Velocity estimates for signal propagation leading to systemic jasmonic acid accumulation in wounded Arabidopsis.

The wound response prohormone jasmonic acid (JA) accumulates rapidly in tissues both proximal and distal to injury sites in plants. Using quantitative liquid chromatography-mass spectrometry after flash freezing of tissues, we found that JA accumulated within 30 s of injury in wounded Arabidopsis leaves (p = 3.5 e(-7)). JA augmentation distal to wounds was strongest in unwounded leaves with direct vascular connections to wounded leaves wherein JA levels increased significantly within 120 s of wounding (p = 0.00027). This gave conservative and statistically robust temporal boundaries for the average velocity of the long distance signal leading to distal JA accumulation in unwounded leaves of 3.4-4.5 cm min(-1). Like JA, transcripts of the JA synthesis gene LIPOXYGENASE2 (LOX2) and the jasmonate response gene JAZ10.3 also accumulated to higher levels in directly interconnected leaves than in indirectly connected leaves. JA accumulation in a lox2-1 mutant plant was initiated rapidly after wounding then slowed progressively compared with the wild type (WT). Despite this, JAZ10.3 expression in the two genotypes was similar. Free cyclopentenone jasmonate levels were similar in both resting WT and lox2-1. In contrast, bound cyclopentenone jasmonates (arabidopsides) were far lower in lox2-1 than in the WT. The major roles of LOX2 are to generate arabidopsides and the large levels of JA that accumulate proximal to the wound. LOX2 is not essential for some of the most rapid events elicited by wounding.

Auditory spatio-temporal brain dynamics and their consequences for multisensory interactions in humans.

Recent multisensory research has emphasized the occurrence of early, low-level interactions in humans. As such, it is proving increasingly necessary to also consider the kinds of information likely extracted from the unisensory signals that are available at the time and location of these interaction effects. This review addresses current evidence regarding how the spatio-temporal brain dynamics of auditory information processing likely curtails the information content of multisensory interactions observable in humans at a given latency and within a given brain region. First, we consider the time course of signal propagation as a limitation on when auditory information (of any kind) can impact the responsiveness of a given brain region. Next, we overview the dual pathway model for the treatment of auditory spatial and object information ranging from rudimentary to complex environmental stimuli. These dual pathways are considered an intrinsic feature of auditory information processing, which are not only partially distinct in their associated brain networks, but also (and perhaps more importantly) manifest only after several tens of milliseconds of cortical signal processing. This architecture of auditory functioning would thus pose a constraint on when and in which brain regions specific spatial and object information are available for multisensory interactions. We then separately consider evidence regarding mechanisms and dynamics of spatial and object processing with a particular emphasis on when discriminations along either dimension are likely performed by specific brain regions. We conclude by discussing open issues and directions for future research.

Evaluation of molecular, cellular and behavioral consequences of serotonin 1A receptor deletion

