Smith-Magenis syndrome (SMS) is a multiple congenital abnormalities intellectual disability syndrome that results from a deletion of chromosome 17p11.2 or mutation of the retinoic acid inducted one gene (RAI1). SMS is characterized by a multitude of phenotypic features including craniofacial defects, short stature, obesity, intellectual disability, self-abusive behavior, sleep disturbance and behavioral abnormalities. Interestingly, although SMS is a clearly defined syndrome with a known molecular change at its foundation, ~40% of all candidate cases sent to the Elsea lab for evaluation do not have a mutation or deletion of RAI1. We hypothesize that at least one other locus must be responsible for this Smith-Magenis-like (SMS-like) phenotype. To address this hypothesis, we first compiled a cohort of 52 subjects who had been referred to the Elsea lab for a clinical diagnosis of SMS. Once these individuals were confirmed to not have an RAI1 mutation or deletion, their phenotypes were compiled and statically analyzed to distinguish whether SMS and SMS-like cohorts are different in the prevalence of the core phenotypes of SMS such as, but not limited to, sleep disturbance, self-abusive behavior and developmental delay. SMS-like and SMS cohorts are not different in prevalence for these core features. Next, all SMS-like subjects were sent for whole genome array comparative genomic hybridization (aCGH) to identify duplications or deletions of each individual’s genome which contribute to the phenotype observed. We identified 6 pathogenic copy number variants (CNVs) in six individuals which contribute directly to the clinical phenotype, including two del(2)(q37). This study enabled us to draw relationships between SMS and other syndromes that had never been appreciated before and helped to identify pathways in which RAI1 may function. Using the data from our SMS-like study we were able to further characterize two known syndromes; Deletion 2q37 syndrome (brachydactyly mental retardation syndrome) and deletion 2q23 syndrome. With regard to deletion 2q37, syndrome we used genomic data from known and new deletion 2q37 subjects to refine the critical region to one gene: the histone deacetylase 4 gene (HDAC4). Using both clinical and molecular clues, we were able to identify one subject from our SMS-like cohort who has an insertion in HDAC4 which results in a premature stop codon. We conclude from this study that mutation of HDAC4 results in brachydactyly mental retardation syndrome. With regard to deletion 2q23 syndrome there were only five known cases in the published literature to which we were able to add two more. Using as similar approach to our del2q37 study we refined the critical region for this syndrome to one gene, the methyl binding domain 5 gene (MBD5). Using a molecular and clinical approach we were able to conclude that haploinsufficiency of MBD5 results in the core phenotypes seen in del2q23 syndrome including microcephaly, intellectual disabilities, severe speech impairment, and seizures. Using all the data generated from the three previous studies we set out to characterize the molecular function of RAI1. We hypothesize that RAI1 is a transcription factor that regulates gene expression of core genes involved in development, neurological function, and circadian rhythm. Using a ChIP-chip based approach we identified 257 transcripts we believe RAI1 regulates. Following up on these transcripts, using in vitro and in vivo methods, we have been able to conclude that RAI1 is a positive regulator of CLOCK, the master regulator of the central circadian cycle. Taken together, these studies have given us insight into the specific molecular changes that contribute to SMS and SMS-like syndromes. We have unveiled pathways and genes which are important to normal human development and behavior and identified novel functions of RAI1. These studies will provide the foundation for the future discovery of the pathways affected.

