Caractérisation moléculaire et fonctionnelle de la protéine DYW1 dans le complexe d'édition chloroplastique d'Arabidopsis thaliana

Dans les organites des plantes, l édition de l ARN consiste majoritairement en une désamination de cytidines à des sites spécifiques de l ARNm. Trente-quatre sites d édition ont été découverts dans les transcrits chloroplastiques d Arabidopsis thaliana et plus de 500 dans les transcrits mitochondriaux. Depuis 2005, beaucoup de facteurs d édition ont été trouvés. La majorité de ces protéines appartiennent à la famille des PentatricoPeptide Repeat (PPR). Parmi ces PPR, certaines contiennent un domaine DYW possédant de faibles similarités avec les cytidines désaminases (CDA), alors que d autres en sont dénuées, générant un doute sur le fait qu il ait une activité CDA. Le gène At1g47580 (DYW1) code une protéine unique chez Arabidopsis thaliana contenant seulement un domaine DYW. Il a été proposé que DYW1 puisse interagir avec les PPR ne contenant pas de domaine DYW, pour former un hétérodimère, capable d éditer spécifiquement un site. En accord avec cette hypothèse, nous avons montré que DYW1 agissait sur le même site d édition que CRR4, une PPR sans domaine DYW, et que ces protéines interagissaient in vivo. De plus, nous avons montré que DYW1 remplaçait les parties manquantes de CRR4 pour l édition. Pour obtenir plus d informations sur la fonction du domaine DYW, des mutations ont été introduites dans DYW1. Nous avons montré que la signature CDA dans les protéines DYW était essentielle à l édition de l ARN ainsi qu à l interaction avec les ions zinc. Les données sont en accord avec l hypothèse d une activité CDA dans le domaine DYW. Cependant, aucune activité CDA n a pu être mise à jour in vitro. Il est vraisemblable qu au moins un cofacteur doive encore être identifié.In plant organelles, RNA editing mostly takes the form of conversions of cytidines to uridines at specific sites in mRNAs. Thirty-four editing sites have been found in Arabidopsis thaliana chloroplast transcripts and more than 500 sites in mitochondrial transcripts. Since 2005, lots of proteins have been found to act as RNA editing factors. Most of these proteins belong to the PentatricoPeptide Repeat (PPR) family. Amongst these PPR, some contain a DYW domain with weak similarity to cytidine deaminases (CDA), whilst others lack such a domain, creating doubts about whether this domain is required for editing. The gene At1g47580 (named DYW1) encodes a protein in Arabidopsis thaliana that contains only a DYW domain. Our initial hypothesis was that DYW1 might interact with PPR proteins that lack a DYW domain, in order to form a heterodimer, able to perform site-specific editing. In accordance with this hypothesis, we discovered that DYW1 is involved in editing the same site as CRR4, a PPR lacking a DYW domain, and that these two proteins interact together in vivo. Moreover, we showed that DYW1 replaces all the missing parts of CRR4 for editing. So, other partners need to be hypothesized for other DYW-lacking editing factors if this hypothesis is to be generalized. The highly conserved residues making up the CDA signature in DYW proteins were found to be essential for RNA editing and are also required for zinc binding, which is a known characteristic of CDAs. All the data so far are consistent with the DYW domain being (part of) a CDA activity; nevertheless, no CDA activity could be detected in vitro. It is likely that at least one required cofactor remains to be identified.EVRY-Bib. électronique (912289901) / SudocSudocFranceF

Individual Differences in Well-Being in Older Breast Cancer Survivors

Older women who survive breast cancer may differ significantly in their long-term well-being. Using a risk and protective factors model, we studied predictors of well-being in 127 women age 70 and above with a history of at least one year\u27s survival of breast cancer. Mean post-cancer survivorship was 5.1 years. Using life satisfaction, depression and general health perceptions as outcome variables, we assessed whether demographic variables, cancer-related variables, health status and psychosocial resources predicted variability in well-being using correlational and hierarchical regression analyses. Higher age predicted increased depression but was not associated with life satisfaction or general health perceptions. Cancer-related variables, including duration of survival, and type of cancer treatment, were not significantly associated with survivors\u27 well-being. Poorer health status was associated with poorer well-being in all three dependent variables. After controlling for demographics, cancer-related variables, and health status, higher levels of psychosocial resources including optimism, mastery, spirituality and social support predicted better outcome in all three dependent variables. While many older women survive breast cancer without severe sequelae, there is considerable variability in their well-being after survivorship. Successful intervention with older breast cancer survivors might include greater attention not only to cancer-specific concerns, but also attention to geriatric syndromes and functional impairment, and enhancement of protective psychosocial resources

Identification des protéines PPR impliquées dans l'épissage des ARN messagers dans les chloroplastes et les mitochondries chez Arabidopsis Thaliana

