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Die Funktionalität β1- und β2-adrenerger Rezeptoren wird durch Polymorphismen in ihrer kodierenden Region moduliert. Wir haben uns die Technik des Fluoreszenz-Resonanz- Energie-Transfers (FRET) zu Nutze gemacht, um den Einfluss der am häufigsten vorkommenden Polymorphismen (Ser49Gly und Gly389Arg im β1AR, Arg16Gly und Gln27Glu im β2AR) auf die Rezeptorkonformation nach Aktivierung zu untersuchen. Dafür wurden FRET-Sensoren für die beiden βAR-Subtypen mit einem gelb-fluoreszierenden Protein (YFP) sowie einem cyan-fluoreszierenden Protein (CFP oder Cerulean) in der dritten intrazellulären Schleife bzw. am C-Terminus verwendet. Nach Stimulierung der βARSensoren konnte die Aktivierung der polymorphen Rezeptorvarianten in lebenden Zellen in Echtzeit untersucht werden. Dabei behielten die FRET-Sensoren sowohl die Bindungsaffinitäten der nativen Rezeptoren als auch eine intakte Funktionalität hinsichtlich der Bildung von sekundären Botenstoffen. Der Vergleich der Aktivierungskinetiken der verschieden polymorphen Varianten des β1AR und β2AR ergab keine signifikanten Unterschiede nach einer einmaligen Stimulation. Es zeigte sich jedoch, dass Rezeptorpolymorphismen die Aktivierungskinetik vorstimulierter βAR erheblich beeinflussen. So konnten wir im Vergleich zur ersten Aktivierung eine schnellere Aktivierung der Gly16-Varianten des β2AR sowie des Gly49Arg389-β1AR feststellen, während die Arg16-β2AR-Variante und der Ser49Gly389-β1AR dagegen bei einer wiederholten Stimulation langsamer aktiviert wurden. Diese Ergebnisse lassen auf ein "Rezeptorgedächtnis" schließen, das spezifisch für bestimmte polymorphe Rezeptorvarianten ist und eine βAR-Subtyp-spezische Ausprägung zeigt. Die Ausbildung der unterschiedlichen Aktivierungskinetiken hing von der Interaktion des Rezeptors mit löslichen intrazellulären Faktoren ab und bedurfte einer Phosphorylierung intrazellulärer Serin- und Threonin-Reste durch G-Protein-gekoppelte Rezeptorkinasen. Die Interaktion mit löslichen intrazellulären Faktoren scheint für den β1AR weniger stark ausgeprägt zu sein als für den β2AR. Die cAMP-Produktion war für die schneller werdenden, “hyperfunktionellen” Gly16-β2ARVarianten signifikant um mehr als 50% höher im Vergleich zur “hypofunktionellen” Arg16- Variante. Die unterschiedliche Funktionalität spiegelte sich im Therapieausgang bei Tokoysepatientinnen wider, dessen Erfolg mit dem Arg16Gly Polymorphismus verknüpft war. Die Daten implizieren eine intrinsische, polymorphismusabhängige Eigenschaft der βAR, die die Aktivierungskinetik der Rezeptoren bei wiederholten Stimulationen determiniert. Diese könnte auch für die zwischen Individuen variierende Ansprechbarkeit auf β-Agonisten und β-Blocker mitverantwortlich sein.
While beneficial sponge-microbe associations have received much attention in recent years, less effort has been undertaken to investigate the interactions of sponges with potentially pathogenic microorganisms. Thus, the aim of this study was to examine two selected Caribbean disease conditions, termed “Sponge Orange Band” and “Sponge White Patch”, via ecological and molecular methods. Sponge Orange Band (SOB) disease affects the prominent Caribbean barrel sponge Xestospongia muta that is counted among the high-microbial-abundance (HMA) sponges, whereas Sponge White Patch (SWP) disease affects the abundant rope sponge Amphimedon compressa that belongs to the low-microbial-abundance (LMA) sponges. I have documented for both Caribbean sponge diseases a disease progression going along with massive tissue destruction as well as loss of the characteristic microbial signatures. Even though new bacteria were shown to colonize the bleached areas, the infection trials revealed in both cases no indication for the involvement of a microbial pathogen as an etiologic agent of disease leaving us still in the dark about the cause of Sponge Orange Band as well as Sponge White Patch disease.