Le succès écologique des organismes dépend principalement de leur phénotype. Une composante important du phénotype est la morphologie fonctionnelle car elle influence la performance d’un organisme donné dans un milieu donné et donc reflète son écologie. Des disparités dans la morphologie fonctionnelle ou dans le développement entre espèces peuvent donc mener à des différences écologiques. Ce projet évalue le rôle des mécanismes de variation morphologique dans la production de différences écologiques entre espèces au sein des poissons hybrides du complexe Chrosomus eos-neogaeus. En utilisant la microtomodensitométrie à rayons X et la morphométrie géométrique 3D, la forme des éléments des mâchoires est décrite pour comparer la variation morphologique et les différences développementales entre les membres du complexe C. eos neogaeus. Les hybrides présentent autant de variation phénotypique que les espèces parentales et présentent des phénotypes nouveaux, dit transgressifs. Les hybrides présentent aussi des différences marquées avec les espèces parentales dans leur allométrie et dans leur intégration phénotypique. Finalement, ceux-ci semblent être plastiques et en mesure de modifier leur phénotype pour occuper plusieurs environnements. L’entièreté de ces résultats suggère que des changements dans le développement des hybrides entraînent une différenciation phénotypique et écologique avec les espèces parentales. / An organism’s success is highly dependent on its phenotype. A major component of the phenotype is functional morphology because of its role in determining performance in a given environment clues as to their ecology. Morphological disparity and developmental differences between species can thus lead to ecological differences. This project assesses the role that mechanisms capable of generating morphological variation have in producing ecological differences between species within the hybrid fishes of the Chrosomus eos-neogaeus complex. Using X-ray based computer-aided micro-tomography and 3D geometric morphometrics, the shape of bones composing the jaws was described to measure morphological variation and developmental differences in the C. eos-neogaeus complex. Chrosomus eos-neogaeus hybrids displayed as much variation as parental species and novel, deemed transgressive, phenotypes. Hybrids were also markedly different from parental species in their patterns of allometric scaling and phenotypic integration. Finally, hybrids appeared to be plastic and able to acclimate to different environments. The results of this study indicate that changes in the development of hybrids could lead to phenotypic and, in turn, ecological differences with parental species.

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Les chênes sessiles et pédonculés sont deux espèces sympatriques interfertiles occupant des places distinctes dans la succession écologique. Elles constituent pour cela un bon modèle pour l’étude de la spéciation écologique. Malgré leur écologie contrastée, les deux espèces de chênes coexistent naturellement dans de nombreuses forêts, rendant possible l’étude de leur système de reproduction intra- et interspécifique. Des travaux précédents ont suggéré que l’hybridation entre ces deux espèces serait fréquence-dépendante. Elle dépendrait de la proportion de pollen allospécifique (c'est-à-dire de pollen de l’autre espèce) que reçoit l’arbre mère étudié. Ce phénomène d’hybridation fréquence-dépendante est appelé effet Hubbs, du nom d’un ichtyologue qui découvrit ce mécanisme en 1955. Mon travail a consisté à étudier, dans une parcelle mixte de chênes, les barrières à l’hybridation qui permettent la coexistence de ces deux espèces. Pour cela, j’ai effectué une étude de paternité de grande ampleur (près de 3500 individus typés sur 12 marqueurs microsatellites). Tout d’abord, je me suis intéressée à la délimitation des deux espèces en appliquant pour la première fois in situ le critère d’interfertilité. Je me suis ensuite concentrée sur les facteurs qui influencent l’hybridation au travers d’une modélisation des croisements à l’échelle de l’individu. Les résultats montrent que le maintien de ces deux espèces est régi par deux composantes environnementales importantes : la fréquence de chaque espèce et leur distribution, qui influencent la quantité de pollen reçue. Grâce à cette étude empirique et à l’approche de modélisation de ces croisements, nous avons désormais une bien meilleure vision de l’effet de l’environnement sur l’hybridation. J’ai par la suite comparé les caractéristiques du système de reproduction de chaque espèce (dispersion du pollen et fécondité mâle) en cherchant si un lien existait avec leur stratégie écologique. Les résultats suggèrent que les différences de dynamique écologique pourraient être à l’origine de la spéciation du fait de l’existence de compromis différents en termes d’allocation de ressources mais qu’à plus court terme la stabilité de l’environnement est essentielle au maintien des espèces. / Quercus petraea and Q. robur are two interfertile sympatric species. They occupy distinct stages during forest succession and constitute therefore good models for ecological speciation studies. Despite their differences, they often grow together in mixed stands, allowing the study of their intra- and interspecific reproductive system. Hybridization between these two oak species has been suggested to be frequency-dependent. The effect of the relative species abundance on hybridization is known as the “Hubbs’ effect”, from the name of an ichthyologist who described this mechanism in 1955. My work was to study the processes that limit hybridization between these two species, thereby allowing their coexistence. I conducted an extensive paternity analysis (almost 3500 individuals genotyped at 12 microsatellite markers). First, I tried to delimitate the two oak species by applying for the first time in situ the interfertility criterion. Then, I focused on interspecific crosses by studying those factors influencing hybridization. Results showed that the maintenance of these two species depends on the frequency of each species and their distribution, as both factors influence the quantity of pollen received by female flowers. Thanks to this empirical study and to this modeling approach, we have now a much better view of environmental effects on hybridization. I then compared the characteristics of the reproductive system of each species (pollen dispersal and male fertility) in relation with their ecological strategies. The results suggest that differences in species ecological dynamics are at the origin of the speciation process but that at a finer scale the stability of the environment is crucial for species’ maintenance.