In 1998, three different research groups simultaneously reported increased anxiety-related behavior in tests of conflict in their serotonin 1a (5-HT1a) receptor knockout (KO) line with male mice being more severely affected by 5-HT1a receptor deletion than female KO. Similarly, in the hippocampus, we observed increased dendritic complexity in the stratum radiatum of CA1 pyramidal neurons in male but not in female 5-HT1a receptor KO mice. These observations prompted us to investigate gender- dependent differences of 5-HT1a receptor deletion in hippocampal-related behavioral tasks. Testing our mice in anxiety-related paradigms, we reproduced the original studies showing increased anxiety- related behavior in male 5-HT1a receptor KO mice when compared to male WT mice, but no difference between female 5-HT1a receptor KO and WT mice. Similarly, male 5-HT1a receptor KO mice were impaired in association of aversive stimuli fear conditioning paradigms. We argue that increased dendritic complexity and increased synaptic strength of CA3-CA1 synapses in the stratum radiatum impaired proper signal propagation attributed to overactivation of CA1 pyramidal neurons leading to impaired fear memory of male 5-HT1a receptor KO mice. Similar mechanisms in the ventral hippocampus are likely to have contributed to gender-dependent differences in anxiety-related behavior in our and the original studies from 1998. In this study, we started to shed light on the 5-HT1a receptor downstream signaling pathways involved in dendritogenesis of pyramidal neurons during early postnatal development. We could show that NR2B-containing NMDA receptor during development acts downstream of 5-HT1a receptor and is responsible for increased amount of branching in male 5-HT1a receptor KO mice. Conversely, protein and NR2B mRNA expression was increased in 5-HT1a receptor KO mice at P15. Although the exact signaling cascade of 5-HT1a receptor regulating NR2B-containing NMDA receptor has not been determined, CaMKII is a potential downstream effector to influence transportation and removal of NR2B-containing NMDA receptors to and from the synapse. In contrast, Erk1/2 likely acts downstream of NR2B-containing NMDA receptors and was shown to be sufficient to regulate dendritic branching. Moreover, increased NR2B-containing NMDA receptor mediated cell death via excitotoxicity during development and is likely to be involved in reduced survival of adult born neurons in the hippocampus of 5-HT1a receptor KO male. The convergence of 5-HT1a receptor signaling onto NR2B-containing NMDA receptor signaling enables estrogen to interfere with its downstream pathway via G-protein coupled estrogen receptor 1 activation resulting in normalization of branching and behavior in female 5-HT1a receptor mice. In conclusion, our data strongly suggests a hormone- regulated mechanism that by converging on NR2B-containing NMDA receptor signaling is able to normalize morphology of pyramidal neurons and behavior of female 5-HT1a receptor KO mice. Our findings provide a possible explanation for gender-dependent differences in the occurrence of mental disorders with 5-HT1a receptor abnormalities as a strong predisposing factor. -- En 1998, trois équipes de recherche ont décrit un comportement de type anxieux dans des tests de conflit pour leur souris transgéniques avec une délétion du gène pour le récepteur 5-HT1a de la sérotonine. De plus, les trois groupes rapportent un phénotype plus sévère pour le comportement anxieux chez les souris transgéniques mâles que femelles. Dans l'hippocampe, la région avec la densité de récepteur 5-HT1a la plus élevée dans le télencéphale, nous avons observé dans le stratum radiatum une complexité accrue des arborisations dendritiques des neurones pyramidaux du secteur CA1 chez les souris transgénique mâles mais pas chez les femelles. Cette observation nous a encouragés à initier cette étude sur les différences en fonction du genre utilisant les tests comportementaux en rapport avec les fonctions de l'hippocampe chez les souris déficientes pour le récepteur 5-HT1a.Testant nos souris avec des paradigmes associés à l'anxiété, nous avons reproduit les données originales montrant que les souris transgéniques mâles ont un phénotype plus sévère que les souris mâles sauvages, mais qu'aucune différence n'est observée entre les femelles sauvages et transgéniques. De même, les souris mâles déficientes pour le récepteur 5-HT1a sont handicapées dans les tests de conditionnement au stress avec des stimuli aversifs. Nous faisons l'hypothèse que l'augmentation de la complexité de l'arborisation dendritique et l'augmentation de la force du signal synaptique entres les régions CA3 et CA1 de l'hippocampe dans le stratum radiatum perturbe la propagation du signal nerveux qui conduit à l'hyperactivation des neurones du secteur CA1. Ceci conduit à une mémoire de stress altérée chez les souris mâles déficientes pour le récepteur 5-HT1a. Un mécanisme similaire dans l'hippocampe ventral contribue probablement aux différences en fonction du genre dans les tests pour le comportement de type anxieux qui ont été rapportés dans les études originales de 1998. Les mesures de protéine et de mRNA ont mis en évidence une augmentation de l'expression du récepteur NMDA contenant la sous- unité NR2B dans les souris déficientes pour le récepteur 5-HT1a à P15. Dans les cultures organotypiques d'hippocampe, nous avons commencé à disséquer les messagers secondaires à l'activation du récepteur 5-HT1a qui sont impliqués dans la régulation de la croissance dendritique des neurones pyramidaux pendant la période postnatale précoce. Nous avons démontré que les récepteurs NR2B sont en aval de l'activation du récepteur 5-HT1a et qu'ils sont impliqués dans l'accroissement du nombre de dendrites chez la souris mâle déficiente pour le récepteur 5-HT1a. Bien que la cascade de signalisation du récepteur 5-HT1a pour réguler les récepteurs NMDA contenant le NR2B ne soit pas établie, CaMKII est identifié comme un effecteur potentiel pour altérer le transport du récepteur NMDA à la synapse. D'autre part, Erk1/2 est probablement un messager en aval du NR2B du récepteur NMDA, et a été documenté comme suffisant pour réguler l'arborisation dendritique. L'augmentation de NR2B à la synapse des souris déficientes pour le récepteur 5-HT1a peut conduire à une augmentation de l'excitotoxicité dans les cellules. Nous avons observé une augmentation chez la souris déficiente pour le récepteur 5-HT1a de la mort cellulaire dans des tranches d'hippocampe stimulées, ce qui peut être en relation avec la réduction de la survie des neurones générés dans l'hippocampe de la souris mâle transgénique adulte par rapport à la souris mâle sauvage. De plus, la convergence de la signalisation du récepteur 5-HT1a sur la signalisation de la sous-unité NR2B du récepteur NMDA permet à l'oestrogène d'interférer avec sa voie de signalisation du récepteur de l'oestrogène couplé à une protéine G (GPER-1), ceci permettant à l'oestrogène de réduire la taille de l'arborisation des neurones pyramidaux de CA1 chez la femelle de la souris déficiente pour le récepteur 5-HT1a. En conclusion, nos observations suggèrent fortement qu'un mécanisme hormonal convergeant sur la voie de signalisation de la sous-unité NR2B du récepteur NMDA permet la normalisation de l'exubérance des dendrites des neurones CA1 de l'hippocampe et du comportement des souris femelles déficientes pour le récepteur 5-HT1a. Ceci donne une explication possible pour la différence en fonction du genre dans l'apparition de troubles mentaux avec les variations du récepteur 5-HT1a comme facteur de prédisposition important.

