Quantitative TCR:pMHC Dissociation Rate Assessment by NTAmers Reveals Antimelanoma T Cell Repertoires Enriched for High Functional Competence.

Experimental models demonstrated that therapeutic induction of CD8 T cell responses may offer protection against tumors or infectious diseases providing that T cells have sufficiently high TCR/CD8:pMHC avidity for efficient Ag recognition and consequently strong immune functions. However, comprehensive characterization of TCR/CD8:pMHC avidity in clinically relevant situations has remained elusive. In this study, using the novel NTA-His tag-containing multimer technology, we quantified the TCR:pMHC dissociation rates (koff) of tumor-specific vaccine-induced CD8 T cell clones (n = 139) derived from seven melanoma patients vaccinated with IFA, CpG, and the native/EAA or analog/ELA Melan-A(MART-1)(26-35) peptide, binding with low or high affinity to MHC, respectively. We observed substantial correlations between koff and Ca(2+) mobilization (p = 0.016) and target cell recognition (p < 0.0001), with the latter independently of the T cell differentiation state. Our strategy was successful in demonstrating that the type of peptide impacted on TCR/CD8:pMHC avidity, as tumor-reactive T cell clones derived from patients vaccinated with the low-affinity (native) peptide expressed slower koff rates than those derived from patients vaccinated with the high-affinity (analog) peptide (p < 0.0001). Furthermore, we observed that the low-affinity peptide promoted the selective differentiation of tumor-specific T cells bearing TCRs with high TCR/CD8:pMHC avidity (p < 0.0001). Altogether, TCR:pMHC interaction kinetics correlated strongly with T cell functions. Our study demonstrates the feasibility and usefulness of TCR/CD8:pMHC avidity assessment by NTA-His tag-containing multimers of naturally occurring polyclonal T cell responses, which represents a strong asset for the development of immunotherapy.

Optimal Exchange Rate Policy in a Growing Semi-Open Economy

This paper considers an alternative perspective to China's exchange rate policy. It studies a semi-open economy where the private sector has no access to international capital markets but the central bank has full access. Moreover, it assumes limited financial development generating a large demand for saving instruments by the private sector. The paper analyzes the optimal exchange rate policy by modeling the central bank as a Ramsey planner. Its main result is that in a growth acceleration episode it is optimal to have an initial real depreciation of the currency combined with an accumulation of reserves, which is consistent with the Chinese experience. This depreciation is followed by an appreciation in the long run. The paper also shows that the optimal exchange rate path is close to the one that would result in an economy with full capital mobility and no central bank intervention.

Postnatal β-cell maturation is associated with islet-specific microRNA changes induced by nutrient shifts at weaning.

Glucose-induced insulin secretion is an essential function of pancreatic β-cells that is partially lost in individuals affected by Type 2 diabetes. This unique property of β-cells is acquired through a poorly understood postnatal maturation process involving major modifications in gene expression programs. Here we show that β-cell maturation is associated with changes in microRNA expression induced by the nutritional transition that occurs at weaning. When mimicked in newborn islet cells, modifications in the level of specific microRNAs result in a switch in the expression of metabolic enzymes and cause the acquisition of glucose-induced insulin release. Our data suggest microRNAs have a central role in postnatal β-cell maturation and in the determination of adult functional β-cell mass. A better understanding of the events governing β-cell maturation may help understand why some individuals are predisposed to developing diabetes and could lead to new strategies for the treatment of this common metabolic disease.

Detecting patterns of species diversification in the presence of both rate shifts and mass extinctions.

BACKGROUND: Recent methodological advances allow better examination of speciation and extinction processes and patterns. A major open question is the origin of large discrepancies in species number between groups of the same age. Existing frameworks to model this diversity either focus on changes between lineages, neglecting global effects such as mass extinctions, or focus on changes over time which would affect all lineages. Yet it seems probable that both lineages differences and mass extinctions affect the same groups. RESULTS: Here we used simulations to test the performance of two widely used methods under complex scenarios of diversification. We report good performances, although with a tendency to over-predict events with increasing complexity of the scenario. CONCLUSION: Overall, we find that lineage shifts are better detected than mass extinctions. This work has significance to assess the methods currently used to estimate changes in diversification using phylogenetic trees. Our results also point toward the need to develop new models of diversification to expand our capabilities to analyse realistic and complex evolutionary scenarios.

