No description available.

Thesis (Ph. D.)--University of Oklahoma. / Bibliography: leaves 139-152.

Thesis (Ph. D.)--University of Washington, 2005. / Vita. Includes bibliographical references (leaves 214-243).

La recombinaison méiotique joue un double rôle de moteur évolutif en participant à la création d'une diversité génétique soumise à la sélection naturelle et de contrôle dans la fabrication des gamètes lors de la méiose. De plus, en association avec certains mécanismes de réparation, la recombinaison, au travers de la conversion génique biaisée manipule les fréquences alléliques au sein des populations. Les connaissances sur le fonctionnement même de ce processus ont considérablement augmenté ces dernières années faisant découvrir un processus complexe, autant dans son fonctionnement que dans son évolution. Le thème général de la thèse est l'analyse, dans un contexte évolutif, des relations entre les différents rôles et caractéristiques fonctionnelles de la recombinaison. Un modèle de la recombinaison prenant en compte des contraintes liées au contrôle de la méiose et le phénomène d'interférence a permis une comparaison entre espèces au sein des vertébrés et des non-vertébrés de même qu'une comparaison entre sexes. Par ailleurs, nous avons montré l'impact de la localisation spécifique aux sexes des points chauds de recombinaison sur l'évolution du contenu en GC des génomes de plusieurs vertébrés. Finalement, nous proposons un modèle à l'échelle de la génétique des populations, permettant d'analyser l'impact de la recombinaison sur la fréquence de mutations délétères dans les populations humaines. Cette thèse, nous l'espérons, apportera sa pierre à l'étude interdisciplinaire de la recombinaison, à la fois au sein de la biologie et par ses relations au travers de la modélisation avec l'informatique et les mathématiques. / Meiotic recombination plays several critical roles in molecular evolution. First, recombination represents a key step in the production and transmission of gametes during meiosis. Second, recombination facilitates the impact of natural selection by shuffling genomic sequences. Furthermore, the action of certain repair mechanisms during recombination affects the frequencies of alleles in populations via biased gene conversion. Lately, the numerous advancements in the study of recombination have unraveled the complexity of this process regarding both its mechanisms and evolution. The main aim of this thesis is to analyze the relationships between the different causes, characteristics, and effects of recombination from an evolutionary perspective. First, we developed a model based on the control mechanisms of meiosis and inter-crossover interference. We further used this model to compare the recombination strategies in multiple vertebrates and invertebrates, as well as between sexes. Second, we studied the impact of the sex-specific localization of recombination hotspots on the evolution of the GC content for several vertebrates. Last, we built a population genetics model to analyze the impact of recombination on the frequency of deleterious mutation in the human population.

