Étude in vivo de la relation entre la structure et la fonction de la boucle variable de l'ARN de transfert de la sélénocystéine d'E. coli

La sélénocystéine est le 21e acide aminé encodé génétiquement et on la retrouve à travers les trois domaines de la vie. Elle est synthétisée sur l'ARNtSec par un processus unique. L'ARNtSec se distingue également au niveau structural. La tige acceptrice possède 8 (procaryotes) et 9 (eucaryotes) paires de bases, contrairement aux ARNt canoniques qui ont invariablement 7 paires de bases dans la tige acceptrice. De plus, la tige D a 2 paires de bases additionnelles qui remplacent les interactions tertiaires universelles 8-14, 15-48 qui sont absentes chez l'ARNtSec. D'autre part, la longueur de la boucle variable de l'ARNtSec est plus longue que la majorité des ARNt de type II. Dans ce mémoire, on se concentre sur la région de la boucle variable de l'ARNtSec . La recherche consiste à distinguer les paires de bases de la boucle variable qui sont essentielles à la biosynthèse et l’insertion de la sélénocystéine. De plus, on regarde si la paire de base additionnelle de la tige acceptrice de l'ARNtSec (procaryote) est essentielle pour l'insertion de la sélénocystéine. Pour répondre à ces questions, on a utilisé l'approche expérimentale Évolution Instantanée qui consiste au criblage in vivo d'ARNtSec fonctionnels chez E. coli. Dans ce travail, on montre que l'insertion de la sélénocystéine ne nécessite pas une spécificité de la longueur ou de la séquence de l'ARNtSec. On montre aussi que ni la longueur de la tige acceptrice ou du domaine tige acceptrice/tige T n'est essentielle pour avoir un ARNtSec fonctionnel.

The study of RNA tertiary interactions in tRNA structure and function

Le rôle des deux paires de bases universelles inverse Hoogsteen U : A ( RHUAs ) présentent chez les ARNt standards , une dans la boucle T et l'autre dans le noyau de la forme en L , a été étudiée. Pour chacun des RHUAs , un criblage génétique spécialisé in vivo chez les bactéries , le système suppresseur ambre ( pour l'étude de la RHUA dans la boucle T ) et le système d'ARNt de la sélénocystéine ( tRNASec ) ( pour l'étude de la RHUA dans le noyau ) , ont été utilisé pour générer des variants fonctionnels à partir de multiples librairies combinatoires . Ces variants ont ensuite été séquencé et soumis à une analyse systématique qui comprend la modélisation informatique et un type d'analyse phylogénétique. Les résultats du système suppresseur ambre ont montré un ensemble de variants fonctionnels qui ne nécessitent pas le motif RHUA dans la boucle T et qui ont remplacé la méthode standard de l'interaction entre les boucles D et T avec une double hélice interboucle , ILDH . D'autres études ont abouti à la détermination d'un modèle In silico de l'alternative à la norme standard de la boucle T, sous le nom de type III . Les résultats du système tRNASec ont révélé que pour cette ARNt exceptionnel, l'absence de RHUA ( dans le noyau ) assure une flexibilité accrue qui est spécifiquement nécessaire pour la fonction de tRNASec . Ainsi, les ARNt standards , à la différence de tRNASec , avec la présence universelle de RHUA dans le noyau , a été naturellement sélectionnée pour être rigide . Pris ensemble, la RHUA joue un rôle essentiel dans la stabilisation des interactions tertiaires.

Evolution and functional characterisation of the IR chemosensory receptors in the Drosophila larval gustatory system