In recent years high-throughput experiments provided a vast amount of data from all areas of molecular biology, including genomics, transcriptomics, proteomics and metabolomics. Its analysis using bioinformatics methods has developed accordingly, towards a systematic approach to understand how genes and their resulting proteins give rise to biological form and function. They interact with each other and with other molecules in highly complex structures, which are explored in network biology. The in-depth knowledge of genes and proteins obtained from high-throughput experiments can be complemented by the architecture of molecular networks to gain a deeper understanding of biological processes. This thesis provides methods and statistical analyses for the integration of molecular data into biological networks and the identification of functional modules, as well as its application to distinct biological data. The integrated network approach is implemented as a software package, termed BioNet, for the statistical language R. The package includes the statistics for the integration of transcriptomic and functional data with biological networks, the scoring of nodes and edges of these networks as well as methods for subnetwork search and visualisation. The exact algorithm is extensively tested in a simulation study and outperforms existing heuristic methods for the calculation of this NP-hard problem in accuracy and robustness. The variability of the resulting solutions is assessed on perturbed data, mimicking random or biased factors that obscure the biological signal, generated for the integrated data and the network. An optimal, robust module can be calculated using a consensus approach, based on a resampling method. It summarizes optimally an ensemble of solutions in a robust consensus module with the estimated variability indicated by confidence values for the nodes and edges. The approach is subsequently applied to two gene expression data sets. The first application analyses gene expression data for acute lymphoblastic leukaemia (ALL) and differences between the subgroups with and without an oncogenic BCR/ABL gene fusion. In a second application gene expression and survival data from diffuse large B-cell lymphomas are examined. The identified modules include and extend already existing gene lists and signatures by further significant genes and their interactions. The most important novelty is that these genes are determined and visualised in the context of their interactions as a functional module and not as a list of independent and unrelated transcripts. In a third application the integrative network approach is used to trace changes in tardigrade metabolism to identify pathways responsible for their extreme resistance to environmental changes and endurance in an inactive tun state. For the first time a metabolic network approach is proposed to detect shifts in metabolic pathways, integrating transcriptome and metabolite data. Concluding, the presented integrated network approach is an adequate technique to unite high-throughput experimental data for single molecules and their intermolecular dependencies. It is flexible to apply on diverse data, ranging from gene expression changes over metabolite abundances to protein modifications in a combination with a suitable molecular network. The exact algorithm is accurate and robust in comparison to heuristic approaches and delivers an optimal, robust solution in form of a consensus module with confidence values. By the integration of diverse sources of information and a simultaneous inspection of a molecular event from different points of view, new and exhaustive insights into biological processes can be acquired.
