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Discovery of Novel N-acylethanolamines in Early Land Plants and Their ImplicationsKilaru, Aruna 01 January 2017 (has links)
N-acylethanolamines (NAEs) are family of lipid-derived signaling molecules in many organisms, which include an endocannabinoid N-arachidonoylethanolamide (anandamide, AEA, NAE 20:4). NAEs, specifically AEA plays pivotal role in mammalian neurological and physiological functions; however their metabolism and functional implications in plants are yet to be fully discovered. Unlike seed plants, bryophytes possess unique fatty acid composition that includes 20:4 and 20:5, which prompted our search for endocannabinods in moss Physcomitrella patens. To this extent, we used targeted lipidomic analyses and discovered long-chain NAEs and their corresponding N-acyl-phosphatidylethalamine (NAPE) precursors in an early land plant species. In protonemal tissues N-arachidonyl-PE and N-20:5-PE contributed to about 49% and 30%, respectively. Matured gametophytes on the other hand showed a 12% increase in N-20:4-PE and 20% decline in N-20:5-PE, relative to NAPE content in protonema. In all haploid developmental stages analyzed NAE 20:4 levels contributed to ~ 23% of the total NAE while NAE 20:5 remained as a minor component (5%). Interestingly, in Selaginella moellendorffi, an early vascular plant N-18:2-PE species was most abundant; although minor amounts of N-20:3-PE, N-20:4-PE and N-20:5-PE were present, only a small quantity of NAE 20:4 was identified among the 20C NAEs. Both AEA and it corresponding fatty acid, arachidonic acid have growth inhibitory effects in a dose dependent manner. Biological implications of anandamide and its metabolic pathway in moss are under investigation. Our data reveals an evolutionarily conserved occurrence of NAE metabolites in early land plants, with an exclusive report of AEA presence in a bryophyte.
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Marijuana to Moss: Discovery and Implications of N-acylethanolaminesKilaru, Aruna 23 April 2018 (has links)
No description available.
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Discovery Of Novel N-Acylethanolamines In Early Land Plants And Their ImplicationsShinde, Suhas, Devaiah, Shivakumar, Welti, Ruth, Kilaru, Aruna 24 June 2017 (has links)
N-acylethanolamines (NAEs) are family of lipid-derived signaling molecules in many organisms, which include an endocannabinoid N-arachidonoylethanolamide (anandamide, AEA, NAE 20:4). NAEs, specifically AEA plays pivotal role in mammalian neurological and physiological functions; however their metabolism and functional implications in plants are yet to be fully discovered. Unlike seed plants, bryophytes possess unique fatty acid composition that includes 20:4 and 20:5, which prompted our search for endocannabinods in moss Physcomitrella patens. To this extent, we used targeted lipidomic analyses and discovered long-chain NAEs and their corresponding N-acyl-phosphatidylethalamine (NAPE) precursors in an early land plant species. In protonemal tissues N-arachidonyl-PE and N-20:5-PE contributed to about 49% and 30%, respectively. Matured gametophytes on the other hand showed a 12% increase in N-20:4-PE and 20% decline in N-20:5-PE, relative to NAPE content in protonema. In all haploid developmental stages analyzed NAE 20:4 levels contributed to ~ 23% of the total NAE while NAE 20:5 remained as a minor component (5%). Interestingly, in Selaginella moellendorffi, an early vascular plant N-18:2-PE species was most abundant; although minor amounts of N-20:3-PE, N-20:4-PE and N-20:5-PE were present, only a small quantity of NAE 20:4 was identified among the 20C NAEs. Both AEA and it corresponding fatty acid, arachidonic acid have growth inhibitory effects in a dose dependent manner. Biological implications of anandamide and its metabolic pathway in moss are under investigation. Our data reveals an evolutionarily conserved occurrence of NAE metabolites in early land plants, with an exclusive report of AEA presence in a bryophyte.
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Composition of N-Acylethanolamines in Physcomitrella Patens at Varying Life StagesFarley, C., Kilaru, Aruna, Devaiah, Shivakumar, Roth, M., Shiva, A., Tamura, P., Welti, Ruth 01 January 2016 (has links)
No description available.