Chemosensory receptor gene families encode divergent proteins capable of detecting a huge diversity of environmental stimuli that are constantly changing over evolutionary time as organisms adapt to distinct ecological niches. While olfaction is dedicated to the detection of volatile compounds, taste is key to assess food quality for nutritional value and presence of toxic substances. The sense of taste also provides initial signals to mediate endocrine regulation of appetite and food metabolism and plays a role in kin recognition. The fruit fly Drosophila melanogaster is a very good model for studying smell and taste because these senses are very important in insects and because a broad variety of genetic tools are available in Drosophila. Recently, a family of 66 chemosensory receptors, the Ionotropic Receptors (IRs) was described in fruit flies. IRs are distantly related to ionotropic glutamate receptors (iGluRs), but their evolutionary origin from these synaptic receptors is unclear. While 16 IRs are expressed in the olfactory system, nothing is known about the other members of this repertoire. In this thesis, I describe bioinformatic, expression and functional analyses of the IRs aimed at understanding how these receptors have evolved, and at characterising the role of the non-olfactory IRs. I show that these have emerged at the basis of the protostome lineage and probably have acquired their sensory function very early. Moreover, although several IRs are conserved across insects, there are rapid and dramatic changes in the size and divergence of IR repertoires across species. I then performed a comprehensive analysis of IR expression in the larva of Drosophila melanogaster, which is a good model to study taste and feeding mechanisms as it spends most of its time eating or foraging. I found that most of the divergent members of the IR repertoire are expressed in both peripheral and internal gustatory neurons, suggesting that these are involved in taste perception. Finally, through the establishment of a new neurophysiological assay in larvae, I identified for the first time subsets of IR neurons that preferentially detect sugars and amino acids, indicating that IRs might be involved in sensing these compounds. Together, my results indicate that IRs are an evolutionarily dynamic and functionally versatile family of receptors. In contrast to the olfactory IRs that are well-conserved, gustatory IRs are rapidly evolving species-specific receptors that are likely to be involved in detecting a wide variety of tastants. - La plupart des animaux possèdent de grandes familles de récepteurs chimiosensoriels dont la fonction est de détecter l'immense diversité de composés chimiques présents dans l'environnement. Ces récepteurs évoluent en même temps que les organismes s'adaptent à leur écosystème. Il existe deux manières de percevoir ces signaux chimiques : l'olfaction et le goût. Alors que le système olfactif perçoit les composés volatiles, le sens du goût permet d'évaluer, par contact, la qualité de la nourriture, de détecter des substances toxiques et de réguler l'appétit et le métabolisme. L'un des organismes modèles les plus pertinents pour étudier le sens du goût est le stade larvaire de la mouche du vinaigre Drosophila melanogaster. En effet, la principale fonction du stade larvaire est de trouver de la nourriture et de manger. De plus, il est possible d'utiliser tous les outils génétiques développés chez la drosophile. Récemment, une nouvelle famille de 66 récepteurs chimiosensoriels appelés Récepteurs Ionotropiques (IRs) a été découverte chez la drosophile. Bien que leur orogine soit peu claire, ces récepteurs sont similaires aux récepteurs ionotropiques glutamatergiques impliqués dans la transmission synaptique. 16 IRs sont exprimés dans le système olfactif de la mouche adulte, mais pour l'instant on ne connaît rien des autres membres de cette famille. Durant ma thèse, j'ai effectué des recherches sur l'évolution de ces récepteurs ainsi que sur l'expression et la fonction des IRs non olfactifs. Je démontre que les IRs sont apparus chez l'ancêtre commun des protostomiens et ont probablement acquis leur fonction sensorielle très rapidement. De plus, bien qu'un certain nombre d'IRs olfactifs soient conservés chez les insectes, d'importantes variations dans la taille et la divergence des répertoires d'IRs entre les espèces ont été constatées. J'ai également découvert qu'un grand nombre d'IRs non olfactifs sont exprimés dans différents organes gustatifs, ce qui leur confère probablement une fonction dans la perception des goûts. Finalement, pour la première fois, des neurones exprimant des IRs ont été identifiés pour leur fonction dans la perception de sucres et d'acides aminés chez la larve. Mes résultats présentent les IRs comme une famille très dynamique, aux fonctions très variées, qui joue un rôle tant dans l'odorat que dans le goût, et dont la fonction est restée importante tout au long de l'évolution. De plus, l'identification de neurones spécialisés dans la perception de certains composés permettra l'étude des circuits neuronaux impliqués dans le traitement de ces informations.