Uric acid is a danger signal activating NALP3 inflammasome in lung injury inflammation and fibrosis.

RATIONALE: Lung injury leads to pulmonary inflammation and fibrosis through myeloid differentiation primary response gene 88 (MyD88) and the IL-1 receptor 1 (IL-1R1) signaling pathway. The molecular mechanisms by which lung injury triggers IL-1beta production, inflammation, and fibrosis remain poorly understood. OBJECTIVES: To determine if lung injury depends on the NALP3 inflammasome and if bleomycin (BLM)-induced lung injury triggers local production of uric acid, thereby activating the NALP3 inflammasome in the lung. Methods: Inflammation upon BLM administration was evaluated in vivo in inflammasome-deficient mice. Pulmonary uric acid accumulation, inflammation, and fibrosis were analyzed in mice treated with the inhibitor of uric acid synthesis or with uricase, which degrades uric acid. MEASUREMENTS AND MAIN RESULTS: Lung injury depends on the NALP3 inflammasome, which is triggered by uric acid locally produced in the lung upon BLM-induced DNA damage and degradation. Reduction of uric acid levels using the inhibitor of uric acid synthesis allopurinol or uricase leads to a decrease in BLM-induced IL-1beta production, lung inflammation, repair, and fibrosis. Local administration of exogenous uric acid crystals recapitulates lung inflammation and repair, which depend on the NALP3 inflammasome, MyD88, and IL-1R1 pathways and Toll-like receptor (TLR)2 and TLR4 for optimal inflammation but are independent of the IL-18 receptor. CONCLUSIONS: Uric acid released from injured cells constitutes a major endogenous danger signal that activates the NALP3 inflammasome, leading to IL-1beta production. Reducing uric acid tissue levels represents a novel therapeutic approach to control IL-1beta production and chronic inflammatory lung pathology.