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The Role of N-Acylethanolamines in the Development of Physcomitrella PatensSante, Richard, Kilaru, Aruna 05 April 2012 (has links)
Global demand for food, which is expected to double by 2050, presents plant biologists with a major challenge to generate higher yields on existing croplands. The long-term goal of the proposed research is to generate stress tolerant plants and improve crop productivity. Stress responses, in plants or animals, most often involve the activation stress-signaling pathways. One pathway is mediated by a group of bioactive lipid molecules, N-acylethanolamines (NAEs), and is highly conserved among eukaryotes. NAEs are composed mainly of amides of ethanolamine and fatty acids. In model Arabidopsis thaliana (At), NAEs inhibit germination and growth in seedlings via abscisic-dependent and independent mechanisms aimed at stress regulation. This work focuses on identifying the NAE pathway in the moss, Physcomitrella patens (P. patens), that shows higher resistant to stress than At and consequently the role played by NAE metabolites in the development of P. patens. Mosses are small seedless and fruitless early plants that have evolved successful mechanisms of surviving prolonged stresses during their transition from aquatic life to land. Previous studies have shown that P. patens is tolerant to high temperature, salinity and osmotic shock, to which most plants are fragile. I hypothesize that the NAE pathway is present in P. patens and that it plays a significant role in its development. NAE composition and metabolizing enzymes like FAAH and NAPE synthase will be measured to ascertain NAE pathway in P. patens. Further quantification of NAE content and composition during development will identify the key developmental time points that are associated with high metabolite levels. Total lipids will be extracted in triplicates from twelve tissue samples harvested at protonema stage; mature gametophyte stage which is subdivided into leafy tissue and rhizoids; antheridia and archegonia stage; zygotic stage and the mature and germinating developmental stages. The high lipid content of the mature spores was previously associated with extreme longevity in ephemeral habitats. It would be of interest to determine if such an association exists with NAE metabolite content. Key time points will then be used to determine the effect of exogenous NAE 12:0. Preliminary analysis shows that P. patens have higher levels of NAEs than other plants examined. Furthermore, fatty acid amide hydrolases (FAAH) that hydrolyses NAEs were identified in silico. Putative PpFAAH1 candidate showed 95 % homology with previously characterized AtFAAH in At. These results show the occurrence of NAEs in P. patens and a possible enzyme for its hydrolysis. Further confirmation of the NAE pathway in P. patens will be done by in silico identification of the precursor enzyme NAPE synthase. Understanding how P. patens tolerates stress using NAEs pathways by identifying key time points in its development will help other researchers know which specific NAEs affect the growth of P. patens and will facilitate the characterization of metabolic enzymes at specific times.
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Novel Polyunsaturated N-acylethanolamines (NAE) and Their Role in Physcomitrella PatensKilaru, Aruna, Haq, Imadadul, Chilufya, Jedaidah, Devaiah, Shivakumar, Shinde, Sushas, Welti, Ruth 22 June 2017 (has links)
Anandamide (N-arachidonoylethanolamide, AEA), a 20C polyunsaturated (PU) N-acylethanolamine (NAE) influences many neurological functions in mammals. Although 20C PU-NAEs are considered unique to animals, they were recently discovered in early land plants but their metabolism and functions remain unknown. Comprehensive lipidomic analyses of Physcomitrella patens revealed not only abundance of arachidonic acid (AA, 20:4) and eicosapentaenoic acid (EPA, 20:5) but also their corresponding ethanolamides (AEA and EPEA, respectively). While moss showed increasing AA with development, 14% and 24% in protonemata and gametophyte tissues, respectively, EPA decreased from 7% in protonemata to ~1.3 % in gametophytes. An increase in 20:4- and decrease in 20:5- ethanolamides and their corresponding membrane precursors, phosphatidylethanolamides, also was observed during gametophyte development. Pharmacological studies revealed that AEA specifically inhibits polarized tip growth, which justifies the low endogenous levels of AEA in protonemata. To further determine the physiological relevance of these 20C PU-NAEs, a fatty acid amide hydrolase that catabolizes NAEs has been heterologously characterized. Furthermore, generation of metabolite mutants with altered NAE levels is underway. Overall, we identified two novel NAEs, AEA and EPEA in Physcomitrella, which may play an important role in regulation of moss growth and development, although the underlying mechanism is still unclear.