Evolution of primordial black holes in a radiation and phantom energy environment

In this work we extend previous work on the evolution of a primordial black hole (PBH) to address the presence of a dark energy component with a super-negative equation of state as a background, investigating the competition between the radiation accretion, the Hawking evaporation and the phantom accretion, the latter two causing a decrease on black hole mass. It is found that there is an instant during the matter-dominated era after which the radiation accretion becomes negligible compared to the phantom accretion. The Hawking evaporation may become important again depending on a mass threshold. The evaporation of PBHs is quite modified at late times by these effects, but only if the generalized second law of thermodynamics is violated.

Prognostic Value Of Dna And Mrna E6/e7 Of Human Papillomavirus In The Evolution Of Cervical Intraepithelial Neoplasia Grade 2.

This study aimed at evaluating whether human papillomavirus (HPV) groups and E6/E7 mRNA of HPV 16, 18, 31, 33, and 45 are prognostic of cervical intraepithelial neoplasia (CIN) 2 outcome in women with a cervical smear showing a low-grade squamous intraepithelial lesion (LSIL). This cohort study included women with biopsy-confirmed CIN 2 who were followed up for 12 months, with cervical smear and colposcopy performed every three months. Women with a negative or low-risk HPV status showed 100% CIN 2 regression. The CIN 2 regression rates at the 12-month follow-up were 69.4% for women with alpha-9 HPV versus 91.7% for other HPV species or HPV-negative status (P < 0.05). For women with HPV 16, the CIN 2 regression rate at the 12-month follow-up was 61.4% versus 89.5% for other HPV types or HPV-negative status (P < 0.05). The CIN 2 regression rate was 68.3% for women who tested positive for HPV E6/E7 mRNA versus 82.0% for the negative results, but this difference was not statistically significant. The expectant management for women with biopsy-confirmed CIN 2 and previous cytological tests showing LSIL exhibited a very high rate of spontaneous regression. HPV 16 is associated with a higher CIN 2 progression rate than other HPV infections. HPV E6/E7 mRNA is not a prognostic marker of the CIN 2 clinical outcome, although this analysis cannot be considered conclusive. Given the small sample size, this study could be considered a pilot for future larger studies on the role of predictive markers of CIN 2 evolution.

Effects Of Therapeutic Approach On The Neonatal Evolution Of Very Low Birth Weight Infants With Patent Ductus Arteriosus.

To analyze the effects of treatment approach on the outcomes of newborns (birth weight [BW] < 1,000 g) with patent ductus arteriosus (PDA), from the Brazilian Neonatal Research Network (BNRN) on: death, bronchopulmonary dysplasia (BPD), severe intraventricular hemorrhage (IVH III/IV), retinopathy of prematurity requiring surgical (ROPsur), necrotizing enterocolitis requiring surgery (NECsur), and death/BPD. This was a multicentric, cohort study, retrospective data collection, including newborns (BW < 1000 g) with gestational age (GA) < 33 weeks and echocardiographic diagnosis of PDA, from 16 neonatal units of the BNRN from January 1, 2010 to Dec 31, 2011. Newborns who died or were transferred until the third day of life, and those with presence of congenital malformation or infection were excluded. Groups: G1 - conservative approach (without treatment), G2 - pharmacologic (indomethacin or ibuprofen), G3 - surgical ligation (independent of previous treatment). Factors analyzed: antenatal corticosteroid, cesarean section, BW, GA, 5 min. Apgar score < 4, male gender, Score for Neonatal Acute Physiology Perinatal Extension (SNAPPE II), respiratory distress syndrome (RDS), late sepsis (LS), mechanical ventilation (MV), surfactant (< 2 h of life), and time of MV. death, O2 dependence at 36 weeks (BPD36wks), IVH III/IV, ROPsur, NECsur, and death/BPD36wks. Student's t-test, chi-squared test, or Fisher's exact test; Odds ratio (95% CI); logistic binary regression and backward stepwise multiple regression. Software: MedCalc (Medical Calculator) software, version 12.1.4.0. p-values < 0.05 were considered statistically significant. 1,097 newborns were selected and 494 newborns were included: G1 - 187 (37.8%), G2 - 205 (41.5%), and G3 - 102 (20.6%). The highest mortality was observed in G1 (51.3%) and the lowest in G3 (14.7%). The highest frequencies of BPD36wks (70.6%) and ROPsur were observed in G3 (23.5%). The lowest occurrence of death/BPD36wks occurred in G2 (58.0%). Pharmacological (OR 0.29; 95% CI: 0.14-0.62) and conservative (OR 0.34; 95% CI: 0.14-0.79) treatments were protective for the outcome death/BPD36wks. The conservative approach of PDA was associated to high mortality, the surgical approach to the occurrence of BPD36wks and ROPsur, and the pharmacological treatment was protective for the outcome death/BPD36wks.