Identification and characterization of novel molecular mechanisms involved in initiation and progression of myelination

Schwann cells synthesize a large amount of membrane that form a specialized structure called myelin that surrounds axons and facilitate the transmission of electrical signal along neurons in peripheral nervous system (PNS). Previous studies demonstrated that both Schwann cell differentiation and de-differentiation (in the situation of a nerve injury or demyelinating disease) are regulated by cell-intrinsic regulators including several transcription factors. In particular, the de-differentiation of mature Schwann cells is driven by the activation of multiple negative regulators of myelination including Sox2, c-Jun, Notch and Pax3, all usually expressed in immature Schwann cells and suppressed at the onset of myelination. In order to identify new regulators of myelination involved in the development of the PNS, we analyzed the gene-expression profiling data from developing PNS and from three models of demyelinating neuropathies. This analysis led to the identification of Sox4, a member of the Sox family of transcription factors, as a potential candidate. To characterize the molecular function of Sox4 in PNS, we generated two transgenic lines of mice, which overexpress Sox4 specifically in Schwann cells. Detailed analysis of these mice showed that the overexpression of Sox4 in Schwann cells causes a delay in progression of myelination between post-natal day 2 (P2) and P5. Our in vitro analysis suggested that Sox4 cDNA can be overexpressed while the protein translation is tightly regulated. Interestingly, we observed that Sox4 protein is stabilized in nerves of the CMT4C mouse, a model of the human neuropathy. We therefore crossed Sox4 transgenic mice with CMT4C mice and we observed that Sox4 overexpression exacerbated the neuropathy phenotype in these mice. While recognized as being crucial for the normal function of both neurons and myelinating glial cells, the processes that regulate the beginning of myelination and the nature of the neuro-glial cross-talk remains mostly unknown. In order to gain insight into the molecular pathways involved in the interactions between neurons and associated glial cells, we developed a neuron-glia co-culture system based on microfluidic chambers and successfully induced myelination in this system by ascorbic acid. Importantly, we observed that in addition to acting on Schwann cells, ascorbic acid also modulate neuronal/axonal NRG1/ErbB2-B3 signalling. The experimental setting used in our study thus allowed us to discover a novel phenomena of propagation for myelination in vitro. The further characterization of this event brought us to identify other compounds able to induce myelination: ADAMs secretases inhibitor GM6001 and cyclic-AMP. The results generated during my thesis project are therefore not only important for the advancement of our understanding of how the PNS works, but may also potentially help to develop new therapies aiming at improvement of PNS myelination under disease conditions. - Les cellules de Schwann synthétisent une grande quantité de membrane formant une structure spécialisée appelée myéline qui entoure les axones et facilite la transmission du signal électrique le long des neurones du système nerveux périphérique (SNP). Des études antérieures ont démontré que la différenciation et la dédifférenciation des cellules de Schwann (dans la situation d'une lésion nerveuse ou d'une maladie démyélinisante) sont régulées par des régulateurs cellulaires intrinsèques, incluant plusieurs facteurs de transcription. En particulier, la dédifférenciation des cellules de Schwann matures est contrôlée par l'activation de plusieurs régulateurs négatifs de la myélinisation dont Sox2, c-Jun, Notch et Pax3, tous habituellement exprimés dans des cellules de Schwann immatures et supprimés au début de la myélinisation. Afin d'identifier de nouveaux régulateurs de myélinisation impliqués dans le développement du SNP, nous avons analysé le profil d'expression génique durant le développement du SNP ainsi que dans trois modèles de neuropathies démyélinisantes. Cette analyse a mené à l'identification de Sox4, un membre de la famille des facteurs de transcription Sox, comme étant un candidat potentiel. Dans le but de caractériser la fonction moléculaire de Sox4 dans le SNP, nous avons généré deux lignées transgéniques de souris qui surexpriment Sox4 spécifiquement dans les cellules de Schwann. L'analyse détaillée de ces souris a montré que la surexpression de Sox4 dans les cellules de Schwann provoque un retard dans la progression de la myélinisation entre le jour postnatal 2 (P2) et P5. Notre analyse in vitro a suggéré que l'ADNc de Sox4 peut être surexprimé alors que la traduction des protéines est quand à elle étroitement régulée. De façon intéressante, nous avons observé que la protéine Sox4 est stabilisée dans les nerfs des souris CMT4C, un modèle de neuropathie humaine. Nous avons donc croisé les souris transgéniques Sox4 avec des souris CMT4C et avons observé que la surexpression de Sox4 exacerbe le phénotype de neuropathie chez ces souris. Bien que reconnus comme étant cruciaux pour le fonctionnement normal des neurones et des cellules gliales myélinisantes, les processus qui régulent le début de la myélinisation ainsi que la nature des interactions neurone-glie restent largement méconnus. Afin de mieux comprendre les mécanismes moléculaires impliqués dans les interactions entre les neurones et les cellules gliales leur étant associés, nous avons développé un système de co-culture neurone-glie basé sur des chambres microfluidiques et y avons induit avec succès la myélinisation avec de l'acide ascorbique. Étonnamment, nous avons remarqué que, en plus d'agir sur les cellules de Schwann, l'acide ascorbique module également la voie de signalisation neuronale/axonale NRG1/ErbB2-B3. Le protocole expérimental utilisé dans notre étude a ainsi permis de découvrir un nouveau phénomène de propagation de la myélinisation in vitro. La caractérisation plus poussée de ce phénomène nous a menés à identifier d'autres composés capables d'induire la myélinisation: L'inhibiteur de sécrétases ADAMs GM6001 et l'AMP cyclique. Les résultats obtenus au cours de mon projet de thèse ne sont donc pas seulement importants pour l'avancement de notre compréhension sur la façon dont le SNP fonctionne, mais peuvent aussi potentiellement aider à développer de nouvelles thérapies visant à l'amélioration de la myélinisation du SNP dans des conditions pathologiques.