Die asexuellen Sporen von Aspergillus fumigatus sind ubiquitär verbreitete Luftkeime. Als Saprophyt ist dieser opportunistisch humanpathogene Pilz darauf spezialisiert, polymere Substanzen aus dem umgebenden Milieu zu zersetzen, um daraus die von ihm benötigten Nährstoffe zu generieren und aufzunehmen. Die Fähigkeit, verschiedene Stickstoff- und Kohlenstoffquellen zu verwerten, trägt dabei zu seiner Virulenz bei und hierbei scheint die extrazelluläre Proteolyse eine wichtige Rolle zu spielen. Sekretierte Proteasen, die das umgebende Gewebe während einer Infektion mit A. fumigatus erschließen, könnten somit zu dessen Pathogenität beitragen. Dementsprechend sollte im Rahmen dieser Arbeit die Bedeutung einer Regulation der extrazellulären proteolytischen Aktivität von A. fumigatus für dessen Virulenz untersucht werden. Dies geschah durch Untersuchungen eines konservierten Transkriptionsfaktors, PrtT. Dabei stellte sich heraus, dass PrtT die Expression der drei Hauptproteasen von A. fumigatus, Alp, Mep und Pep stark beeinflusst, in einem murinen Tiermodell der pulmonaren Aspergillose scheint dieser Regulator jedoch keine Rolle für die Pathogenität von A. fumigatus zu spielen. Um einen weiteren Aspekt des pilzlichen Aminosäurestoffwechsels zu beleuchten, wurde die Biosynthese der aromatischen Aminosäuren als mögliche Virulenzdeterminate untersucht. Für den Menschen sind diese Aminosäuren essentiell, weshalb dieser Syntheseweg ein mögliches Ziel für antimykotische Substanzen darstellen könnte. Es konnten mehrere für A. fumigatus essentielle Komponenten des Shikimatweges identifiziert werden, des Weiteren wurden Deletionsmutanten in den Genen aroC und trpA, die für die Chorismatmutase bzw. Anthranilatsynthase der Biosynthese von Phenylalanin und Tyrosin bzw. Tryptophan kodieren, erzeugt und phänotypisch charakterisiert. Deren Untersuchung in einem alternativen Tiermodell der Aspergillose zeigte eine deutlich attenuierte Virulenz. Diese Ergebnisse verdeutlichen, wie wichtig die Biosynthese der aromatischen Aminosäuren für das Wachstum von A. fumigatus ist, und dass ein Eingriff in diesen Syntheseweg eine lohnende Strategie zur Entwicklung neuer Antimykotika sein könnte. Die hier präsentierten Ergebnisse unterstreichen die für den Schimmelpilz A. fumigatus typische Redundanz bezüglich extrazellulärer proteolytischer Enzyme und dass diese nur bedingt hinsichtlich ihres Virulenzbeitrags untersucht werden können. Im Gegensatz hierzu lassen sich bestimmte Stoffwechselwege, die oftmals durch einzigartige Genprodukte katalysiert werden, unter Umständen besser als unspezifische aber vielversprechende Virulenzdeterminanten identifizieren.
Die Amygdala ist ein Kernkomplex, der dicht von serotonergen Afferenzen innerviert wird. Sowohl bei Tieren als auch beim Menschen spielen Interaktionen zwischen dem serotonergen System und der Amygdala bei der Verarbeitung von Reizen, die mit Angst oder Stress assoziiert sind, eine zentrale Rolle. Genetische Variationen im serotonergen System und/oder dauerhafter Stress können dazu führen, dass diese Verarbeitungsprozesse fehlerhaft ablaufen, wodurch Verhaltensanormalitäten bzw. die Entstehung psychiatrischer Erkrankungen begünstigt werden. Die Zielneurone der serotonergen Transmission in der Amygdala, die molekularen Mechanismen möglicher Interaktionen und strukturelle Konsequenzen der Störungen dieser Interaktionen sind jedoch bis zum heutigen Zeitpunkt noch nicht vollständig bekannt. Daher bestand ein Ziel der vorliegenden Arbeit darin, den Einfluss eines Ungleichgewichts im serotonergen System (5-Htt KO) sowie von wiederholtem, sozialem Stress auf die neuronale Morphologie der Amygdala zu analysieren und Zielneurone serotonerger Afferenzen zu identifizieren und zu charakterisieren, um die neuronalen Netzwerke der