin hemoproteins, proteins of nitrogen metabolism, and a wide range of viral proteins associated with these endosymbiont-enriched samples. Also present were 21 unusual peptide/protein toxins thought to originate from either microbial consorts or from contamination by coral nematocysts. Of particular interest are the proteins of apoptosis, vesicular transport, and endo/exocytosis, which are discussed in context of the cellular processes of coral bleaching. Notably, the protein complement provides evidence that, rather than being expelled by the host, stressed endosymbionts may mediate their own departure.

In this communication, we introduce a novel biomarker of aquatic contamination based on the xenobiotic-induced response of the hepatic coenzyme Q (CoQ) redox balance of fishes to polycyclic aromatic hydrocarbons (PAHs). The method is demonstrated by comparing changes in the liver CoQ redox balance with that measured using the CYP1A-based, 7-ethoxyresofurin-O-deethylase activity assay, on administration of benzo[a]pyrene (BaP) and β-naphthoflavone (BNF) to Barramundi ( Lates calcarifer ). Both assays showed comparable dose-dependent effects in fish treated with BaP or BNF. Perturbation in the constitutive hepatic CoQ redox balance of fishes may thus provide a simple biomarker of aquatic PAH contamination.

Exposing the coral Stylophora pistillata to seawater depleted in available iron by complexation with the strong chelator desferrioxamine B reduces the photosynthetic efficiency and alters the pigment composition in its symbiotic algae at high temperatures. Similar effects of iron limitation are known for free‐living algae, but this is the first demonstration of low‐iron stress in a dinoflagellate living endosymbiotically. Maintaining corals at elevated temperature (30°C and 31°C) under diminished iron availability leads to reduced maximum quantum yields (Fv:Fm) of photosystem II (PSII), specifically on brightly illuminated surfaces of the coral. This reduction in maximum quantum yield is due in part to increased photoprotection, indicated by an increase in the photoprotective xanthophyll diatoxanthin, which promotes nonphotochemical quenching of excess light energy to restrict oxidative damage under conditions that impair photosynthetic electron transfer. However, photophysiological changes associated with the lowered maximum quantum yield did not prevent photodamage to PSII under the combined effects of elevated temperature and low iron availability, as shown by the decrease in maximum (Fm) that was not accompanied by significant change in the minimum (Fo) fluorescence yield. All of the foregoing is consistent with the potential of iron limitation to contribute to the underlying conditions by which thermal stress evokes the physiological response by corals that culminates in the symbiotic dysfunction of natural bleaching.