Investigation of field and diffusion time dependence of the diffusion-weighted signal at ultrahigh magnetic fields.

Over the last decade, there has been a significant increase in the number of high-magnetic-field MRI magnets. However, the exact effect of a high magnetic field strength (B0 ) on diffusion-weighted MR signals is not yet fully understood. The goal of this study was to investigate the influence of different high magnetic field strengths (9.4 T and 14.1 T) and diffusion times (9, 11, 13, 15, 17 and 24 ms) on the diffusion-weighted signal in rat brain white matter. At a short diffusion time (9 ms), fractional anisotropy values were found to be lower at 14.1 T than at 9.4 T, but this difference disappeared at longer diffusion times. A simple two-pool model was used to explain these findings. The model describes the white matter as a first hindered compartment (often associated with the extra-axonal space), characterized by a faster orthogonal diffusion and a lower fractional anisotropy, and a second restricted compartment (often associated with the intra-axonal space), characterized by a slower orthogonal diffusion (i.e. orthogonal to the axon direction) and a higher fractional anisotropy. Apparent T2 relaxation time measurements of the hindered and restricted pools were performed. The shortening of the pseudo-T2 value from the restricted compartment with B0 is likely to be more pronounced than the apparent T2 changes in the hindered compartment. This study suggests that the observed differences in diffusion tensor imaging parameters between the two magnetic field strengths at short diffusion time may be related to differences in the apparent T2 values between the pools. Copyright © 2013 John Wiley & Sons, Ltd.

A UV signal of offspring condition mediates context-dependent parental favouritism.

As fitness returns during a breeding attempt are context-dependent, parents are predicted to bias their food allocation within a brood from poor towards good condition nestlings when environmental conditions deteriorate. We tested this prediction in the Alpine swift and the European starling, two migratory bird species, by modifying an ultraviolet (UV) visual signal of condition in nestlings and exploring how parents allocate food to their young as the season progresses. We show in both species that: (i) UV light reflected by the body skin of offspring positively correlates with their stature (i.e. body mass and skeletal size) and (ii) parental favouritism towards young with more UV reflective skin gradually increases as the season progresses. Early-breeding parents supplied food preferentially to UV pale (i.e. small stature) nestlings, whereas late-breeding parents favoured UV bright offspring (i.e. large stature). These results emphasize that parents use UV signals of offspring condition to adjust their feeding strategies depending on the ecological context.

TRAIL receptors 1 (DR4) and 2 (DR5) signal FADD-dependent apoptosis and activate NF-kappaB.

