Copper carbenes alkylate guanine chemoselectively through a substrate directed reaction

Cu(I) carbenes derived from α-diazocarbonyl compounds lead to selective alkylation of the O6 position in guanine (O6-G) in mono- and oligonucleotides. Only purine-type lactam oxygens are targeted – other types of amides or lactams are poorly reactive under conditions that give smooth alkylation of guanine. Mechanistic studies point to N7G as a directing group that controls selectivity. Given the importance of O6-G adducts in biology and biotechnology we expect that Cu(I)-catalyzed O6-G alkylation will be a broadly used synthetic tool. While the propensity for transition metals to increase redox damage is well-appreciated, our results suggest that transition metals might also increase the vulnerability of nucleic acids to alkylation damage

Uso e implementación de TIC por los docentes de biología: Sede Leónidas Anastasi (Drago) del Ciclo Básico Común (Universidad de Buenos Aires)

Las nuevas tecnologías de la información y la comunicación (TIC) pueden actuar como recursos potentes para la enseñanza de las Ciencias Biológicas en nuestros cursos del Ciclo Básico Común (CBC) de la Universidad de Buenos Aires (UBA). Sin embargo, hemos observado que en el grupo de docentes de Biología hay cierta resistencia e incluso a veces se percibe forzada su implementación en el aula. Esta actitud podría conducir a que el uso de la tecnología pierda su sentido didáctico, pasando a ser un añadido o anexo, en vez de estar integrado y formar parte del proceso de enseñanza. Con el objeto de mejorar la enseñanza promoviendo la implementación de la tecnología en futuras estrategias de intervención docente en las aulas de Biología del CBC, nos propusimos: realizar una investigación exploratoria descriptiva que permitiera conocer la situación actual de los docentes de Biología en nuestra Sede Leónidas Anastasi (Drago) en cuanto al uso e implementación de las nuevas tecnologías en las aulas. Los resultados obtenidos de las entrevistas llevadas a cabo dan cuenta que hay un incremento en la incorporación de los recursos tecnológicos y TIC; no obstante, también se logra develar la presencia de problemáticas, como son la adquisición de la infraestructura tecnológica y la cultura de su utilización, más allá de los ofrecimientos de capacitación que haya. Nuestro desafío es proyectar, en una segunda instancia y a partir de los resultados obtenidos en esta investigación, políticas de acción tanto formadoras como promovedoras de diseños de propuestas didácticas que integren TIC para la enseñanza de Biología en el CBC-Drago. Por tal motivo consideramos de suma importancia este primer abordaje exploratorio puesto que estamos convencidos que las propuestas didácticas serias deben gestarse a partir de conocer primero las necesidades de cada contexto socio institucional docente.The new information and communications technology (ICT) can act as powerful teaching respurce of Biological Sciences at our CBC courses at the University of Buenos Aires (UBA). However, we have observed that in the group of biology teachers there is some resistance and sometimes even it is perceived as forced its implementation in the classroom. This attitude could lead to that the use of technology loses its didactic sense, becoming an addition or annex, instead of being integrated and become part of the teaching process. In order to improve teaching by promoting the implementation of technology in future strategies of educational intervention in the CBC Biology classroom, we proposed: make a descriptive exploratory research to allow to know the current situation of Biology teachers in our Leonidas Headquarters Anastasi site (Drago) in the use and implementation of new technologies in the classroom. The results show that there is a significant increase in the incorporation of technology and ICT resources; however, it does not ignore the presence of problems, such as the acquisition of technological infrastructure and culture of use, beyond the existing training offers. Our challenge is to project, in a second instance and from the results obtained in this investigation, action policies both forming and promotive of didactic proposal designs that integrate ICT for teaching biology in the CBC-Drago. We consider important to account this first exploratory approach because we believe that the educational proposals should be appropriate to the needs of each partner institution teaching context.Fil: Sturla, Andrés. Universidad de Buenos Aires. Ciclo Básico Común; ArgentinaFil: Baldoni, Gabriela. Universidad de Buenos Aires. Ciclo Básico Común; ArgentinaFil: Todaro, Laura Beatriz. Universidad de Buenos Aires. Ciclo Básico Común; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Garofalo, Sofia Judith. Universidad de Buenos Aires. Ciclo Básico Común; Argentin

The High Resolution Structure of GDP-4-keto-6-deoxy-D-mannose epimerase/reductase

