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Kubanek, Julia

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https://hdl.handle.net/1853/71428
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School of Biological Sciences
School established in 2016 with the merger of the Schools of Applied Physiology and Biology
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Warding Off Disease on Coral Reefs: Antimicrobial Chemical Cues and their Future in Drug Discovery

2012-05-08 , Kubanek, Julia

Unlike many animals, seaweeds do not possess adaptive immune systems to ward off disease. In many cases, they produce small molecules – natural antibiotics – that prevent colonization or infection by pathogens. We have found that seaweeds produce unusual secondary metabolites against pathogen attack, including complex isoprenoid-and shikimate-derived macrolides not seen in any other organisms. Surprisingly, these natural antifungals are not distributed evenly across algal surfaces; instead they are concentrated at discrete surface patches where they provide bursts of protection at sites that may be especially vulnerable to infection due to prior wounding. Working with the Fernandez lab at Georgia Tech, we applied surface imaging mass spectrometry to intact algal surfaces to show that antifungal defenses are heterogeneously distributed, with compound concentrations high enough at localized patches to block infection. This patchy distribution may represent an optimal defense strategy, in which the most vulnerable parts of the alga are best defended. Chemical defenses of seaweeds also serve as valuable leads for pharmaceutical development. Members of one group of algal antifungal agents, the bromophycolides, exhibit potent in vitro and in vivo antimalarial activity with an unexpected mechanism of action, inhibiting growth of the malarial parasite Plasmodium falciparum at sub-micromolar concentrations. Using a molecular probe designed from the seaweed’s antifungal agent, we identified a major molecular target and drug binding mechanism within the malaria parasite that is helping guide our synthesis of novel analogs for future development.

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Genetic determinants of mate recognition in Brachionus manjavacas (Rotifera)

2009-09-09 , Snell, Terry W. , Shearer, Tonya L , Smith, Hilary A. , Kubanek, Julia , Gribble, Kristin E. , Welch , David B. Mark

Background: Mate choice is of central importance to most animals, influencing population structure, speciation, and ultimately the survival of a species. Mating behavior of male brachionid rotifers is triggered by the product of a chemosensory gene, a glycoprotein on the body surface of females called the mate recognition pheromone. The mate recognition pheromone has been biochemically characterized, but little was known about the gene(s). We describe the isolation and characterization of the mate recognition pheromone gene through protein purification, N-terminal amino acid sequence determination, identification of the mate recognition pheromone gene from a cDNA library, sequencing, and RNAi knockdown to confirm the functional role of the mate recognition pheromone gene in rotifer mating. Results: A 29 kD protein capable of eliciting rotifer male circling was isolated by high-performance liquid chromatography. Two transcript types containing the N-terminal sequence were identified in a cDNA library; further characterization by screening a genomic library and by polymerase chain reaction revealed two genes belonging to each type. Each gene begins with a signal peptide region followed by nearly perfect repeats of an 87 to 92 codon motif with no codons between repeats and the final motif prematurely terminated by the stop codon. The two Type A genes contain four and seven repeats and the two Type B genes contain three and five repeats, respectively. Only the Type B gene with three repeats encodes a peptide with a molecular weight of 29 kD. Each repeat of the Type B gene products contains three asparagines as potential sites for N-glycosylation; there are no asparagines in the Type A genes. RNAi with Type A double-stranded RNA did not result in less circling than in the phosphate-buffered saline control, but transfection with Type B double-stranded RNA significantly reduced male circling by 17%. The very low divergence between repeat units, even at synonymous positions, suggests that the repeats are kept nearly identical through a process of concerted evolution. Information-rich molecules like surface glycoproteins are well adapted for chemical communication and aquatic animals may have evolved signaling systems based on these compounds, whereas insects use cuticular hydrocarbons. Conclusion: Owing to its critical role in mating, the mate recognition pheromone gene will be a useful molecular marker for exploring the mechanisms and rates of selection and the evolution of reproductive isolation and speciation using rotifers as a model system. The phylogenetic variation in the mate recognition pheromone gene can now be studied in conjunction with the large amount of ecological and population genetic data being gathered for the Brachionus plicatilis species complex to understand better the evolutionary drivers of cryptic speciation.