La recombinaison méiotique initiée en prophase I de méiose génère soit des crossing-over (COs), qui sont des échanges réciproques entre segments chromosomiques, ou des conversions géniques non associées aux COs (NCOs). Les deux types d'événements se produisent dans de petites régions (moins de 10 kilobases) appelées points chauds, qui sont distribuées de manière non homogène le long des chromosomes. L'objectif de ma thèse était la caractérisation d'un point chaud de recombinaison méiotique (nommée 14a) chez Arabidopsis thaliana (i) dans différentes accessions (ii) dans le mutant msh4, un gène impliqué dans la formation des COs. Dans les deux hybrides ColxLer et ColxWs (i) 14a a un taux très élevé de COs 0,85% et 0,49%, respectivement (ii) Les COs sont regroupés dans deux petites régions de quelques kilobases, 14a1 et 14a2 avec une distribution de type gaussienne observée aux points chauds décrits dans d'autres espèces (iii) 14a1 est aussi un point chaud de NCO avec un taux aussi élevé que celui des COs (0,5%) dans ColxLer (iv) un biais de l'initiation de recombinaison a été trouvé dans 14a1 aussi bien pour les COs que les NCOs dans le fond génétique ColxLer.Une réduction de la fréquence de CO a été observée dans le mutant msh4 dans le fond génétique ColxLer à 14a1 et 14a2 (6,4% et 18,7% par rapport au sauvage). Cela confirme le rôle précédemment connu de la protéine MSH4 impliqué dans la formation de CO. La fréquence de NCO à 14a1 est similaire à celle observéedans le fond sauvage. Le rôle des H3K4 histones trimethyltransferase d’Arabidopsis dans la recombinaison méiotique (comme précédemment observé comme Set1 chez S. cerevisiae ou PRDM9 chez les mammifères) a également été étudiée. Aucun des dix gènes d’histones méthyltransférase étudié n'a montré de rôle dans la méiose. Cela pourrait être dû à (i) une forte redondance de la fonction entre les protéines (ii) une autre histone méthyltransférase en charge de l'étiquetage des points chauds de recombinaison méiotique (plus de 29 putatif histone méthyltransférase ont été identifiés dans le génome d'Arabidopsis!) (iii) contrairement à S. cerevisiae, les souris et l'homme, un autre mécanisme de contrôle épigénétique de la recombinaison méiotique. / Meiotic recombination initiated in prophase I of meiosis generates either crossovers (COs), which are reciprocal exchanges between chromosome segments, or gene conversion not associated to crossovers (NCOs). Both kinds of events occur in narrow regions (less than 10 kilobases) called hotspots, which are distributed non-homogenously along chromosomes. The aim of my PhD was the characterization of a hotspot of meiotic recombination (named 14a) in Arabidopsis thaliana (i) across different accessions (ii) in msh4 mutant, a gene involved in CO formation. In both ColxLer and ColxWs hybrids (i) 14a had a very high rate of COs 0.85% and 0.49%, respectively (ii) COs clustered in two small regions of a few kilobases, 14a1 and 14a2 with typical Gaussian curve distribution observed in other organisms (iii) 14a1 was also a hotspot of NCO with high rate (0.5%) in ColxLer (iv) a bias of recombination initiation at 14a1 CO and NCO hotspot was found in ColxLer. A reduction of CO frequency was observed in msh4 mutant in ColxLer background at 14a1 (6.4%) and 14a2 (18.7%) compared to wild type. This confirmed previously known role of MSH4 protein in CO formation. Frequency of NCO at 14a1 was similar to wild type. The role of putative Arabidopsis histone H3K4 trimethyltransferase in meiotic recombination as previously observed like Set1 in S.cerevisiae or PRDM9 in mammals (mice and human) was also studied. None of ten putative histone methyltransferase genes was involved in meiosis. This could be due to (i) a strong redundancy of function between gene products (ii) another histone methyltransferase in charge of labeling meiotic recombination hotspots (more than 29 putative histone methyltransferase have been identified in the Arabidopsis genome!) (iii) contrary to S. cerevisiae, mice and humans, another mechanism for epigenetic control of meiotic recombination

En génomique comparative, on considère classiquement trois forces déterminant l'évolution des séquences : la mutation, la sélection et la dérive génétique. Récemment, lors de l'étude de l'origine évolutive des variations de la composition en base des génomes, un quatrième agent a été identifié : la conversion génique biaisée (BGC). Le BGC est intimement lié à la recombinaison méiotique et semble présent chez la plupart des eucaryotes. Ce phénomène introduit une surreprésentation de certains allèles dans les produits méiotiques aboutissant à une augmentation de la fréquence de ces variants dans la population. Ce processus est capable de mimer et d'interférer avec la sélection naturelle. Il est donc important de le caractériser afin de pouvoir le distinguer efficacement de la sélection dans l'étude de l'adaptation à l'échelle moléculaire. C'est ce que nous nous attachons à faire dans le cadre de ce travail. Pour cela nous utilisons deux espèces modèles. Premièrement la levure Saccharomyces cerevisiae pour laquelle une carte de recombinaison haute résolution permettant l'analyse du processus de conversion, est disponible. L'étude approfondie de cette carte nous a permis de lever le voile sur les mécanismes moléculaires qui sous-tendent le BGC. Deuxièmement, grâce à des découvertes récentes sur la détermination des patrons de recombinaison via la protéine PRDM9 chez les mammifères, nous avons quantifié la dynamique et l'intensité de ce processus dans l'histoire évolutive récente de l'homme. Ces résultats nous ont permis de confirmer la place du BGC comme quatrième force d'évolution moléculaire, mais aussi de discuter de l'origine évolutive de ce phénomène / Usually, three main forces are considered when studying sequences evolution in comparative genomics : mutation, selection and genetic drift. Recently, a fourth process has been identified during the study of base composition landscapes in genomes : biased gene conversion (BGC). This phenomenon introduces an overrepresentation of certain alleles in meiosis products (gametes or spores) leading to an increase of the frequency of those variants in the population. Thus, it is able to mimic and interfere with natural selection. Hence, it is important to describe this phenomenon in order to be able to trustfully distinguish BGC and selection in the study of adaptation at the molecular scale. So, the main goal of this work is to analyze the molecular origin, the intensity and the dynamics of BGC. To do so, we use two model species. First, we use the yeast Saccharomyces cerevisiae because, for this specie, a high-resolution recombination map is available which allows a fine study of the conversion process. Analyzing this map led us to shed the light on the molecular mechanisms of BGC. Secondly, recent discoveries on the role of the PRDM9 protein in the determination of recombination landscapes in mammals allowed us to quantify the dynamics and intensity of BGC in the recent human history. Thanks to those two studies, we first confirmed that BGC is the fourth force of molecular evolution and we also provided hypotheses about the evolutionary origin of this process