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Analysis of N-Acylethanolamines in the Oilseed Crop Camelina sativaCorley, Chase D 08 1900 (has links)
To better understand the nature and function of N-acylethanolamines (NAEs) in Camelina sativa, we used mass spectrometry analysis to identify and quantify NAE types in developing seeds, desiccated seeds and seedlings. Developing seeds showed a differential increase in individual NAE species and an overall increase in NAE content with seed development and maturation. The NAE composition in mature, desiccated seeds mostly reflected the total fatty acid composition in the seed tissues, except for a noted absence of 11-eicosenoic (20C monounsaturated) fatty acid in the NAE pool. During seed stratification and seedling growth, individual NAE species were depleted at similar rates. Simulated drought treatments during seedling development resulted in a significant rise in NAE levels for the major 18C NAE types compared with untreated seedlings. Arabidopsis and Camelina mutants with reported altered fatty acid profiles were analyzed for their NAE compositions; both Arabidopsis and Camelina had relatively similar changes between compositions of total seed fatty acids and NAEs. Furthermore, seeds were analyzed from transgenic Arabidopsis and Camelina with engineered, non-native, long-chain polyunsaturated fatty acids (18C, 20C and 22C), and the results showed the production of novel N-acylphosphatidylethanolamines (presumed precursors of NAEs) and NAEs with the same long acyl chains. These results demonstrate i) that NAE levels change dramatically with seed and seedling development in Camelina, ii) that NAE levels in Camelina seedlings can be elevated in response to environmental stress, and iii) that the pool of NAEs can be manipulated by making alterations to seed fatty acid compositions, some of which may have promise for the production of novel, bioactive NAEs.
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Endocannabinoid System in a Planarian ModelMustonen, Katie Lynn 12 1900 (has links)
In this study, the presence and possible function of endocannabinoid ligands in the planarian is investigated. The endocannabinoids ananadamide (AEA) and 2-arachidonoylglycerol (2-AG) and entourage NAE compounds palmitoylethanolamide (PEA), stearoylethanolamide (SEA) and oleoylethanolamide (OEA) were found in Dugesia dorotocephala. Changes in SEA, PEA, and AEA levels were observed over the initial twelve hours of active regeneration. Exogenously applied AEA, 2-AG and their catabolic inhibition effected biphasic changes in locomotor velocity, analogous to those observed in murines. The genome of a close relative, Schmidtea mediterranea, courtesy of the University of Utah S. med genome database, was explored for cannabinoid receptors, none were found. A putative fatty acid amide hydrolase (FAAH) homolog was found in Schmidtea mediterranea.
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Novel Polyunsaturated N-acylethanolamines and Their Implications in Physcomitrella patensShinde, S., Welti, R., Kilaru, Aruna 01 January 2018 (has links)
No description available.
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Biochemical Characterization of Tomato Fatty Acid Amide HydrolaseShrestha, Sujan 01 August 2018 (has links) (PDF)
Fatty acid amide hydrolase (FAAH) is an enzyme that terminates the signaling role played by the lipid mediators, N- acylethanolamines (NAEs), present both in plants and animals. FAAH is responsible for NAE hydrolysis and has been extensively studied in mammalian systems and the model plant Arabidopsis thaliana; it has been reported in various organisms as well as some crop plants such as rice and Medicago truncatula. To understand the role of FAAH in diverse organisms, here we report the identification and biochemical characterization of a FAAH homolog in tomato. Previously identified and cloned candidate FAAH from tomato was expressed in Escherichia coli as a fused protein with 6X his-tag for identification. Supernatant containing recombinant FAAH showed the ability to hydrolyze NAE substrates. The optimal reaction conditions for enzyme assay and kinetic parameters for tomato FAAH were determined and effect of inhibitor on enzyme was determined. Characterization of FAAH in tomato will contribute to further understanding of NAE metabolic pathway and its implications.
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