TRAIL induces apoptosis through two closely related receptors, TRAIL-R1 (DR4) and TRAIL-R2 (DR5). Here we show that TRAIL-R1 can associate with TRAIL-R2, suggesting that TRAIL may signal through heteroreceptor signaling complexes. Both TRAIL receptors bind the adaptor molecules FADD and TRADD, and both death signals are interrupted by a dominant negative form of FADD and by the FLICE-inhibitory protein FLIP. The recruitment of TRADD may explain the potent activation of NF-kappaB observed by TRAIL receptors. Thus, TRAIL receptors can signal both death and gene transcription, functions reminiscent of those of TNFR1 and TRAMP, two other members of the death receptor family.

Recombinant " Physcomitrella patens " as a sensitive tool to study heat sensing and heat-shock signal transduction in plants

SUMMARY When exposed to heat stress, plants display a particular set of cellular and molecular responses, such as chaperones expression, which are highly conserved in all organisms. In chapter 1, I studied the ability of heat shock genes to become transiently and abundantly induced under various temperature regimes. To this aim, I designed a highly sensitive heat-shock dependent conditional gene expression system in the moss Physcomitrella patens, using the soybean heatinducible promoter (hsp17.3B). Heat-induced expression of various reporter genes was over three orders of magnitude, in tight correlation with the intensity and duration of the heat treatments. By performing repeated heating/cooling cycles, a massive accumulation of recombinant proteins was obtained. Interestingly, the hsp17.3B promoter was also activated by specific organic chemicals. Thus, in chapter 2, I took advantage of the extreme sensitivity of this promoter to small temperature variations to further address the role of various natural and organic chemicals and develop a plant based-bioassay that can serve as an early warning indicator of toxicity by pollutants and heavy metals. A screen of several organic pollutants from textile and paper industry showed that chlorophenols as well as sulfonated anthraquinones elicited a heat shock like response at noninducing temperatures. Their effects were synergistically amplified by mild elevated temperatures. In contrast to standard methods of pollutant detection, this plant-based biosensor allowed to monitor early stress-responses, in correlation with long-term toxic effect, and to attribute effective toxicity thresholds for pollutants, in a context of varying environmental cues. In chapter 3, I deepened the study of the primary mechanism by which plants sense mild temperature variations and trigger a cellular signal leading to the heat shock response. In addition to the above described heat-inducible reporter line, I generated a P. patens transgenic line to measure, in vivo, variations of cytosolic calcium during heat treatment, and another line to monitor the role of protein unfolding in heat-shock sensing and signalling. The heat shock signalling pathway was found to be triggered by the plasma membrane, where temperature up shift specifically induced the transient opening of a putative high afimity calcium channel. The calcium influx triggered a signalling cascade leading to the activation of the heat shock genes, independently on the presence of misfolded proteins in the cytoplasm. These results strongly suggest that changes in the fluidity of the plasma membrane are the primary trigger of the heatshocksignalling pathway in plants. The present thesis contributes to the understanding of the basic mechanism by which plants perceive and respond to heat and chemical stresses. This may contribute to developing appropriate better strategies to enhance plant productivity under the increasingly stressful environment of global warming. RÉSUME Les plantes exposées à des températures élevées déclenchent rapidement des réponses cellulaires qui conduisent à l'induction de gènes codant pour les heat shock proteins (HSPs). En fonction de la durée d'exposition et de la vitesse à laquelle la température augmente, les HSPs sont fortement et transitoirement induites. Dans le premier chapitre, cette caractéristique aété utilisée pour développer un système inductible d'expression de gènes dans la mousse Physcomitrella patens. En utilisant plusieurs gènes rapporteurs, j'ai montré que le promoteur du gène hsp17.3B du Soja est activé d'une manière. homogène dans tous les tissus de la mousse proportionnellement à l'intensité du heat shock physiologique appliqué. Un très fort taux de protéines recombinantes peut ainsi être produit en réalisant plusieurs cycles induction/recovery. De plus, ce promoteur peut également être activé par des composés organiques, tels que les composés anti-inflammatoires, ce qui constitue une bonne alternative à l'induction par la chaleur. Les HSPs sont induites pour remédier aux dommages cellulaires qui surviennent. Étant donné que le promoteur hsp17.3B est très sensible à des petites augmentations de température ainsi qu'à des composés chimiques, j'ai utilisé les lignées développées dans le chapitre 1 pour identifier des polluants qui déclenchent une réaction de défense impliquant les HSPs. Après un criblage de plusieurs composés, les chlorophénols et les antraquinones sulfonés ont été identifiés comme étant activateurs du promoteur de stress. La détection de leurs effets a été réalisée seulement après quelques heures d'exposition et corrèle parfaitement avec les effets toxiques détectés après de longues périodes d'exposition. Les produits identifiés montrent aussi un effet synergique avec la température, ce qui fait du biosensor développé dans ce chapitre un bon outil pour révéler les effets réels des polluants dans un environnement où les stress chimiques sont combinés aux stress abiotiques. Le troisième chapitre est consacré à l'étude des mécanismes précoces qui permettent aux plantes de percevoir la chaleur et ainsi de déclencher une cascade de signalisation spécifique qui aboutit à l'induction des gènes HSPs. J'ai généré deux nouvelles lignées afin de mesurer en temps réel les changements de concentrations du calcium cytosolique ainsi que l'état de dénaturation des protéines au cours du heat shock. Quand la fluidité de la membrane augmente après élévation de la température, elle semble induire l'ouverture d'un canal qui permet de faire entrer le calcium dans les cellules. Ce dernier initie une cascade de signalisation qui finit par activer la transcription des gènes HSPs indépendamment de la dénaturation de protéines cytoplasmiques. Les résultats présentés dans ce chapitre montrent que la perception de la chaleur se fait essentiellement au niveau de la membrane plasmique qui joue un rôle majeur dans la régulation des gènes HSPs. L'élucidation des mécanismes par lesquels les plantes perçoivent les signaux environnementaux est d'une grande utilité pour le développement de nouvelles stratégies afin d'améliorer la productivité des plantes soumises à des conditions extrêmes. La présente thèse contribue à décortiquer la voie de signalisation impliquée dans la réponse à la chaleur.