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Chemical Cues in the Ocean

2006-04-25 , Kubanek, Julia

We are all at the mercy of molecules: hormones affect our sexual interests, toxins give us food poisoning, polluting chemicals give us asthma, and the smell of Krispy Kreme tugs on the steering wheel whenever we drive along Ponce de Leon Ave. Molecules play an even greater role in the lives of marine plants, animals, and microorganisms. On coral reefs and in the open ocean, chemicals affect who eats who, who settles down next to who, and who can fight off a potentially deadly infection. In lab and field experiments, Georgia Tech faculty and students are working to understand the role of natural chemical compounds in the survival and reproduction of marine organisms.

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A biochemical, genetic, and genomic investigation of the evolution and ecology of sexual reproduction

2010-11-17 , Snell, Terry W. , Kubanek, Julia

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Chemically mediated competition between microbes and animals: microbes as consumers in food webs

2006-11 , Burkepile, Deron E. , Parker, John D. , Woodson, Clifton Brock , Mills, Heath Jordan , Kubanek, Julia , Sobecky, Patricia A. , Hay, Mark E.

Microbes are known to affect ecosystems and communities as decomposers, pathogens, and mutualists. However, they also may function as classic consumers and competitors with animals if they chemically deter larger consumers from using rich food-falls such as carrion, fruits, and seeds that can represent critical windfalls to both microbes and animals. Microbes often use chemicals (i.e., antibiotics) to compete against other microbes. Thus using chemicals against larger competitors might be expected and could redirect significant energy subsidies from upper trophic levels to the detrital pathway. When we baited traps in a coastal marine ecosystem with fresh vs. microbe-laden fish carrion, fresh carrion attracted 2.6 times as many animals per trap as microbe-laden carrion. This resulted from fresh carrion being found more frequently and from attracting more animals when found. Microbe-laden carrion was four times more likely to be uncolonized by large consumers than was fresh carrion. In the lab, the most common animal found in our traps (the stone crab Menippe mercenaria) ate fresh carrion 2.4 times more frequently than microbe-laden carrion. Bacteria-removal experiments and feeding bioassays using organic extracts of microbe-laden carrion showed that bacteria produced noxious chemicals that deterred animal consumers. Thus bacteria compete with large animal scavengers by rendering carcasses chemically repugnant. Because food-fall resources such as carrion are major food subsidies in many ecosystems, chemically mediated competition between microbes and animals could be an important, common, but underappreciated interaction within many communities.

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Ambiguous role of phlorotannins as chemical defenses in the brown alga Fucus vesiculosus

2004-08-16 , Kubanek, Julia , Lester, Sarah E. , Fenical, William , Hay, Mark E.

Brown seaweeds (Fucales) produce phlorotannins that are often considered chemical defenses against herbivores. The many correlative and fewer direct tests conducted have shown effects of phlorotannins on herbivore feeding behavior to be variable. In an attempt to clarify the roles of phlorotannins versus other metabolites in defending brown algae, we conducted bioassay-guided fractionation of herbivore-deterrent extracts from the commonly studied brown alga Fucus vesiculosus. Feeding by the amphipods Ampithoe valida and A. longimana and the sea urchin Arbacia punctulata was suppressed by crude and water-soluble extracts of F. vesiculosus, but this deterrence was lost following storage or fractionation of the active, water-soluble extract. Phlorotannins in these extracts did not decompose in parallel with the loss of feeding deterrence. F. vesiculosus phlorotannins were fed to herbivores at 3 to 12× the isolated yield (or 4.2 to 16.8% of plant dry mass). No herbivore was deterred from feeding by concentrations of 3 or 6×, but A. valida (the only test herbivore that readily consumes F. vesiculosus in the field) was deterred at 12× isolated yield. When juvenile A. valida were raised on an artificial diet containing F. vesiculosus phlorotannins at 3× isolated yield, the phlorotannin-rich diet significantly enhanced, rather than reduced, amphipod survivorship and growth relative to an equivalent diet without phlorotannins. Females ovulated only on the phlorotannin-rich diet. Compounds other than phlorotannins appear to defend the F. vesiculosus populations we investigated, but we were unable to identify these unstable compounds.