This thesis deals with interspecific hybridization in sympatric populations of Pulsatilla patens and P. pratensis. The key aim was to assess karyological and phenotypic variation in the study group. DNA flow cytometry revealed three non-overlapping groups of genome sizes, corresponding to P. patens, P. × hackelii and P. pratensis. The frequency of interspecific hybridization was low, accounting to approx. 3% of analyzed plants. Morphometric analyses identified a set a reliable species-specific characters, including number of divisions the leaf, length of petiole the part of leaf, number of smaller parts of leaf and colour of blossom. Although the majority of hybrid individuals flowered and set fruits, their fertility was reduced considerably. The threat from interspecific hybridization to parental species is therefore rather low.

Master thesis investigates reproductive barriers in diploid-polyploid complex of Vicia cracca. Complex with basic chromosome number x=7 consists of diploid (2x=14), tetraploid (4x=28) and rare triploid (3x=21) cytotype. I studied prereproducitve barriers between diploid and tetraploid cytotype: phenology of flowering, pollinators' behavior (preference of species of pollinators to cytotypes, sequence of visited cytotypes), variables, that could explain pollinators' behavior (amount of nectar as the main reward, size and amount of pollen grains as a potentional reward). To find out how strong the triploid block is I analyzed ploidy of seeds and seedlings from mixed-ploidy population. The habitat isolation showed up to be the strongest reproductive barrier. Pollinator's behavior meaningfully contributes to isolation, phenology of flowering contributes only minimally. Index expressing rate of prereproctive barriers is 0,956. Pollinator Bombus pascuorum visited on one locality preferably tetraploid plants and Andrena sp. preferred diploid plants. Even though tetraploid plants produce more nectar, no other analysis showed pollinators' preference to tetraploid plants. I prepared one squash of diploid V. cracca using method of in situ hybridization. This method needs to be optimilized for studied taxon.

The presented study tries to reveal the evolutionary history of polyploid complex Galium pusillum aiming on a rare czech endemic species G. sudeticum, an ideal model to study allopatric speciation in the context of Central European (post)glacial development. The species G. sudeticum grows in Krkonose Mts. and on serpentines in western Bohemia (Slavkovský les) more than 200 kilometers away. Our morphological and molecular data suggest a clearly different history of its two isolated areas. The populations in Krkonose Mts. are probable relics of mountainous species G. anisophyllon which was there on its northern border of occurence and hybridized with lowland species G. valdepilosum during postglacial vegetations shifts. This fact is well supported by intermediate position of Krkonose Mts. populations in both morphological and molecular AFLP analyses and by sharing the same chloroplast haplotype with geographically close lowland populations. On the other hand the serpentine lowland populations in western Bohemia are both morphologicaly and genetically indistinguishable from G. valdepilosum. Distinct genetic lineage of few populations belonging to G. valdepilosum (incl. one traditionaly referred as G. sudeticum) was found in western Bohemia and Bavaria, located on relic stands (calcareous and serpentine...

Master thesis investigates reproductive barriers in diploid-polyploid complex of Vicia cracca. Complex with basic chromosome number x=7 consists of diploid (2x=14), tetraploid (4x=28) and rare triploid (3x=21) cytotype. I studied prezygotic barriers between diploid and tetraploid cytotype: phenology of flowering, pollinators' behavior (preference of spieces of pollinators to cytotypes, sequence of visited cytotypes), variables, that could explain pollinators' behavior (amount of nectar as the main reward, size and amount of pollen grains as a potentional reward). To find out how strong the triploid block is, I analyzed ploidy of seeds and seedlings from mixed-ploidy population. The habitat isolation showed up to be the strongest reproductive barrier. Pollinator's behavior meaningfully contributes to isolation, phenology of flowering contributes only minimally. Index expressing rate of prereproductive barriers is 0,990. Pollinator Bombus pascuorum visited on one locality preferably tetraploid plants and Andrena sp. preferred diploid plants. Even though tetraploid plants produce more nectar, no other analysis showed pollinators' preference to tetraploid plants.