GDP-4-keto-6-deoxy-D-mannose epimerase/reductase is a bifunctional enzyme involved in the biosynthesis of cell-surface structures, such as blood group antigens. Each subunit in the homodimeric enzyme consists of two domains. The N-terminal domain displays a Rossmann-fold topology and binds the NADP+ coenzyme. The C-terminal domain is held to bind the substrate. The holo-enzyme structure has been refined at 1.45 Å resolution, based on synchrotron data, to a final R-factor of 0.127 (Rfree = 0.167). The refined protein model highlights several residues involved in coenzyme recognition and binding and suggests that the enzyme belongs to the short-chain dehydrogenase protein homology family. Implications of the catalytic mechanism are discussed

The use of an artificial nucleotide for polymerase-based recognition of carcinogenic O6-alkylguanine DNA adducts.

Enzymatic approaches for locating alkylation adducts at single-base resolution in DNA could enable new technologies for understanding carcinogenesis and supporting personalized chemotherapy. Artificial nucleotides that specifically pair with alkylated bases offer a possible strategy for recognition and amplification of adducted DNA, and adduct-templated incorporation of an artificial nucleotide has been demonstrated for a model DNA adduct O(6)-benzylguanine by a DNA polymerase. In this study, DNA adducts of biological relevance, O(6)-methylguanine (O(6)-MeG) and O(6)-carboxymethylguanine (O(6)-CMG), were characterized to be effective templates for the incorporation of benzimidazole-derived 2'-deoxynucleoside-5'-O-triphosphates ( BENZI: TP and BIM: TP) by an engineered KlenTaq DNA polymerase. The enzyme catalyzed specific incorporation of the artificial nucleotide BENZI: opposite adducts, with up to 150-fold higher catalytic efficiency for O(6)-MeG over guanine in the template. Furthermore, addition of artificial nucleotide BENZI: was required for full-length DNA synthesis during bypass of O(6)-CMG. Selective incorporation of the artificial nucleotide opposite an O(6)-alkylguanine DNA adduct was verified using a novel 2',3'-dideoxy derivative of BENZI: TP. The strategy was used to recognize adducts in the presence of excess unmodified DNA. The specific processing of BENZI: TP opposite biologically relevant O(6)-alkylguanine adducts is characterized herein as a basis for potential future DNA adduct sequencing technologies

Uso e implementación de TIC por los docentes de Biología. Sede Leónidas Anastasi (Drago) del Ciclo Básico Común (Universidad de Buenos Aires)

Las nuevas tecnologías de la información y la comunicación (TIC) pueden actuar como recursos potentes para la enseñanza de las Ciencias Biológicas en nuestros cursos del Ciclo Básico Común (CBC) de la Universidad de Buenos Aires (UBA). Sin embargo, hemos observado que en el grupo de docentes de Biología hay cierta resistencia e incluso a veces se percibe forzada su implementación en el aula. Esta actitud podría conducir a que el uso de la tecnología pierda su sentido didáctico, pasando a ser un añadido o anexo, en vez de estar integrado y formar parte del proceso de enseñanza. Con el objeto de mejorar la enseñanza promoviendo la implementación de la tecnología en futuras estrategias de intervención docente en las aulas de Biología del CBC, nos propusimos: realizar una investigación exploratoria descriptiva que permitiera conocer la situación actual de los docentes de Biología en nuestra Sede Leónidas Anastasi (Drago) en cuanto al uso e implementación de las nuevas tecnologías en las aulas. Los resultados obtenidos de las entrevistas llevadas a cabo dan cuenta que hay un incremento en la incorporación de los recursos tecnológicos y TIC; no obstante, también se logra develar la presencia de problemáticas, como son la adquisición de la infraestructura tecnológica y la cultura de su utilización, más allá de los ofrecimientos de capacitación que haya. Nuestro desafío es proyectar, en una segunda instancia y a partir de los resultados obtenidos en esta investigación, políticas de acción tanto formadoras como promovedoras de diseños de propuestas didácticas que integren TIC para la enseñanza de Biología en el CBC-Drago. Por tal motivo consideramos de suma importancia este primer abordaje exploratorio  puesto que estamos convencidos que las propuestas didácticas serias deben gestarse a partir de conocer primero las  necesidades de cada contexto socio institucional docente.  Palabras clave: enseñanza; biología; TIC; superior; docentes The new information and communications technology (ICT) can act as powerful teaching respurce of Biological Sciences at our CBC courses at the University of Buenos Aires (UBA). However, we have observed that in the group of biology teachers there is some resistance and sometimes even it is perceived as forced its implementation in the classroom. This attitude could lead to that the use of technology loses its didactic sense, becoming an addition or annex, instead of being integrated and become part of the teaching process. In order to improve teaching by promoting the implementation of technology in future strategies of educational intervention in the CBC Biology classroom, we proposed: make a descriptive exploratory research to allow to know the current situation of Biology teachers in our Leonidas Headquarters Anastasi site (Drago) in the use and implementation of new technologies in the classroom. The results show that there is a significant increase in the incorporation of technology and ICT resources; however, it does not ignore the presence of problems, such as the acquisition of technological infrastructure and culture of use, beyond the existing training offers. Our challenge is to project, in a second instance and from the results obtained in this investigation, action policies both forming  and promotive of didactic proposal designs that integrate ICT for teaching biology in the CBC-Drago. We consider important to account this first exploratory approach because we believe that the educational proposals should be appropriate to the needs of each partner institution teaching context. Keywords: education; biology; ICT; higher education teachers

