Identification of genes coding for putative wax ester synthase/diacylglycerol acyltransferase enzymes in terrestrial and marine environments

Synthesis of neutral lipids such as triacylglycerols (TAG) and wax esters (WE) is catalyzed in bacteria by wax ester synthase/diacylglycerol acyltransferase enzymes (WS/DGAT). We investigated the diversity of genes encoding this enzyme in contrasting natural environments from Patagonia (Argentina). The content of petroleum hydrocarbons in samples collected from oil-producing areas was measured. PCR-based analysis covered WS/DGAT occurrence in marine sediments and soil. No product was obtained in seawater samples. All clones retrieved from marine sediments affiliated with gammaproteobacterial sequences and within them, most phylotypes formed a unique cluster related to putative WS/DGAT belonging to marine OM60 clade. In contrast, soils samples contained phylotypes only related to actinomycetes. Among them, phylotypes affiliated with representatives largely or recently reported as oleaginous bacteria, as well as with others considered as possible lipid-accumulating bacteria based on the analysis of their annotated genomes. Our study shows for the first time that the environment could contain a higher variety of ws/ dgat than that reported from bacterial isolates. The results of this study highlight the relevance of the environment in a natural process such as the synthesis and accumulation of neutral lipids. Particularly, both marine sediments and soil may serve as a useful source for novel WS/DGAT with biotechnological interest.Fil: Lanfranconi, Mariana Patricia. Universidad Nacional de la Patagonia "San Juan Bosco". Facultad de Ciencias Naturales - Sede Comodoro; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Alvarez, Adrián F.. Universidad Nacional Autónoma de México; MéxicoFil: Alvarez, Hector Manuel. Universidad Nacional de la Patagonia "San Juan Bosco". Facultad de Ciencias Naturales - Sede Comodoro; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Identification of a novel ATP-binding cassette transporter involved in long-chain fatty acid import and its role in triacylglycerol accumulation in Rhodococcus jostii RHA1

Members of Rhodococcus genus are specialists in the biosynthesis and accumulation of triacylglycerols (TAG). Since no transport protein related with TAG metabolism has been characterized until now in these bacteria, we used the available genomic information of Rhodococcus jostii RHA1 to perform a broad survey of genes coding for putative lipid transporter proteins in this oleaginous microorganism. Among the seven genes encoding for putative lipid transporters, ro05645 (now called ltp1: lipid transporter protein) coding for an ABC protein was found clustered with others genes encoding for enzymes catalyzing the three putative acylation reactions of the Kennedy pathway for TAG synthesis. Overexpression of ltp1 gene in the RHA1 strain led to an increase of approximately six- and three-folds in biomass and TAG production; respectively, when cells were cultivated on palmitic acid and oleic acid. Moreover, overexpression of ltp1 also promoted a significant increase in the uptake of a fluorescently labeled long-chain fatty acid in comparison with the wild type strain RHA1, and its further incorporation into the TAG fraction. Gluconate-grown cells showed increasing amounts of intracellular free fatty acids, but not of TAG, after overexpressing ltp1 gene. Thus, for the first time to our knowledge, a transporter functionally related to TAG metabolism was identified in oleaginous rhodococci. Our results suggest that Ltp1 is an importer of long chain fatty acids, which plays a functional role in lipid homeostasis of R. jostii RHA1.Fil: Villalba, María S.. Universidad Nacional de la Patagonia "San Juan Bosco". Facultad de Ciencias Naturales - Sede Comodoro; ArgentinaFil: Alvarez, Hector Manuel. Universidad Nacional de la Patagonia "San Juan Bosco". Facultad de Ciencias Naturales - Sede Comodoro; Argentina. Oil m&s; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Nacional Patagónico; Argentin

The pleiotropic transcriptional regulator NlpR contributes to the modulation of nitrogen metabolism, lipogenesis and triacylglycerol accumulation in oleaginous rhodococci