Study of criteria influencing the success rate of DNA swabs in operational conditions: a contribution to an evidence-based approach to crime scene investigation and triage

DNA is nowadays swabbed routinely to investigate serious and volume crimes, but research remains scarce when it comes to determining the criteria that may impact the success rate of DNA swabs taken on different surfaces and situations. To investigate these criteria in fully operational conditions, DNA analysis results of 4772 swabs taken by the forensic unit of a police department in Western Switzerland over a 2.5-year period (2012-2014) in volume crime cases were considered. A representative and random sample of 1236 swab analyses was extensively examined and codified, describing several criteria such as whether the swabbing was performed at the scene or in the lab, the zone of the scene where it was performed, the kind of object or surface that was swabbed, whether the target specimen was a touch surface or a biological fluid, and whether the swab targeted a single surface or combined different surfaces. The impact of each criterion and of their combination was assessed in regard to the success rate of DNA analysis, measured through the quality of the resulting profile, and whether the profile resulted in a hit in the national database or not. Results show that some situations - such as swabs taken on door and window handles for instance - have a higher success rate than average swabs. Conversely, other situations lead to a marked decrease in the success rate, which should discourage further analyses of such swabs. Results also confirm that targeting a DNA swab on a single surface is preferable to swabbing different surfaces with the intent to aggregate cells deposited by the offender. Such results assist in predicting the chance that the analysis of a swab taken in a given situation will lead to a positive result. The study could therefore inform an evidence-based approach to decision-making at the crime scene (what to swab or not) and at the triage step (what to analyse or not), contributing thus to save resource and increase the efficiency of forensic science efforts.

Environmental control of the Pom1-dependent cell-size regulation pathway in fission yeast