G-protein coupling and nuclear translocation of the human abscisic acid receptor LANCL2

Abscisic acid (ABA), a long known phytohormone, has been recently demonstrated to be present also in humans, where it targets cells of the innate immune response, mesenchymal and hemopoietic stem cells and cells involved in the regulation of systemic glucose homeostasis. LANCL2, a peripheral membrane protein, is the mammalian ABA receptor. We show that N-terminal glycine myristoylation causes LANCL2 localization to the plasmamembrane and to cytoplasmic membrane vesicles, where it interacts with the \u3b1 subunit of a Gi protein and starts the ABA signaling pathway via activation of adenylate cyclase. Demyristoylation of LANCL2 by chemical or genetic means triggers its nuclear translocation. Nuclear enrichment of native LANCL2 is also induced by ABA treatment. Therefore human LANCL2 is a non-transmembrane G protein-coupled receptor susceptible to hormone-induced nuclear translocation

Galanin n-terminal fragment (1-15) decreases the voluntary alcohol intake in rats

Galanin (GAL) is involved in drug abuse and addiction including alcohol intake. In this work, we have analysed the role of the N-terminal GAL fragment (1-15) [GAL(1-15)] in voluntary ethanol consumption in rats using the two-bottle choice paradigm as well as compare the effects of GAL(1-15) with GAL. The two-bottle choice test was used to determine the voluntary ethanol consumption of rats (Castilla-Ortega et al., 2016). Three sets of experiments were conducted. In the first set of experiments, a dose-response curve of GAL(1-15) was performed. Groups of rats (n=7-9) received i.c.v. GAL(1-15) 1 nmol, 3 nmol or vehicle 2, 14 and 24 hours before the measures. In the second set of experiments, the effects in two-bottle choice test of GAL 3 nmol, and GAL(1-15) 3 nmol were compared. In the last set of experiments rats received i.c.v. GAL(1-15) 3nmol combined with GALR2 antagonist M871 3 nmol 2 hours before the measures. GAL(1-15) 3nmol significantly decreased the alcohol intake 2 (p<0.05), 14 (p<0.05) and 24 (p<0.05) hours after its administration. Moreover, 2 hours after i.c.v. GAL(1-15) 3nmol a significantly decreased by 90% in preference was observed (p<0.05). This effect was maintained 24hours. GAL(1-15) also significantly reduced the alcohol intake (p<0.05) and preference (p<0.05) compared with GAL. GALR2 antagonist M871 significantly blocked the decreased in the ethanol intake (p<0.05) and preference (p<0.05) induced by GAL(1-15) 2 hours after its administration. These results indicates that GAL(1-15) induces a strong reduction in preference and alcohol consumption in rat, showing a differential role than GAL. These results may give basis for the development of novel therapeutic strategies using GAL(1-15) for treatment of alcohol addiction.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. This study was supported by Spanish SAF2016-79008-P and PPIT.UMA.B1.2017/17

Identification of a high affinity binding site for abscisic acid on human lanthionine synthetase component C-like protein 2