Emotionsverarbeitung besser verstehen zu können. Um vom 5-Htt–Genotyp abhängige und stressbedingte neuromorphologische Veränderungen zu untersuchen, wurden dreidimensionale Rekonstruktionen von Neuronen der laterobasalen Amygdala von männlichen, adulten Wildtyp (WT)- und 5-Htt KO-Mäusen angefertigt und bezüglich verschiedener morphologischer Parameter ausgewertet. An den Pyramidenzellen wurden nur geringfügige Veränderungen der dendritischen Komplexität, jedoch, im Vergleich zu WT-Mäusen, eine wesentliche Erhöhung der Dornendichte an spezifischen dendritischen Kompartimenten bei gestressten WT-Mäusen, sowie nicht gestressten und gestressten 5-Htt KO-Mäusen nachgewiesen. Im Vergleich zu nicht gestressten WT–Mäusen war die dendritische Dornendichte aller anderen Gruppen gleichermaßen erhöht. Die Sternzelle, zeigten bezüglich der untersuchten Parameter keine morphologischen Veränderungen auf. Eine besondere Subpopulation der Interneurone stellen die NeuropeptidY (NPY)–Neurone der laterobasalen Amygdala dar, da sie in diesen Nuclei anxiolytisch wirken. Es gibt nur wenige Anhaltspunkte darüber, durch welche Systeme NPY–Neurone moduliert werden. Da sowohl NPY–Neurone in der laterobasalen Amygdala als auch das serotonerge System an angstregulierenden Prozessen beteiligt sind, sollte im zweiten Teil der vorliegenden Arbeit untersucht werden, ob es sich bei diesen Neuronen um Zielstrukturen des serotonergen Systems handelt. Mittels licht- und elektronenmikroskopischer Analysen wurden synaptische Kontakte zwischen serotonergen Afferenzen und NPY-immunreaktiven Neuronen in der laterobasalen Amygdala von Ratten verifiziert. Da der funktionelle Einfluss der serotonergen Innervation auf diese Zielneurone von deren Serotoninrezeptor (5-HTR)-Ausstattung abhängt, wurden Koexpressionsanalysen von NPY mRNA mit den mRNAs verschiedener 5-HTR durchgeführt. Die Analysen ergaben, dass NPY mRNA–reaktive Neurone in der laterobasalen Amygdala 5-HT1A und 5-HT2C, jedoch nicht 5-HT3 mRNA koexprimieren. Die in der vorliegenden Arbeit erzielten Resultate liefern neue Erkenntnisse über den Einfluss des serotonergen Systems auf die laterobasale Amygdala von Mäusen und Ratten. Bei den Veränderungen der dendritischen Dornendichte nach sozialen Stresserfahrungen könnte es sich um neuroadaptive bzw. kompensatorische Mechanismen der Pyramidenzellen handeln, die WT-Mäusen eine Anpassung an sich ändernde, negative Umweltbedingungen ermöglicht. Die erhöhte Dornendichte könnte dabei die Ausbildung eines „emotionalen Gedächtnisses“ repräsentieren, das eine flexible Verhaltensantwort auf ein erneutes Auftauchen von Gefahr erlaubt. Eine solche Modulation der Erregbarkeit der laterobasalen Amygdala könnte beispielsweise über eine situationsentsprechende Hemmung des Outputs der Pyramidenzellen durch differentiell aktive inhibitorische Netzwerke erfolgen. Eine differentielle Aktivierung kann z. B. über unterschiedliche Rezeptorausstattungen, wie es in der Subpopulation der NPY–Neurone in der vorliegenden Arbeit nachgewiesen wurde, erfolgen. Das erhöhte angstähnliche Verhalten der 5-Htt KO-Mäuse nach wiederholtem Stress könnte mit der Unfähigkeit zusammenhängen, in entsprechenden Situationen durch Neubildung von Dornen zu reagieren, da die Dornendichte bei diesen Tieren schon unter stressarmen Umweltbedingungen ihr Maximum erreicht hat. Sowohl Fehlfunktionen der neuronalen Plastizität als auch mögliche Fehlfunktionen der differentiellen Inhibierung der Pyramidenzellen durch Interneurone, die durch genetische Variationen und/oder Stress bedingt sein können, könnten eine „offene Tür“ repräsentieren, die zu manifesten Auffälligkeiten im Verhalten bei Tieren führt bzw. auch zur Entstehung bestimmter psychiatrischer Erkrankungen beim Menschen beiträgt.