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Biochemical Warfare on the Reef: The Role of Glutathione Transferases in Consumer Tolerance of Dietary Prostaglandins

2010-01-06 , Whalen, Kristen E. , Lane, Amy L. , Kubanek, Julia , Hahn, Mark E.

Background: Despite the profound variation among marine consumers in tolerance for allelochemically-rich foods, few studies have examined the biochemical adaptations underlying diet choice. Here we examine the role of glutathione Stransferases (GSTs) in the detoxification of dietary allelochemicals in the digestive gland of the predatory gastropod Cyphoma gibbosum, a generalist consumer of gorgonian corals. Controlled laboratory feeding experiments were used to investigate the influence of gorgonian diet on Cyphoma GST activity and isoform expression. Gorgonian extracts and semipurified fractions were also screened to identify inhibitors and possible substrates of Cyphoma GSTs. In addition, we investigated the inhibitory properties of prostaglandins (PGs) structurally similar to antipredatory PGs found in high concentrations in the Caribbean gorgonian Plexaura homomalla. Principal Findings: Cyphoma GST subunit composition was invariant and activity was constitutively high regardless of gorgonian diet. Bioassay-guided fractionation of gorgonian extracts revealed that moderately hydrophobic fractions from all eight gorgonian species examined contained putative GST substrates/inhibitors. LC-MS and NMR spectral analysis of the most inhibitory fraction from P. homomalla subsequently identified prostaglandin A2 (PGA2) as the dominant component. A similar screening of commercially available prostaglandins in series A, E, and F revealed that those prostaglandins most abundant in gorgonian tissues (e.g., PGA2) were also the most potent inhibitors. In vivo estimates of PGA2 concentration in digestive gland tissues calculated from snail grazing rates revealed that Cyphoma GSTs would be saturated with respect to PGA2 and operating at or near physiological capacity. Significance: The high, constitutive activity of Cyphoma GSTs is likely necessitated by the ubiquitous presence of GST substrates and/or inhibitors in this consumer’s gorgonian diet. This generalist’s GSTsmay operate as ‘all-purpose’ detoxification enzymes, capable of conjugating or sequestering a broad range of lipophilic gorgonian compounds, thereby allowing this predator to exploit a range of chemically-defended prey, resulting in a competitive dietary advantage for this species.

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Drugs from the Sea

2006-08-26 , Ludovice, Peter J. , Hunt, William D. , Kubanek, Julia

Our guest Prof. Julia Kubanek from the School of Biology and the School of Chemistry and Biochemistry at Georgia Tech will discuss how pharmaceutical agents can be extracted from marine organisims.

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Palatability and defense of some tropical infaunal worms: alkylpyrrole sulfamates as deterrents to fish feeding

2003-11-28 , Kicklighter, Cynthia Ellen , Kubanek, Julia , Barsby, Todd , Hay, Mark E.

Numerous studies have investigated chemical defenses among sessile species growing on hard substrates, but few have addressed this for mobile species in soft-sediment communities. We investigated the palatability and potential chemical defenses of 11 worm species from soft-sediment systems in southern Florida, USA. Three species were unpalatable to the bluehead wrasse Thalassoma bifasciatum. The polychaete Cirriformia tentaculata and the hemichordate Ptychodera bahamensis were uniformly unpalatable. For the polychaete Eupolymnia crassicornis, the exposed tentacles were unpalatable, but the body, which remains protected in a deeply buried tube, was palatable. These unpalatable worms were chemically defended; extracts of C. tentaculata, P. bahamensis, and the tentacles of E. crassicornis deterred fish feeding. For C. tentaculata, bioassay-guided fractionation demonstrated that a mixture of 3 closely related alkylpyrrole sulfamates deterred fish at naturally occurring concentrations (2-n-hexylpyrrole sulfamate [1.6% of worm dry mass], 2-n-heptylpyrrole sulfamate [3.1% dry mass], and 2-n-octylpyrrole sulfamate [0.8% dry mass]). This appears to be the first documentation of characterized natural products defending a marine worm from consumers. For P. bahamensis and the tentacles of E. crassicornis, deterrent effects of crude extracts decomposed before specific compounds could be identified

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