Lanthionine synthetase component C-like protein 2 (LANCL2) has been identified as the mammalian receptor mediating the functional effects of the universal stress hormone abscisic acid (ABA) in mammals. ABA stimulates insulin independent glucose uptake in myocytes and adipocytes via LANCL2 binding in vitro, improves glucose tolerance in vivo and induces brown fat activity in vitro and in vivo. The emerging role of the ABA/LANCL2 system in glucose and lipid metabolism makes it an attractive target for pharmacological interventions in diabetes mellitus and the metabolic syndrome. The aim of this study was to investigate the presence of ABA binding site(s) on LANCL2 and identify the amino acid residues involved in ABA binding. Equilibrium binding assays ([3H]-ABA saturation binding and surface plasmon resonance analysis) suggested multiple ABA-binding sites, prompting us to perform a computational study that indicated one putative high-affinity and two low-affinity binding sites. Site-directed mutagenesis (single mutant R118I, triple mutants R118I/R22I/K362I and R118I/S41A/E46I) and equilibrium binding experiments on the mutated LANCL2 proteins identified a high-affinity ABA-binding site involving R118, with a KD of 2.6 nM ± 1.2 nM, as determined by surface plasmon resonance. Scatchard plot analysis of binding curves from both types of equilibrium binding assays revealed a Hill coefficient >1, suggesting cooperativity of ABA binding to LANCL2. Identification of the high-affinity ABA-binding site is expected to allow the design of ABA agonists/antagonists, which will help to understand the role of the ABA/LANCL2 system in human physiology and disease

Abscisic Acid: A Novel Nutraceutical for Glycemic Control

Abscisic acid is naturally present in fruits and vegetables, and it plays an important role in managing glucose homeostasis in humans. According to the latest U.S. dietary survey, about 92% of the population might have a deficient intake of ABA due to their deficient intake of fruits and vegetables. This review summarizes the in vitro, preclinical, mechanistic, and human translational findings obtained over the past 15 years in the study of the role of ABA in glycemic control. In 2007, dietary ABA was first reported to ameliorate glucose tolerance and obesity-related inflammation in mice. The most recent findings regarding the topic of ABA and its proposed receptor lanthionine synthetase C-like 2 in glycemic control and their interplay with insulin and glucagon-like peptide-1 suggest a major role for ABA in the physiological response to a glucose load in humans. Moreover, emerging evidence suggests that the ABA response might be dysfunctional in diabetic subjects. Follow on intervention studies in healthy individuals show that low-dose dietary ABA administration exerts a beneficial effect on the glycemia and insulinemia profiles after oral glucose load. These recent findings showing benefits in humans, together with extensive efficacy data in mouse models of diabetes and inflammatory disease, suggest the need for reference ABA values and its possible exploitation of the glycemia-lowering effects of ABA for preventative purposes. Larger clinical studies on healthy, prediabetic, and diabetic subjects are needed to determine whether addressing the widespread dietary ABA deficiency improves glucose control in human

Extracellular NAD + Is an Agonist of the Human P2Y 11 Purinergic Receptor in Human Granulocytes

Micromolar concentrations of extracellular beta-NAD+ (NAD(e)+) activate human granulocytes (superoxide and NO generation and chemotaxis) by triggering: (i) overproduction of cAMP, (ii) activation of protein kinase A, (iii) stimulation of ADP-ribosyl cyclase and overproduction of cyclic ADP-ribose (cADPR), a universal Ca2+ mobilizer, and (iv) influx of extracellular Ca2+. Here we demonstrate that exposure of granulocytes to millimolar rather than to micromolar NAD(e)+ generates both inositol 1,4,5-trisphosphate (IP3) and cAMP, with a two-step elevation of intracellular calcium levels ([Ca2+]i): a rapid, IP3-mediated Ca2+ release, followed by a sustained influx of extracellular Ca2+ mediated by cADPR. Suramin, an inhibitor of P2Y receptors, abrogated NAD(e)+-induced intracellular increases of IP3, cAMP, cADPR, and [Ca2+]i, suggesting a role for a P2Y receptor coupled to both phospholipase C and adenylyl cyclase. The P2Y(11) receptor is the only known member of the P2Y receptor subfamily coupled to both phospholipase C and adenylyl cyclase. Therefore, we performed experiments on hP2Y(11)-transfected 1321N1 astrocytoma cells: micromolar NAD(e)+ promoted a two-step elevation of the [Ca2+]i due to the enhanced intracellular production of IP3, cAMP, and cADPR in 1321N1-hP2Y(11) but not in untransfected 1321N1 cells. In human granulocytes NF157, a selective and potent inhibitor of P2Y(11), and the down-regulation of P2Y(11) expression by short interference RNA prevented NAD(e)+-induced intracellular increases of [Ca2+]i and chemotaxis. These results demonstrate that beta-NAD(e)+ is an agonist of the P2Y(11) purinoceptor and that P2Y(11) is the endogenous receptor in granulocytes mediating the sustained [Ca2+]i increase responsible for their functional activation