The Nuclear Factors of Activated T cells (NFATs) are critical transcription factors playing major roles in the control of the cell cycle, apoptosis and, probably, also cancerogenesis. Of all the four genuine NFATc family members, NFATc1 has the unique induction property which appears to be essential for T and B cell development, along with its considerable role in cytokine gene expression and function in non-lymphoid tissues and during organ development (such as in the development of muscle and heart cells). A number of studies have proved the potential role of NFATc1 protein in development of lymphomas and leukemias and provided evidence of differential expression of the same gene in different tumours (Suppression in classical Hodgkin lymphomas but overexpression in T-ALLs). Although the most commonly accepted pathway is the dephosphorylation of NFAT by calcineurin upon a rise in intracellular Ca++ leading to nuclear translocation followed by transcription of Il2 gene and related cytokines, it is quite possible that signaling mechanisms other than (or in addition to) calcineurin activation lead to NFATc1 induction as well. One of the major isoforms of NFATc1, NFATc1/αA, is the short inducible factor, produced upon full T and B cell activation. Here we used two different conditional knock-out mice as our study model. Inactivation of the murine Nfatc1 gene in bone marrow (of Cd79a/mb-1-cre x Nfatc1flx/flx mice) and spleen (of Cd23-cre x Nfatc1flx/flx mice) resulted in complete ablation of NFATc1 expression in splenic B cells. Although no severe developmental defects were found for the generation of ‘conventional’ B2 cells, NFATc1 inactivation in bone marrow B-cells led to a strong decrease in the peritoneal B1a cell population. In-vitro studies showed a clear-cut decrease in proliferation and an increase in Activation Induced Cell Death (AICD) of NFATc1-/- splenic B cells upon BCR stimulation. While NFATc1 appears to control directly the AICD of peripheral B cells, further studies revealed an effect of NFATc1 on proliferation by a sustained differentiation program controlling Ca++ flux and calcineurin activity which are needed to maintain transcription and proliferation of primary B cells. Re-expression of NFATc1 at a low dose could protect cells against AICD, whereas at a higher dose it initiated AICD. These data suggest an important dual role of NFATc1 in controlling proliferation and apoptosis of peripheral B lymphocytes. NFATc1 ablation also impaired the Ig class switch to IgG3 by T cell-independent (TI) type II antigens and impaired IgG3+ plasmablast formation when studied in-vivo by NP-Ficoll immunization or in-vitro using an in-vitro class-switch model. Contrary to the immunizations with TI-type II antigen, no significant differences were documented in Ig class switch upon immunization with NP-KLH, a T-cell dependent (TD) antigen. Taken together, the data indicate NFATc1/αA as a crucial player in the activation and function of splenic B cells upon BCR stimulation. Missing or incomplete NFATc1/αA induction appears to be one reason for the generation of B cell unresponsiveness, whereas uncontrolled NFATc1/αA expression could lead to unbalanced immune reactions and autoimmune diseases.
The present work investigated the neural mechanisms underlying cognitive inhibition/thought suppression in Anderson’s and Green’s Think/No-Think paradigm (TNT), as well as different variables influencing these mechanisms at the cognitive, the neurophysiological, the electrophysiological and the molecular level. Neurophysiological data collected with fNIRS and fMRI have added up to the existing evidence of a fronto-hippocampal network interacting during the inhibition of unwanted thoughts. Some evidence has been presented suggesting that by means of external stimulation of the right dlPFC through iTBS thought suppression might be improved, providing further evidence for an implication of this region in the TNT. A combination of fNIRS with ERP has delivered evidence of a dissociation of early condition-independent attentional and later suppression-specific processes within the dlPFC, both contributing to suppression performance. Due to inconsistencies in the previous literature it was considered how stimulus valence would influence thought suppression by manipulating the emotional content of the to-be-suppressed stimuli. Findings of the current work regarding the ability to suppress negative word or picture stimuli have, however, been inconclusive as well. It has been hypothesized that performance in the TNT might depend on the combination of valence conditions included in the paradigm. Alternatively, it has been suggested that inconsistent findings regarding the suppression of negative stimuli or suppression at all might be due to certain personality traits and/or genetic variables, found in the present work to contribute to thought inhibition in the TNT. Rumination has been shown to be a valid predictor of thought suppression performance. Increased ruminative tendencies led to worse suppression performance which, in the present work, has been linked to less effective recruitment of the dlPFC and in turn less effective down-regulation of hippocampal activity during suppression trials. Trait anxiety has also been shown to interrupt thought suppression despite higher, however, inefficient recruitment of the dlPFC. Complementing the findings regarding ruminative tendencies and decreased thought inhibition a functional polymorphism in the KCNJ6 gene, encompassing a G-to-A transition, has been shown to disrupt thought suppression despite increased activation of the dlPFC. Through the investigation of thought suppression at different levels, the current work adds further evidence to the idea that the TNT reflects an executive control mechanism, which is sensitive to alterations in stimulus valence to some extent, neurophysiological functioning as indicated by its sensitivity to iTBS, functional modulations at the molecular level and personality traits, such as rumination and trait anxiety.