Evolution Of The Deaths Registry System In Brazil: Associations With Changes In The Mortality Profile, Under-registration Of Death Counts, And Ill-defined Causes Of Death.

This paper examines the spatial pattern of ill-defined causes of death across Brazilian regions, and its relationship with the evolution of completeness of the deaths registry and changes in the mortality age profile. We make use of the Brazilian Health Informatics Department mortality database and population censuses from 1980 to 2010. We applied demographic methods to evaluate the quality of mortality data for 137 small areas and correct for under-registration of death counts when necessary. The second part of the analysis uses linear regression models to investigate the relationship between, on the one hand, changes in death counts coverage and age profile of mortality, and on the other, changes in the reporting of ill-defined causes of death. The completeness of death counts coverage increases from about 80% in 1980-1991 to over 95% in 2000-2010 at the same time the percentage of ill-defined causes of deaths reduced about 53% in the country. The analysis suggests that the government's efforts to improve data quality are proving successful, and they will allow for a better understanding of the dynamics of health and the mortality transition.

Digestive Peptidase Evolution In Holometabolous Insects Led To A Divergent Group Of Enzymes In Lepidoptera.

Trypsins and chymotrypsins are well-studied serine peptidases that cleave peptide bonds at the carboxyl side of basic and hydrophobic l-amino acids, respectively. These enzymes are largely responsible for the digestion of proteins. Three primary processes regulate the activity of these peptidases: secretion, precursor (zymogen) activation and substrate-binding site recognition. Here, we present a detailed phylogenetic analysis of trypsins and chymotrypsins in three orders of holometabolous insects and reveal divergent characteristics of Lepidoptera enzymes in comparison with those of Coleoptera and Diptera. In particular, trypsin subsite S1 was more hydrophilic in Lepidoptera than in Coleoptera and Diptera, whereas subsites S2-S4 were more hydrophobic, suggesting different substrate preferences. Furthermore, Lepidoptera displayed a lineage-specific trypsin group belonging only to the Noctuidae family. Evidence for facilitated trypsin auto-activation events were also observed in all the insect orders studied, with the characteristic zymogen activation motif complementary to the trypsin active site. In contrast, insect chymotrypsins did not seem to have a peculiar evolutionary history with respect to their mammal counterparts. Overall, our findings suggest that the need for fast digestion allowed holometabolous insects to evolve divergent groups of peptidases with high auto-activation rates, and highlight that the evolution of trypsins led to a most diverse group of enzymes in Lepidoptera.

Sensory evaluation of black instant coffee beverage with some volatile compounds present in aromatic oil from roasted coffee

The oil obtained from Brazilian roasted coffee by supercritical CO2 extraction shows considerable aromatic properties, mainly composed by five aromatic compounds, 2-methylpyrazine; 2-furfurylalcohol, 2,5-dimethylpyrazine; γ-butyrolactone and 2-furfurylacetate. Sensory analyses were used to verify the influence of a mixture of these important classes of aromatic coffee compounds (pyrazines, furans and lactones) and of the roasted coffee aromatic oil on the coffee aroma and flavour of black instant freeze and spray-dried coffee beverages. In the acceptance evaluation of the aroma, the samples prepared with freeze-dried instant coffee without the mixture of volatile compounds (sample 4) were not significantly different from the freeze-dried instant coffee in which the aromatic coffee oil was added (sample 5) and from the sample prepared with freeze-dried coffee in which the mixture of the five volatile was added (sample 3), coincidentally from the same drying process. Therefore, sample (3) did not differ from samples prepared with spray dried instant coffee without (sample 1) and to which (sample 2) the mixture of volatile was added. Therefore, with respect to this attribute, the addition of this mixture did not interfere in this drink acceptance. Taking into consideration the flavor, samples prepared with freeze-dried instant coffee in which the aromatic coffee oil was added (5) and the samples with (3) and without (4) the mixture of the five volatile was added did not differ significantly, however sample (4) did not differ from samples (1) and (2). Regarding this attribute, the addition of the aromatic oil of roasted coffee or a mixture of volatile in samples of freeze-dried instant coffee had a better acceptance than those dried by spray dryer (1) and (2). Thus, the enrichment of drinks with the aromatic oil of roasted coffee, or even with the mixture of the five components did not influence the consumer acceptance with respect to the aroma, but exerts influence with respect to flavour.