MR spectroscopy of the human brain with enhanced signal intensity at ultrashort echo times on a clinical platform at 3T and 7T.

Recently, the spin-echo full-intensity acquired localized (SPECIAL) spectroscopy technique was proposed to unite the advantages of short TEs on the order of milliseconds (ms) with full sensitivity and applied to in vivo rat brain. In the present study, SPECIAL was adapted and optimized for use on a clinical platform at 3T and 7T by combining interleaved water suppression (WS) and outer volume saturation (OVS), optimized sequence timing, and improved shimming using FASTMAP. High-quality single voxel spectra of human brain were acquired at TEs below or equal to 6 ms on a clinical 3T and 7T system for six volunteers. Narrow linewidths (6.6 +/- 0.6 Hz at 3T and 12.1 +/- 1.0 Hz at 7T for water) and the high signal-to-noise ratio (SNR) of the artifact-free spectra enabled the quantification of a neurochemical profile consisting of 18 metabolites with Cramér-Rao lower bounds (CRLBs) below 20% at both field strengths. The enhanced sensitivity and increased spectral resolution at 7T compared to 3T allowed a two-fold reduction in scan time, an increased precision of quantification for 12 metabolites, and the additional quantification of lactate with CRLB below 20%. Improved sensitivity at 7T was also demonstrated by a 1.7-fold increase in average SNR (= peak height/root mean square [RMS]-of-noise) per unit-time.