In this thesis I studied psychological aspects in the behaviour of Drosophila, and especially Drosophila larvae. After an introduction where I present the general scientific context and describe the mechanisms of olfactory perception as well as of classical and operant conditioning, I present the different experiments that I realised during my PhD. Perception The second chapter deals with the way adult Drosophila generalise between single odours and binary mixtures of odours. I found that flies perceive a mixture of two odours as equally similar to the two elements composing it; and that the intensity as well as the physico-chemical nature of the elements composing a mixture affect the degree of generalisation between this mixture and one of its elements. These findings now call for further investigation on the physiological level, using functional imaging. Memory The third chapter presents a series of experiments in Drosophila larvae in order to define some characteristics of a new protocol for classical aversive learning which involves associating odours with mechanical disturbance as a punishment. The protocol and the first results should open new doors for the study of classical conditioning in Drosophila larvae, by allowing the comparison between two types of aversive memory (gustatory vs. mechanical reinforcement), including a comparison of their neurogenetic bases. It will also allow enquiries into the question whether these respective memories are specific for the kind of reinforcer used. Agency The fourth chapter documents our attempts to establish operant memory in Drosophila larvae. By analysing the first moments of the test, I could reveal that the larvae modified their behaviour according to their previous operant training. However, this memory seems to be quickly extinguished during the course of the test. We now aim at repeating these results and improving the protocol, in order to be able to systematically study the mechanisms allowing and underlying operant learning in Drosophila larvae. In the fifth chapter, I use the methods developed in chapter four for an analysis of larval locomotion. I determine whether larval locomotion in terms of speed or angular speed is affected by a treatment with the “cognitive enhancer” Rhodiola rosea, or by mutations in the Synapsin or SAP47 genes which are involved in the formation of olfactory memory. I also characterize the modifications induced by the presence of gustatory stimuli in the substrate on which the larvae are crawling. This thesis thus brings new elements to the current knowledge of Drosophila
There is evidence that pheromones are communicative signals in animals. However, the existence and function of human pheromones are still under discussion. During the last years several substances have been labeled as putative human pheromones and especially 4,16–androstadien-3-one (androstadienone), found in male and female sweat, became subject of intense investigation. In contrast to common odors androstadienone presumably modulates human physiological and psychological reactions. Data suggest that androstadienone might influence the processing of visual cues, specifically faces or affective stimuli, via projections from the fusiform gyrus and the amygdala. Moreover, attentional processes may be modulated, which is supported by explicit and implicit behavioral data. This thesis includes three experimental studies examining effects of androstadienone exposure on behavioral and cortical reactions to visual and emotional stimuli. The main hypotheses were that androstadienone might influence human behavior to and perception of visual cues. The first study sought to clarify androstadienone effects on attention-related reactions as well as on behavioral tendencies. Motoric approach-avoidance reactions in response to happy and angry facial expressions were investigated in 30 women and 32 men. Participants either inhaled androstadienone or a control solution, without knowing the real content, while performing the following task: they had to push away or to pull towards them a joystick as fast as possible in reaction to either an angry or a happy cartoon face, which was presented on a computer screen. Results showed that androstadienone modulated the participant´s task performance by accelerating the reaction speed compared to the control compound. Faster reactions were observed particularly when reacting to angry faces but not when reacting to happy faces. This might be explained by the finding that human body odors, the source of androstadienone, were found to activate the human fear system, i.e. modulating fear-related attentional processes. Therefore, the quicker reaction towards angry faces with exposure to androstadienone could be due to an enhanced allocation of attentional resources towards fear-related cues like angry faces. Results also showed that androstadienone enhanced men´s approach tendency towards faces independent of emotional expressions. This observation might be explained by androstadienone´s former shown ability to improve attractiveness ratings of other persons. In this regard, the endogenous odor might enhance evaluations of faces in men and, thus, might improve their willingness to approach social stimuli. In contrast