Cells couple their growth and division rate in response to nutrient availability to maintain a constant size. This co-ordination happens either at the G1-S or the G2-M transition of the cell cycle. In the rod-shaped fission yeast, size regulation happens at the G2-M transition prior to mitotic commitment. Recent studies have focused on the role of the DYRK-family protein kinase Pom1, which forms gradients emanating from cell poles and inhibits the mitotic activator kinase Cdr2, present at the cell middle. Pom1 was proposed to inhibit Cdr2 until cells reached a critical size before division. However when and where Pom1 inhibits Cdr2 is not clear as medial Pom1 levels do not change during cell elongation. Here I show that Pom1 gradients are susceptible to environmental changes in glucose. Specifically, upon glucose limitation, Pom1 re-localizes from the poles to the cell sides where it delays mitosis through regulating Cdr2. This re-localization occurs due to microtubule de- stabilization and lateral catastrophes leading to transient deposition of the Pom1 gradient nucleator Tea4 along the cell cortex. As Tea4 localization to cell sides is sufficient to recruit Pom1, this explains the mechanism of Pom1 re-localization. Microtubule destabilization and consequently Tea4 and Pom1 spread depends on the activity of the cAMP-dependent Protein Kinase A (PKA/Pka1), as pka1 mutant cells have stable microtubules and retain polar Tea4 and Pom1 under limited glucose. PKA signaling negatively regulates the microtubule rescue factor CLASP/Cls1, thus reducing its ability to stabilize microtubules. Thus PKA signaling tunes CLASP activity to promote microtubule de-stabilization and Pom1 re-localization upon glucose limitation. I show that the side-localized Pom1 delays mitosis and balances the role of the mitosis promoting, mitogen-associated protein kinase (MAPK) protein Sty1. Thus Pom1 re-localization may serve to buffer cell size upon glucose limitation. -- Afin de maintenir une taille constante, les cellules régulent leur croissance ainsi que leur taux de division selon les nutriments disponibles dans le milieu. Dans la levure fissipare, cette régulation de la taille précède l'engagement mitotique et se fait à la transition entre les phases G2 à M du cycle cellulaire. Des études récentes se sont focalisées sur le rôle de la protéine Pom1, membre de la famille des DYRK kinase. Celle-ci forme un gradient provenant des pôles de la cellule et inhibe l'activateur mitotique Cdr2 présent au centre de la cellule. Le model propose que Pom1 inhibe Cdr2 jusqu'à atteindre une taille critique avant la division. Cependant quand et à quel endroit dans la cellulle Pom1 inhibe Cdr2 n'était pas clair car les niveaux médians de Pom1 ne changent pas au cours de la l'élongation des cellules. Dans cette étude, je montre que les gradients de Pom1 sont sensibles aux changements environnementaux du taux de glucose. Plus spécifiquement, en conditions limitantes de glucose, Pom1 se relocalise des pôles de la cellule pour se distribuer sur les côtés de celle-ci. Par conséquent, un délai d'entrée en mitose est observé dû à l'inhibition Cdr2 par Pom1. Cette délocalisation est due à la déstabilisation des microtubules qui va conduire à une déposition transitoire de Tea4, le nucléateur du gradient de Pom1, tout au long du cortex de la cellule. Comme la localisation de Tea4 sur les côtés de la cellule est suffisante pour recruter la protéine Pom1, ceci explique le mécanisme de relocalisation de celle-ci. La déstabilisation des microtubules et par conséquent la diffusion de Tea4 et Pom1 dépendent de l'activité de la protéine kinase A dépendante de l'AMP cyclique (PKA/Pka1). En absence de pka1, la stabilité des microtubules n'est pas affectée ce qui permet la rétention de Tea4 et Pom1 aux pôles de la cellule même en conditions limitantes de glucose. La signalisation via PKA régule négativement le facteur de sauvetage des microtubules CLASP/Cls1 et permet donc de réduire sa fonction de déstabilisation des microtubules. Ainsi la signalisation via PKA affine l'activité des CLASP pour promouvoir la déstabilisation des microtubules et la relocalisation de Pom1 en conditions limitantes de glucose. Je montre que la localisation sur les côtés retarde l'entrée en mitose et compense l'action de la protéine Sty1, connue pour être une MAPK qui induit l'entrée en mitose. Ainsi, la relocalisation de Pom1 pourrait servir à tamponner la taille de la cellule en condition limitantes de glucose. -- Various cell types in the environment such as bacterial, plant or animal cells have a distinct cellular size. Maintaining a constant cell size is important for fitness in unicellular organisms and for diverse functions in multicellular organisms. Cells regulate their size by coordinating their growth rate to their division rate. This coupling is important otherwise cells would get progressively smaller or larger after each successive cell cycle. In their natural environment cells may face fluctuations in the available nutrient supply. Thus cells have to coordinate their division rate to the variable growth rates shown under different nutrient conditions. During my PhD, I worked with a single-celled rod shaped yeast called the fission yeast. These cells are longer when the nutrient supply is abundant and shorter when the nutrient supply is scarce. A protein that senses changes in the external carbon source (glucose) is called Protein Kinase A (PKA). The rod shape of fission yeast cells is maintained thanks to a structural backbone called the cytoskeleton. One of the components of this backbone is called microtubules, which are small tube like structures spanning the length of the cell. They transport a protein called Tea4, which in turn is important for the proper localization of another protein Pom1 to the cell ends. Pom1 helps to maintain proper shape and size of these rod shaped yeast cells. My thesis work showed that upon reduction in the external nutrient (glucose) levels, microtubules become less stable and show an alteration in their organization. A significant percentage of the microtubules contact the side of the cell instead of touching only the cell tip. This leads to the spreading of the protein Pom1 away from the tips all around the cell periphery. This helps fission yeast cells to maintain the proper size required under these conditions of limited glucose supply. I further showed that the protein PKA regulates microtubule stability and organization and thus Pom1 spreading and maintenance of proper cell size. Thus my work led to the discovery of a novel pathway by which fission yeast cells maintain their size under limited supply of glucose. -- Divers types cellulaires dans l'environnement tels que les bactéries, les plantes ou les cellules animales ont une taille précise. Le maintien d'une taille cellulaire constante est importante pour le fitness des organismes unicellulaire ainsi que pour multiples fonctions dans les organismes multicellulaires. Les cellules régulent leur taille en coordonnant le taux de croissance avec le taux de division. Ce couplage est essentiel sinon les cellules deviendraient progressivement plus petites ou plus grandes après chaque cycle cellulaire. Dans leur habitat naturels les cellules peuvent faire face a des fluctuations dans le taux de nutriment disponible. Les cellules doivent donc coordonner leur taux de division aux taux variables de croissances perçus dans les différentes conditions nutritionnels. Pendant ma thèse, j'ai travaillée sur une levure unicellulaire, en forme de bâtonnet, nommé levure fissipare ou levure de fission. La taille de ces cellules est plus grande quand le taux de nutriments est grand et plus courte quand celui-ci est plus faible. Une protéine qui perçoit les changements dans le taux externe de la source de carbone (glucose) est nommée PKA pour protéine kinase A. La forme en bâtonnet de la cellule est due aux caractères structuraux du cytosquelette. Une composante importante de ce cytosquelette sont les microtubules, dont la structures ressemble à des petit tubes qui vont d'un bout à l'autre de la cellule. Ces microtubules transportent une protéine importante nommée Tea4 qui à leur tour importante pour la bonne localisation d'une autre protéine Pom1 aux extrémités de la cellule. La protéine Pom1 aide à maintenir la taille appropriée des levures fissipares. Mon travail de thèse a montré qu'en présence de taux faible de nutriments (glucose) les microtubules deviennent de moins en moins stables et montrent une désorganisation globale. Un pourcentage significatif des microtubules touche les côtés de la cellule aux lieu d'atteindre uniquement les extrémités. Ceci a pour conséquence une diffusion de Pom1 tout au long du cortex de la cellule. Ceci aide les levures fissipares à maintenir la taille appropriée pendant ce stress nutritionnel. De plus, je montre que PKA régule la stabilité et l'organisation des microtubules et par conséquent la diffusion de Pom1 et le maintien d'une taille constante. En conclusion, mon travail a conduit à la découverte d'un nouveau mécanisme par lequel la levure fissipare maintient sa taille dans des conditions limitantes en glucose.