The regulatory mechanisms involved in lipogenesis and triacylglycerol (TAG) accumulation are largely unknown in oleaginous rhodococci. In this study a regulatory protein (here called NlpR: Nitrogen lipid Regulator), which contributes to the modulation of nitrogen metabolism, lipogenesis and triacylglycerol accumulation in oleaginous rhodococci was identified. Under nitrogen deprivation conditions, in which TAG accumulation is stimulated, the nlpR gene was significantly upregulated, whereas a significant decrease of its expression and TAG accumulation occurred when cerulenin was added. The nlpR disruption negatively affected the nitrate/nitrite reduction as well as lipid biosynthesis under nitrogen-limiting conditions. In contrast, its overexpression increased TAG production during cultivation of cells in nitrogen-rich media. A putative ‘NlpR-binding motif’ upstream of several genes related to nitrogen and lipid metabolisms was found. The nlpR disruption in RHA1 strain led to a reduced transcription of genes involved in nitrate/nitrite assimilation, as well as in fatty acid and TAG biosynthesis. Purified NlpR was able to bind to narK, nirD, fasI, plsC and atf3 promoter regions. It was suggested that NlpR acts as a pleiotropic transcriptional regulator by activating of nitrate/nitrite assimilation genes and others genes involved in fatty acid and TAG biosynthesis, in response to nitrogen deprivation.Fil: Hernández, Martín Alejandro. Universidad Nacional de la Patagonia "san Juan Bosco". Instituto de Biociencias de la Patagonia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Instituto de Biociencias de la Patagonia; ArgentinaFil: Lara, María Julia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Gago, Gabriela Marisa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Gramajo, Hugo Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Alvarez, Hector Manuel. Universidad Nacional de la Patagonia "san Juan Bosco". Instituto de Biociencias de la Patagonia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Instituto de Biociencias de la Patagonia; Argentin

Enantiomeric Ratio Changes of Terpenes in Essential Oils from Hybrid Eucalyptus grandis × E. tereticornis and its Pure Species

Some Eucalyptus species produce oils with biological activities and the effect of their interspecifc hybridization on the enantiomeric composition of terpenes has not been reported. The enantiomeric excesses of monoterpenes in the essential oil of Eucalyptus grandis × E. tereticornis and its parental taxa were determined by gas chromatography-mass spectrometry (GC-MS), and to resolve coelutions problems by preparative high performance liquid chromatography (HPLC)and GC-MS with two columns in series. The essential oil composition of the hybrid presented qualitative and quantitative differences with the composition of its parental taxa. Great differences were found for the enantiomeric ratio in monoterpene alcohols among the three essential oils. Our results suggest that the enantiomeric analysis can be a reliable method for the study of how theinterspecifc hybridization can module the enantiomeric chemical profle in Eucalyptus essential oils. These results suggest the use of interspecifc hybridization to improve or expand the source of bioactive compounds.Fil: Naspi, Cecilia Veronica. Consejo Nacional de Investigaciones Científicas y Técnicas. Unidad de Investigación y Desarrollo Estratégico para la Defensa. Ministerio de Defensa. Unidad de Investigación y Desarrollo Estratégico para la Defensa; ArgentinaFil: Alvarez Costa, Agustin. Consejo Nacional de Investigaciones Científicas y Técnicas. Unidad de Investigación y Desarrollo Estratégico para la Defensa. Ministerio de Defensa. Unidad de Investigación y Desarrollo Estratégico para la Defensa; ArgentinaFil: Lucia, Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Unidad de Investigación y Desarrollo Estratégico para la Defensa. Ministerio de Defensa. Unidad de Investigación y Desarrollo Estratégico para la Defensa; ArgentinaFil: Gonzalez Audino, Paola Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Unidad de Investigación y Desarrollo Estratégico para la Defensa. Ministerio de Defensa. Unidad de Investigación y Desarrollo Estratégico para la Defensa; ArgentinaFil: Masuh, Hector Mario. Consejo Nacional de Investigaciones Científicas y Técnicas. Unidad de Investigación y Desarrollo Estratégico para la Defensa. Ministerio de Defensa. Unidad de Investigación y Desarrollo Estratégico para la Defensa; Argentin