Protein-binding site at the immunoglobulin mu membrane polyadenylylation signal: possible role in transcription termination.

mRNAs specifying immunoglobulin mu and delta heavy chains are encoded by a single large, complex transcription unit (mu + delta gene). The transcriptional activity of delta gene segments in terminally differentiated, IgM-secreting B lymphocytes is 10-20 times lower than in earlier B-lineage cells expressing delta mRNA. We find that transcription of the mu + delta gene in IgM-secreting murine myeloma cells terminates within a region of 500-1000 nucleotides immediately following the mu membrane (mu m) polyadenylylation site. Transcription decreases only minimally through this region in murine cell lines representative of earlier stages in B-cell development. A DNA fragment containing the mu m polyadenylylation signal gives protein-DNA complexes with different mobilities in gel retardation assays with nuclear extracts from myeloma cells than with nuclear extracts from earlier B-lineage cells. However, using a recently developed "footprinting" procedure in which protein-DNA complexes resolved in gel retardation assays are subjected to nucleolytic cleavage while still in the polyacrylamide gel, we find that the DNA sequences protected by factors from the two cell types are indistinguishable. The factor-binding site on the DNA is located 5' of the mu m polyadenylylation signal AATAAA and includes the 15-nucleotide-long A + T-rich palindrome CTGTAAACAAATGTC. This type of palindromic binding site exhibits orientation-dependent activity consistent with the reported properties of polymerase II termination signals. This binding site is followed by two sets of directly repeated DNA sequences with different helical conformation as revealed by their reactivity with the chemical nuclease 1,10-phenanthroline-copper. The close proximity of these features to the signals for mu m mRNA processing may reflect a linkage of the processes of developmentally regulated mu m polyadenylylation and transcription termination.

Echolocation calls of the bats of Trinidad, West Indies: is guild membership reflected in echolocation signal design?

Time-expanded echolocation calls were recorded from 29 species of Neotropical bats in lowland moist tropical forest in Trinidad, West Indies with three aims (I) to describe the echolocation calls of the members of a diverse Neotropical bat community, especially members of the family Phyllostomidae, whose calls are not well documented (2) to investigate whether multivariate analysis of calls allows species and foraging guilds to be identified and (3) to evaluate the use of bat detectors in surveying the phyllostomids of Neotropical forests. The calls of 12 species of the family Phyllostomidae are described here for the first time and a total of 29 species, belonging to five families (Emballonuridae, Mormoopidae, Phyllostomidae, Molossidae and Vespertilionidae) were recorded Quadratic discriminant function analysis (DFA) was used to obtain classification rates for each one of 11 individual species and for six guilds (based on diet, foraging mode and habitat) comprising 26 species Overall classification rates were low compared to similar studies conducted in the Palaeotropics We suggest that this may be due to a combination of ecological plasticity for certain species and a loose relationship between echolocation call shape, fine-grained resource partitioning and resource acquisition in phyllostomids

Ezrin-Radixin-Moesin-binding Sequence of PSGL-1 Glycoprotein Regulates Leukocyte Rolling on Selectins and Activation of Extracellular Signal-regulated Kinases.

P-selectin glycoprotein ligand-1 (PSGL-1) mediates the capture (tethering) of free-flowing leukocytes and subsequent rolling on selectins. PSGL-1 interactions with endothelial selectins activate Src kinases and spleen tyrosine kinase (Syk), leading to α(L)β(2) integrin-dependent leukocyte slow rolling, which promotes leukocyte recruitment into tissues. In addition, but through a distinct pathway, PSGL-1 engagement activates ERK. Because ezrin, radixin and moesin proteins (ERMs) link PSGL-1 to actin cytoskeleton and because they serve as adaptor molecules between PSGL-1 and Syk, we examined the role of PSGL-1 ERM-binding sequence (EBS) on cell capture, rolling, and signaling through Syk and MAPK pathways. We carried out mutational analysis and observed that deletion of EBS severely reduced 32D leukocyte tethering and rolling on L-, P-, and E-selectin and slightly increased rolling velocity. Alanine substitution of Arg-337 and Lys-338 showed that these residues play a key role in supporting leukocyte tethering and rolling on selectins. Importantly, EBS deletion or Arg-337 and Lys-338 mutations abrogated PSGL-1-induced ERK activation, whereas they did not prevent Syk phosphorylation or E-selectin-induced leukocyte slow rolling. These studies demonstrate that PSGL-1 EBS plays a critical role in recruiting leukocytes on selectins and in activating the MAPK pathway, whereas it is dispensable to phosphorylate Syk and to lead to α(L)β(2)-dependent leukocyte slow rolling.