Time evolution of the South Atlantic Magnetic Anomaly

The South Atlantic Magnetic Anomaly (SAMA) is one of the most outstanding anomalies of the geomagnetic field. The SAMA secular variation was obtained and compared to the evolution of other anomalies using spherical harmonic field models for the 1590-2005 period. An analysis of data from four South American observatories shows how this large scale anomaly affected their measurements. Since SAMA is a low total field anomaly, the field was separated into its nondipolar, quadrupolar and octupolar parts. The time evolution of the non-dipole/total, quadrupolar/total and octupolar/total field ratios yielded increasingly high values for the South Atlantic since 1750. The SAMA evolution is compared to the evolution of other large scale surface geomagnetic features like the North and the South Pole and the Siberia High, and this comparison shows the intensity equilibrium between these anomalies in both hemispheres. The analysis of non-dipole fields in historical period suggests that SAMA is governed by (i) quadrupolar field for drift, and (ii) quadrupolar and octupolar fields for intensity and area of influence. Furthermore, our study reinforces the possibility that SAMA may be related to reverse fluxes in the outer core under the South Atlantic region.

Sob o signo de Darwin? Sobre o mau uso de uma quimera

No ano de 2003 Francisco de Oliveria publicou um artigo intitulado "O Ornitorrinco" no qual fez considerações críticas sobre a conjectura politico-social daquele momento histórico. Tal artigo é permeado por um paralelo entre o evolucionismo darwinista e a visão do autor sobre a sociedade brasileira contemporânea. Entretanto, ao fazer tal analogia ele incorre numa série de equívocos teóricos sobre a teoria evolucionista. Tais equívocos consistem, em grande parte, numa substuição indevida entre aquilo que ficou conhecido como Darwinismo Social e a teoria neodarwinista como entendida pelos seus atuais proponentes. O presente trabalho identifica estes equívocos e os contextualiza dentro da teoria neodarwiniana. Além disso, fazemos um recorte histórico do processo de formação do pensamento evolucionista para enfatizar que a associação entre biologia e darwinismo social é mais complexa do que geralmente se assume.

The Southern Brazilian shelf: general characteristics, quaternary evolution and sediment distribution

Extending from latitude 34ºS to 22ºS the Southern Brazilian shelf constitutes the only part of the Brazilian shelf with a subtropical to temperate environment. The studies on the different geological aspects of the area began in the 1960's and have recently been reassessed after studies related to the determination of the Economic Exclusive Zone. In terms of morphology, the Southern Brazilian shelf may be divided into three sectors, the São Paulo Bight, the Florianópolis-Mostardas Sector and the Rio Grande Cone, characterized by conspicuous differences in terms of geological determining factors, bathymetry, declivities and the presence of canyons and channels. Despite the existence of hundreds of radiocarbon datings the sea level changes curve of southern Brazil during the Last Glacial Cycle is still a matter of debate. A recent controversy on the Middle and late Holocene sea level changes curve raised the question of the amplitude of the oscillations which occurred in the period. Also, a few but relatively consistent radiocarbon datings suggest the occurrence of a high sea level during Isotope Stage 3. In terms of sedimentary cover the Southern Brazilian shelf exhibits a very strong hydrodynamic control, both latitudinal and bathymetrical. The sector southward from 25ºS is characterized by the influence of the plume of water carrying sediments originating from the Río de La Plata. Actually its presence is conspicuous up to 28ºS, with the area between this latitude and 25ºS constituting a transitional zone. In terms of bathymetry the outer shelf is marked by the "floor-polisher" effect of the Brazil Current, which is responsible for the maintenance of a relict facies in areas deeper than 100 meters.