On the kinetic and allosteric regulatory properties of the ADP-glucose pyrophosphorylase from Rhodococcus jostii: An approach to evaluate glycogen metabolism in oleaginous bacteria

Rhodococcus spp. are oleaginous bacteria that accumulate glycogen during exponential growth. Despite the importance of these microorganisms in biotechnology, little is known about the regulation of carbon and energy storage, mainly the relationship between glycogen and triacylglycerols metabolisms. Herein, we report the molecular cloning and heterologous expression of the gene coding for ADP-glucose pyrophosphorylase (EC 2.7.7.27) of Rhodococcus jostii, strain RHA1. The recombinant enzyme was purified to electrophoretic homogeneity to accurately characterize its oligomeric, kinetic, and regulatory properties. The R. jostii ADP-glucose pyrophosphorylase is a homotetramer of 190 kDa exhibiting low basal activity to catalyze synthesis of ADP-glucose, which is markedly influenced by different allosteric effectors. Glucose-6P, mannose-6P, fructose-6P, ribose-5P, and phosphoenolpyruvate were major activators; whereas, NADPH and 6P-gluconate behaved as main inhibitors of the enzyme. The combination of glucose-6P and other effectors (activators or inhibitors) showed a cross-talk effect suggesting that the different metabolites could orchestrate a fine regulation of ADP-glucose pyrophosphorylase in R. jostii. The enzyme exhibited some degree of affinity toward ATP, GTP, CTP, and other sugar-1P substrates. Remarkably, the use of glucosamine-1P was sensitive to allosteric activation. The relevance of the fine regulation of R. jostii ADP-glucose pyrophosphorylase is further analyzed in the framework of proteomic studies already determined for the bacterium. Results support a critical role for glycogen as a temporal reserve that provides a pool of carbon able of be re-routed to produce long-term storage of lipids under certain conditions.Fil: Cereijo, Antonela Estefanía. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; ArgentinaFil: Asención Diez, Matías Damián. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; ArgentinaFil: Dávila Costa, José Sebastián. Universidad Nacional de la Patagonia Austral. Centro de Investigaciones y Transferencia Golfo San Jorge. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro de Investigaciones y Transferencia Golfo San Jorge. Universidad Nacional de la Patagonia ; ArgentinaFil: Alvarez, Hector Manuel. Universidad Nacional de la Patagonia Austral. Centro de Investigaciones y Transferencia Golfo San Jorge. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro de Investigaciones y Transferencia Golfo San Jorge. Universidad Nacional de la Patagonia ; ArgentinaFil: Iglesias, Alberto Alvaro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentin

Early evolutionary differentiation of morphological variation in the mandible of South American caviomorph rodents (Rodentia, Caviomorpha)

Caviomorphs are a clade of South American rodents recorded at least since the early Oligocene (>31.5Ma) that exhibit ample eco-morphological variation. It has been proposed that phylogenetic structure is more important than ecological factors for understanding mandibular shape variation in this clade. This was interpreted as a result of the long-standing evolutionary history of caviomorphs and the early divergence of major lineages. In this work, we test this hypothesis through the analysis of morphological variation in the mandible of living and extinct species and compare this information with that obtained through comparative phylogenetic analyses. Our results support the hypothesis of early origin of mandibular variation; moreover, they suggest the conservation of early differentiated morphologies, which could indicate the existence of constrained evolutionary diversification. © 2011 The Authors.Fil: Alvarez, Alicia. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. División Zoología de Vertebrados. Sección de Mastozoología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; ArgentinaFil: Perez, Sergio Ivan. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. División Antropología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; ArgentinaFil: Verzi, Diego Hector. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. División Zoología de Vertebrados. Sección de Mastozoología; Argentin