Le mécanisme d épissage dans les organites est décrit comme étant l ancêtre du spliceosome nucléaire. Cependant même si les protéines composant ce dernier sont bien connues, seulement quelques facteurs d épissage ont été identifiés et caractérisés dans les chloroplastes et les mitochondries. Beaucoup de protéines ayant la faculté de se lier à l ARN ont acquis des fonctions dans l épissage, en effet un certain nombre de protéines sans véritable lien ont un rôle essentiel, avec différents degrés de spécificité dans l épissage de la plupart des introns chloroplastiques chez les plantes. La plus grande famille de protéines se liant à l ARN est la famille des protéines à domaines pentatricopetide repeat (PPR). Ces protéines sont impliquées dans la plupart des processus post-transcriptionnels dans les organites. En 2006, parmi les centaines de protéines PPR décrites chez les plantes, seulement une PPR avait été décrite comme nécessaire à l épissage d un intron. Ainsi, PPR4 est absolument et spécifiquement nécessaire pour l épissage en trans de l intron 1 de rps12 dans les plastes (Schmitz-Linneweber et al., 2006), suggérant que d autres protéines PPR pourraient être impliquées dans l épissage des ARN des organites. Le sujet de cette thèse porte sur la caractérisation d autres protéines PPR impliquées dans ce processus. En utilisant des approches de génétique inverse et des outils mis en place dans le cadre de la thèse afin de détecter des défauts d épissage par PCR quantitative, sept nouvelles PPRs impliquées dans l épissage d un certain nombre d introns dans les plastes et les mitochondries ont pu être caractérisées. Dans l optique de rechercher si des protéines PPR, impliquées dans l épissage mais aussi dans l édition des ARN, interagissent avec d autres protéines, des approches de TAP-TAG ont été réalisées et sont également présentées dans ce manuscrit. L identification de partenaires protéiques pour 3 PPRs impliquées, nous a ainsi permis de redessiner nos modèles et d émettre de nouvelles hypothèses. Enfin, une dernière partie est consacrée à la découverte d isoformes d épissage pour des gènes PPR sans introns. Phénomène qui permettrait de réguler l expression des gènes PPR, et/ou d augmenter la diversité des protéines PPR.The RNA splicing mechanism in organelles is described to be ancestral to that of the nuclear spliceosome. However, whereas this last complex is well known, only very few splicing factors have been identified and characterized in chloroplasts and mitochondria. Many RNA binding proteins have acquired roles in RNA splicing, and indeed a variety of often unrelated RNA binding proteins have essential functions in splicing of many plastid introns in plants, with varying degrees of specificity. The largest family of RNA binding proteins in plant organelles is the pentatricopeptide repeat (PPR) family. PPR proteins are involved in diverse post-transcriptional processes in organelles. In 2006, among hundreds of higher plant proteins of this family, only one was described as being required for a splicing event - PPR4 was shown to be absolutely and specifically required for the trans-splicing of the rps12 intron 1 in plastids (Schmitz-Linneweber et al., 2006). The main purpose of this PhD thesis was to characterize other PPR proteins involved in this process. By using a reverse genetics approach and by developing tools for the detection of splicing defects, seven new PPR proteins involved in RNA splicing of a subset of chloroplast or mitochondria introns have been characterized. In parallel, in order to characterize proteins involved in PPR-containing complexes, a TAP-TAG approach has been carried out on a few PPR proteins involved in splicing or editing of organellar RNA. The identification of partner proteins of 3 PPR proteins allows us to draw new mechanistic models and new hypotheses. Finally, the final part of the manuscript describes the discovery of splicing isoforms of PPR-encoding mRNAs. Alternative splicing may be involved in regulation of PPR gene expression and/or in increasing the diversity of the PPR protein family.EVRY-Bib. électronique (912289901) / SudocSudocFranceF

Variables associated with successful outcome after anterior cruciate ligament reconstruction in recreational athletes: A prospective cohort study

Background: Anterior cruciate ligament (ACL) injury and subsequent reconstruction is common and has a profound effect on health-related quality of life. There is currently limited understanding as to which variables are associated with a successful outcome post-ACL reconstruction (ACLR) in recreational athletes. Purpose: Explore the association between both patient-reported and performance-based measures, and successful outcome, post-ACLR in recreational athletes. Procedures: We sought to recruit recreational athletes within one month of a primary-ACLR for a prospective cohort study. A dichotomised patient specific functional scale of ≥9 points determined a successful outcome at nine-months post-operative. Secondary patient-reported and performance-based data were collected at baseline, three-, six-, and nine-months post-operative. The association between secondary data and the primary outcome was determined using binomial logistic regression, expressed using odds ratio (OR) and 95% confidence intervals (CI). Main results: 90 participants were recruited (males: 58, females: 32, mean age 32.8 years [±7.9], mean height 173.5 [±10.0], mean body mass 74.0 kg [±15.8]), 87 consented to baseline measures. 47 participants completed full data collection and 21 (45%) reported a successful outcome. Higher knee osteoarthritis outcome score (OR range 1.07–1.12) and anterior cruciate ligament quality of life (ACL-QoL) scores (OR range 1.06–1.10) were associated with a successful outcome post-ACLR at various timepoints. Conclusions: Patient-reported, rather than performance-based, measures were associated with successful outcome nine-months post-ACLR in recreational athletes. Both patient-reported and performance-based characteristics are advocated to guide optimal return to function in clinical practice