Evolution of magnetic properties and crystallographic texture in electrical steel with large plastic deformation

Deformation leads to a hardening of steel due to an increase in the density of dislocations and a reduction in their mobility, giving rise to a state of elevated residual stresses in the crystal lattice. In the microstructure, one observes an increase in the contribution of crystalline orientations which are unfavorable to the magnetization, as seen, for example, by a decrease in B(50), the magnetic flux density at a field of 50 A/cm. The present study was carried out with longitudinal strips of fully processed non-oriented (NO) electrical steel, with deformations up to 70% resulting from cold rolling in the longitudinal direction. With increasing plastic deformation, the value of B(50) gradually decreases until it reaches a minimum value, where it remains even for larger deformations. On the other hand, the coercive field H(c) continually increases. Magnetometry results and electron backscatter diffraction results are compared and discussed. (C) 2011 American Institute of Physics. [doi: 10.1063/1.3560895]

The four hexamerin genes in the honey bee: structure, molecular evolution and function deduced from expression patterns in queens, workers and drones

Background: Hexamerins are hemocyanin-derived proteins that have lost the ability to bind copper ions and transport oxygen; instead, they became storage proteins. The current study aimed to broaden our knowledge on the hexamerin genes found in the honey bee genome by exploring their structural characteristics, expression profiles, evolution, and functions in the life cycle of workers, drones and queens. Results: The hexamerin genes of the honey bee (hex 70a, hex 70b, hex 70c and hex 110) diverge considerably in structure, so that the overall amino acid identity shared among their deduced protein subunits varies from 30 to 42%. Bioinformatics search for motifs in the respective upstream control regions (UCRs) revealed six overrepresented motifs including a potential binding site for Ultraspiracle (Usp), a target of juvenile hormone (JH). The expression of these genes was induced by topical application of JH on worker larvae. The four genes are highly transcribed by the larval fat body, although with significant differences in transcript levels, but only hex 110 and hex 70a are re-induced in the adult fat body in a caste-and sex-specific fashion, workers showing the highest expression. Transcripts for hex 110, hex 70a and hex70b were detected in developing ovaries and testes, and hex 110 was highly transcribed in the ovaries of egg-laying queens. A phylogenetic analysis revealed that HEX 110 is located at the most basal position among the holometabola hexamerins, and like HEX 70a and HEX 70c, it shares potential orthology relationship with hexamerins from other hymenopteran species. Conclusions: Striking differences were found in the structure and developmental expression of the four hexamerin genes in the honey bee. The presence of a potential binding site for Usp in the respective 5' UCRs, and the results of experiments on JH level manipulation in vivo support the hypothesis of regulation by JH. Transcript levels and patterns in the fat body and gonads suggest that, in addition to their primary role in supplying amino acids for metamorphosis, hexamerins serve as storage proteins for gonad development, egg production, and to support foraging activity. A phylogenetic analysis including the four deduced hexamerins and related proteins revealed a complex pattern of evolution, with independent radiation in insect orders.

A molecular phylogeny and the evolution of nest architecture and behavior in Trigona s.s. (Hymenoptera : Apidae : Meliponini)

Stingless bees exhibit extraordinary variation in nest architecture within and among species. To test for phylogenetic association of behavioral traits for species of the Neotropical stingless bee genus Trigona s.s., a phylogenetic hypothesis was generated by combining sequence data of 24 taxa from one mitochondrial (16S rRNA) and four nuclear gene fragments (long-wavelength rhodopsin copy 1 (opsin), elongation factor-1 alpha copy F2, arginine kinase, and 28S rRNA). Fifteen characteristics of the nest architecture were coded and tested for phylogenetic association. Several characters have significant phylogenetic signal, including type of nesting substrate, nest construction material, and hemipterophily, the tending of hemipteroid insects in exchange for sugar excretions. Phylogenetic independent habits encountered in Trigona s.s. include coprophily and necrophagy.