to men, women already showed in the control condition higher approach tendency towards faces. Therefore, androstadienone might rather maintain than enhance the approach score in women. In the second study event-related brain potentials (ERPs) triggered by social and non-social visual stimuli were investigated by means of electroencephalography. In a double-blind between-subjects design 51 women participated. Twenty-eight women inhaled androstadienone, whereas 23 women inhaled a control solution. Four different picture categories, i.e. real faces, pictures with couples, pictures with social and non-social scenes, each including three different valence categories, i.e. positive, negative and neutral, should clarify the stimulus type or context androstadienone is acting on. The androstadienone compared to the control odor did not influence brain responses significantly. Explorative analyses, however, suggested that androstadienone influences the processing of faces. While in the control group angry faces elicited larger P300 amplitudes than happy faces, the androstadienone group showed similar P300 amplitudes concerning all emotional expressions. This observation tentatively indicates that the endogenous odor might indeed affect the neuronal responses to emotional facial stimuli, especially late components reflecting evaluative processes. However, this observation has to be verified and further investigated, in particular whether androstadienone caused reduced responses to angry faces or enhanced responses to happy faces. The third study investigated androstadienone effects on face processing especially in men. ERPs elicited by happy, angry and neutral cartoon faces, which were presented on a computer screen, were measured while 16 men, not knowing the applicated odor, inhaled either androstadienone or a control solution. Exposure to androstadienone significantly increased later neuronal responses, the P300 amplitude. This belated component of the ERP reflects attention allocation and evaluative processes towards important stimuli. Therefore, androstadienone might facilitate central nervous face processing by enhancing attention towards these stimuli. In sum, the current results corroborate the notion of androstadienone as an active social chemosignal. In minute amounts and not detectable as an odor it influenced cortical and motoric reactions. Therefore, it might be concluded that androstadienone indeed affects cognitive functions like attentional processes and in turn affects our behavior. The current results further support the notion that androstadienone acts like a human modulator pheromone, namely modulating ongoing behavior or a psychological reaction to a particular context, changing stimulus sensitivity, salience and sensory-motor integration. However, these conclusions remain tentative until further replication takes place, best in ecologically valid environments. Furthermore, one has to keep in mind that the current studies could not replicate several previous findings and could not verify some hypotheses assuming communicative effects of androstadienone. Thus, the main assumption of this thesis that androstadienone is an active chemosignal is still challenged. Also, whether the term “pheromone” is indeed suitable to label androstadienone remains open.
Investigation on Distinct Roles of Smad Proteins in Mediating Bone Morphogenetic Proteins Signals
(2011)
Bone morphogenetic proteins (BMPs) belong to the transforming growth factor-β (TGF-β) superfamily and play important roles in numerous biological events in the development of almost all multi-cellular organisms. Dysregulated BMP signaling is the underlying causes of numerous heritable and non-heritable human diseases including cancer. The vast range of biological responses induced by BMPs converges on three closely related Smad proteins that convey intracellular signals from BMP receptors to the nucleus. The specificity of BMP signaling has been intensively investigated at the level of ligand-receptor interactions, but how the different Smad proteins contribute to differential signals elicited by BMPs remains unclear. In this work, we investigated the BMP/Smad signaling in different aspects. In search for an appropriate fluorescence reporter in zebrafish, we compared different photo-switchable proteins and found EosFP the best candidate this model system for its fast maturation and fluorescence intensity. We modified and created appropriate vectors enabling Tol2-transposon based trangenesis in zebrafish, with which transgenic zebrafish lines were generated. We combined fluorescence protein tagging with high resolution microscopy and investigate the dynamics of Smad proteins in model system zebrafish. We observed that Smad5 undergoes nucleo-translocation as BMP signal transmitter during zebrafish gastrulation. We explored the Smad involvement during myogenic-to-osteogenic conversion of C2C12 cell line induced by BMP4. We created transient loss-of-function of Smads by siRNA-mediated knockdowns and analyzed the effects on these coupled yet distinct procedures by quantitative real-time PCR and terminal marker staining. We found that different Smad-complex stoichiometry might be responsible for distinct cellular signals elicited by BMPs.