A model of membrane contraction predicting initiation and completion of bacterial cell division

Bacterial cell division involves a complex and dynamic sequence of events whereby polymers of the protein FtsZ assemble at the division plane and rearrange to achieve the goal of contracting the cell membrane at the site of cell division, thus dividing the parent cell into two daughter cells. We present a mathematical model (which we refer to as CAM-FF: Critical Accumulation of Membrane-bound FtsZ Fibres) of the assembly of the contractile ring in terms of the accumulation of short linear polymers of FtsZ that associate and dissociate from the cell membrane. In prokaryotes, the biochemical function of FtsZ is thought to underpin the assembly and at least the initial kinetic force of ring contraction. Our model extends earlier work of Surovtsev et al. [PLoS Comput. Biol., 2008, 4, e1000102] by adding (i) the kinetics of FtsZ accumulation on cell membrane anchor proteins and (ii) the physical forces required to deform the cell against its surface tension. Moreover, we provide a more rigorous treatment of intracellular diffusion and we revise some of the model parameter values in light of the experimental evidence now available. We derive a critical contraction parameter which links the chemical population dynamics of membrane-bound FtsZ molecules to the force of contraction. Using this parameter as a tool to predict the ability of the cell to initiate division, we are able to predict the division outcome in cells depleted of key FtsZ-binding proteins

Using stable isotopes of hydrogen to quantify biogenic and thermogenic atmospheric methane sources: A case study from the Colorado Front Range

Global atmospheric concentrations of methane (CH4), a powerful greenhouse gas, are increasing, but because there are many natural and anthropogenic sources of CH4, it is difficult to assess which sources may be increasing in magnitude. Here we present a data set of δ2H-CH4 measurements of individual sources and air in the Colorado Front Range, USA. We show that δ2H-CH4, but not δ13C, signatures are consistent in air sampled downwind of landfills, cattle feedlots, and oil and gas wells in the region. Applying these source signatures to air in ground and aircraft samples indicates that at least 50% of CH4 emitted in the region is biogenic, perhaps because regulatory restrictions on leaking oil and natural gas wells are helping to reduce this source of CH4. Source apportionment tracers such as δ2H may help close the gap between CH4 observations and inventories, which may underestimate biogenic as well as thermogenic sources

Drivers and Challenges in Raising the Achievement of Pupils from Bangladeshi, Somali and Turkish Backgrounds

In this report we examine why attainment rates amongst Bangladeshi pupils have significantly improved over recent years while Turkish and Somali pupils continue to perform below the national average. We explore the experiences of pupils within the education system, the characteristics and policies of their schools, the support received from the local authorities and the impact of parental involvement and parents’ attitudes to school. We also suggest the factors and school strategies likely to have contributed to improved outcomes for underperforming ethnic minority pupils. Evidence will include examples of good practice in schools with high performing pupils from these groups. Wherever possible we identify factors that are specific to the three groups in this study, and in particular the factors that explain increasing attainment rates amongst Bangladeshi pupils

Concordance for clonal hematopoiesis is limited in elderly twins.

Although acquisition of leukemia-associated somatic mutations by 1 or more hematopoietic stem cells is inevitable with advancing age, its consequences are highly variable, ranging from clinically silent clonal hematopoiesis (CH) to leukemic progression. To investigate the influence of heritable factors on CH, we performed deep targeted sequencing of blood DNA from 52 monozygotic (MZ) and 27 dizygotic (DZ) twin pairs (aged 70-99 years). Using this highly sensitive approach, we identified CH (variant allele frequency ≥0.5%) in 62% of individuals. We did not observe higher concordance for CH within MZ twin pairs as compared with that within DZ twin pairs, or to that expected by chance. However, we did identify 2 MZ pairs in which both twins harbored identical rare somatic mutations, suggesting a shared cell of origin. Finally, in 3 MZ twin pairs harboring mutations in the same driver genes, serial blood samples taken 4 to 5 years apart showed substantial twin-to-twin variability in clonal trajectories. Our findings propose that the inherited genome does not exert a dominant influence on the behavior of adult CH and provide evidence that CH mutations may be acquired in utero