Effect of Smoking on Blood Pressure and Resting Heart Rate: A Mendelian Randomization Meta-Analysis in the CARTA Consortium.

BACKGROUND: Smoking is an important cardiovascular disease risk factor, but the mechanisms linking smoking to blood pressure are poorly understood. METHODS AND RESULTS: Data on 141 317 participants (62 666 never, 40 669 former, 37 982 current smokers) from 23 population-based studies were included in observational and Mendelian randomization meta-analyses of the associations of smoking status and smoking heaviness with systolic and diastolic blood pressure, hypertension, and resting heart rate. For the Mendelian randomization analyses, a genetic variant rs16969968/rs1051730 was used as a proxy for smoking heaviness in current smokers. In observational analyses, current as compared with never smoking was associated with lower systolic blood pressure and diastolic blood pressure and lower hypertension risk, but with higher resting heart rate. In observational analyses among current smokers, 1 cigarette/day higher level of smoking heaviness was associated with higher (0.21 bpm; 95% confidence interval 0.19; 0.24) resting heart rate and slightly higher diastolic blood pressure (0.05 mm Hg; 95% confidence interval 0.02; 0.08) and systolic blood pressure (0.08 mm Hg; 95% confidence interval 0.03; 0.13). However, in Mendelian randomization analyses among current smokers, although each smoking increasing allele of rs16969968/rs1051730 was associated with higher resting heart rate (0.36 bpm/allele; 95% confidence interval 0.18; 0.54), there was no strong association with diastolic blood pressure, systolic blood pressure, or hypertension. This would suggest a 7 bpm higher heart rate in those who smoke 20 cigarettes/day. CONCLUSIONS: This Mendelian randomization meta-analysis supports a causal association of smoking heaviness with higher level of resting heart rate, but not with blood pressure. These findings suggest that part of the cardiovascular risk of smoking may operate through increasing resting heart rate.

Typology of "Fatigue" by Heart Rate Variability Analysis in Elite Nordic-skiers.

This study investigated changes in heart rate variability (HRV) in elite Nordic-skiers to characterize different types of "fatigue" in 27 men and 30 women surveyed from 2004 to 2008. R-R intervals were recorded at rest during 8 min supine (SU) followed by 7 min standing (ST). HRV parameters analysed were powers of low (LF), high (HF) frequencies, (LF+HF) (ms(2)) and heart rate (HR, bpm). In the 1 063 HRV tests performed, 172 corresponded to a "fatigue" state and the first were considered for analysis. 4 types of "fatigue" (F) were identified: 1. F(HF(-)LF(-))SU_ST for 42 tests: decrease in LFSU (- 46%), HFSU (- 70%), LFST (- 43%), HFST (- 53%) and increase in HRSU (+ 15%), HRST (+ 14%). 2. F(LF(+) SULF(-) ST) for 8 tests: increase in LFSU (+ 190%) decrease in LFST (- 84%) and increase in HRST (+ 21%). 3. F(HF(-) SUHF(+) ST) for 6 tests: decrease in HFSU (- 72%) and increase in HFST (+ 501%). 4. F(HF(+) SU) for only 1 test with an increase in HFSU (+ 2161%) and decrease in HRSU (- 15%). Supine and standing HRV patterns were independently modified by "fatigue". 4 "fatigue"-shifted HRV patterns were statistically sorted according to differently paired changes in the 2 postures. This characterization might be useful for further understanding autonomic rearrangements in different "fatigue" conditions.