The Perceptions of Adults 35 and Older On Online Learning

This qualitative case study examined the level of satisfaction among a group of adults ages 35 years and older enrolled in an online education program (OLEP) in a university in Puerto Rico. Although the current literature revealed that adult students 35 years and older are the fastest growing population enrolling in online college education programs in Puerto Rico, prior satisfaction studies conducted by this institution did not focus on this population. The theoretical framework of this study was guided by Holsapple and Lee\u27s Post e-learning success model. The goal of this study was to understand students\u27 satisfaction with the online program and determine if the program was helping them accomplish their goals. Data were collected through semi-structured individual interviews with 8 adult students, 35 years of age or older, who were currently enrolled at the institution. Data were analyzed using the category construction approach, open coding, and thematic analysis. Results indicated that the participants had a positive perception of the online program and its impact on their academic development and educational success. The data also revealed issues related to faculty-student communication and course design, which the participants believed needed to improve. The study\u27s findings helped in the development of a best practice manual for the OLEP faculty. The manual will provide OLEP faculty with the tools needed to improve faculty-student communication and online course design, thereby increasing the student satisfaction among the fastest growing online student population. Improving its OLEP shows promise for the university to continue to be an agent of social change for Puerto Rico\u27s economic growth and social progress

The democratic basis for the revocation of the representative mandate

La revocación del mandato representativo es un derecho político fundamental, imbricado en el principio democrático, porque las facultades o atribuciones que confiere integran el estatuto jurídico-político del ciudadano al estar referidas a la participación popular en los asuntos públicos, concretamente al control y exigencia de responsabilidad política de los representantes públicos. Esta tesis doctoral presenta la hipótesis referida en el título: el fundamento de la revocación del mandato representativo reside en la soberanía popular. ¿Qué implica? Primero: la revocación tiene alma democrática y su no reconocimiento constitucional entraña una restricción de la libertad política, inasumible en el Estado democrático; segundo: la institución revocatoria no es propia ni exclusiva del mandato imperativo y tercero: el mandato representativo no implica transmisión o alienación de la titularidad ni del ejercicio de la soberanía sino sólo la delegación o la comisión del ejercicio de los poderes públicos. Este trabajo de investigación pretende dar respuesta a cuestiones capitales de la teoría democrática: ¿por qué el pueblo es el soberano?, ¿cuál es el origen del fundamento democrático de la soberanía? y ¿qué significa la soberanía popular? Debemos partir de estos sólidos cimientos para poder explicar el significado y las implicaciones jurídico-constitucionales del mandato representativo. Y, finalmente, analizar por qué la revocación del mandato representativo es una exigencia indeclinable del principio democrático. En esta tesis surcaremos la Historia de las ideas políticas, atravesaremos la ideología constitucional, nos sumergiremos en la teoría del Derecho y nos asomaremos a la antropología en busca de los argumentos que nos permitan aseverar, con rigor científico, que la revocación del mandato representativo es inherente a la soberanía popular, clave de bóveda del constitucionalismo democrático.The revocation of the representative mandate is a fundamental political right, embedded in the democratic principle, as it confers powers and functions that integrate the citizen’s legal-political status by regulating popular participation in public affairs, particularly with regard to the control and the political accountability of public representatives. This doctoral thesis presents the assumptions outlined in the title: the basis for the revocation of the representative mandate lies on popular sovereignty. What does this imply? Firstly, the repeal has a democratic soul and its non-recognition in the constitution involves a restriction of political freedom, which is untenable in the democratic State; secondly, the revocative institution is not part nor it is exclusive of the imperative mandate; thirdly, the representative mandate does not imply the transmission or the alienation neither of the ownership nor of the exercise of sovereignty, but the delegation or commission of the exercise of public powers. This research seeks to answer crucial questions of the democratic theory: why are people sovereign? What is the origin of the democratic foundation of sovereignty? What does popular sovereignty mean? It is from this sound basis that the meaning and the legal and constitutional implications of the representative mandate should be explained. Finally, it is also important to analyze the reason why the revocation of representative mandate is an irrevocable requirement of the democratic principle. Throughout this thesis we will retrace the history of political ideas, we will go through the constitutional ideology, we will dive into the theory of law and we will take a look at anthropology, in search of arguments that will allow us to assert with scientific accuracy that the revocation of the representative mandate is pertinent to the people's sovereignty, a keystone of democratic constitutionalism