Depuis les années 1990, l'aulne glutineux, espèce clé des ripisylves, est affecté par un oomycète qui cause son dépérissement : Phytophthora alni subsp. alni (Paa). La genèse de Paa est liée à un événement d'hybridation interspécifique entre deux espèces proches, improprement nommées P. alni subsp. uniformis (Pau) et P. alni subsp. multiformis (Pam), car initialement considérées comme des variants de Paa. L'objectif de cette thèse était d'identifier les facteurs ayant pu jouer un rôle dans l'émergence de la maladie en Europe. Par une approche de génétique des populations, nous avons montré que Pau est une espèce invasive en Europe, probablement originaire d'Amérique du Nord. Après son introduction, l'hybridation de Pau avec Pam serait l'un des facteurs essentiels de l'apparition de Paa. Nos résultats confirment que Paa aurait été généré suite à des hybridations récurrentes, qui ont structuré géographiquement les populations en Europe. L'analyse de la variabilité génétique de Paa, révélée par des marqueurs microsatellites, a toutefois montré un faible polymorphisme, avec un génotype dominant largement répandu en Europe. Par ailleurs grâce à la modélisation, nos résultats ont établi que le dépérissement du houppier des aulnes est lié à la température. En particulier l'incidence de la maladie augmente lors des hivers doux, qui pourraient favoriser la survie du mycélium de Paa, celui-ci ne présentant pas de structure de survie hivernale (chlamydospores ou oospores). La température estivale joue également un rôle, plus complexe à interpréter. On constate en effet que l'incidence de la maladie diminue avec l'augmentation des températures estivales, mais ce phénomène pourrait dépendre d'autres facteurs tels que l'état physiologique des arbres ou le type de communautés microbiologiques présentes dans les sols. Inversement, le phénomène de guérison des arbres est favorisé par des températures hivernales basses et par des températures estivales élevées. L'émergence de la maladie ne peut pas être expliquée par le changement climatique. Cependant, une augmentation des températures hivernales dans le futur dans le cadre du changement climatique aggraverait très probablement l'épidémie / Since the early 1990's alder decline caused by the oomycete Phytophthora alni subsp. alni (Paa) is one of the most important threats to riparian ecosystems in Europe. The emergence of Paa is related to an interspecific hybridization event between two related species -initially considered as Paa variants- misnamed as Phytophthora alni subsp. uniformis (Pau) and Phytophthora alni subsp. multiformis (Pam). The objective of this thesis was to identify the factors that may have contributed to the emergence of the disease in Europe. Following a population genetics approach we showed that Pau is likely to be an invasive species in Europe, probably native to North America. Its introduction would have enabled hybridization with Pam and, consequently be a major cause on the emergence of Paa. Our results confirm that Paa has arisen from several hybridization events, which have geographically structured its European populations. Paa's genetic variability, revealed by microsatellite markers, showed low levels of polymorphism, with a dominant genotype scattered throughout Europe. In addition, a modelling approach revealed that alders' crown decline is linked to temperature. In particular, the disease incidence increases during mild winters which favours mycelium survival as Paa does not produce resistant spores (chlamydospores or oospores). The effect of summer temperatures is more complex to explain. Disease incidence decreases when summer temperatures are higher, but this phenomenon can also be linked to the physiological conditions of trees or changes in soil microbiological communities. Conversely, tree recovery is favoured by lower winter and higher summer temperatures. Climate change does not explain the emergence of the disease. However, increases in winter temperatures du to climate change may strengthen the epidemic

Acrocephalus warblers have gone through the adaptive radiation during last severalmillion years, which gave rise to thirty one species occupying mainly Eurasia,Africa and Australia. The majority of species are morphologically very similar,however, they differ in ecological requirements, migration strategy, and song.Interspecific hybridization seems to be quite common among Acrocephaluswarblers, not only between sister species, but also between more distantly relatedtaxa. The main goal of this study was to determine whether this hybridization leadsto gene flow between species and which factors affect the rate of interspecific geneflow. For this purpose we conducted population-genetic analysis in three Europeanspecies of the Acrocephalus warblers of the subgenus Notiocichla: reed warbler (A.scirpaceus), marsh warbler (A. palustris), and blyth's reed warbler (A. dumetorum).Our results based on the analysis of sequence data from eight nuclear loci indicate,that gene flow between the studied species occurs, but only in one direction. Thegene flow is higher between genetically more related species than betweenecologically more similar species. We also estimated that the reed warbler and themarsh warbler diverged approximately 1,1 million years ago. The blyth's warblerand ancestor of the reed and marsh warbler...