Hemos analizado la variabilidad genética de una zona no codificante autosómica, el pseudogén homólogo al gen de la glucocerebrosidasa (psGBA). Parte del análisis se ha realizado desde la perspectiva de la genética de poblaciones humanas. Desde un punto de vista más genómico hemos establecido la dinámica de la región, a fin de entender las causas del espectro de variación. Hemos analizado el papel de la mutación, recombinación, conversión génica y, especialmente, selección. Por otra parte, psGBA es importante en la producción de alelos complejos GBA-psGBA, que provocan los tipos más severos de la enfermedad de Gaucher. Mostramos cómo el conocimiento de la variabilidad en psGBA ayuda al reconocimiento de estos alelos complejos. Finalmente, con los datos de variabilidad de dos regiones parálogas situadas en la misma región cromosómica (gen GBA / pseudogen psGBA) hemos comparado los patrones de mutación que presenta una misma secuencia bajo diferentes presiones selectivas. / We have analyzed the genetic variability in a non-coding autosomal region, the pseudogene homologous to the glucocerebrosidase gene (psGBA).Part of the analysis has been performed from the human populations point of view.From a more genomic perspective, we have established the region dynamics in order to understand the causes of the variability pattern. We have analyzed the role of mutation, recombination, gene conversion and, especially, selection.On the other hand, psGBA is important in the production of complex alleles GBA-psGBA, that lead to the most severe types of Gaucher disease. We show how the knowledge of psGBA variability helps to the identification of those complex alleles.Last, from the variability data from two paralogous regions located in the same chromosomal region (GBA gene /psGBA pseudogene) we have compared the mutation patterns shown by the same sequence under different selective pressures.

Primates are one of the best characterized phylogenies with vast amounts of comparative data available, including genomic sequences, gene expression, and epigenetic modifications. Thus, they provide an ideal system to study sequence evolution, regulatory evolution, epigenetic evolution as well as their interplays. Comparative studies of primate genomes can also shed light on molecular basis of human-specific traits. This dissertation is mainly composed of three chapters studying human and non-human primate evolution. The first study investigated evolutionary rate difference between sex chromosome and autosomes across diverse primate species. The second study developed an unbiased approach without the need of prior information to identify genomic segments under accelerated evolution. The third study investigated interplay between genomic and epigenomic evolution of humans and chimpanzees. Research advance 1: evolutionary rates of the X chromosome are predicted to be different from those of autosomes. A theory based on neutral mutation predicts that the X chromosome evolves slower than autosomes (slow-X evolution) because the numbers of cell division differ between spermatogenesis and oogenesis. A theory based on natural selection predicts an opposite direction (fast-X evolution) because newly arising beneficial mutations on the autosomes are usually recessive or partially recessive and not exposed to natural selection. A strong slow-X evolution is also predicted to counteract the effect of fast-X evolution. In our research, we simultaneously studied slow-X evolution, fast-X evolution as well as their interaction in a phylogeny of diverse primates. We showed that slow-X evolution exists in all the examined species, although their degrees differ, possibly due to their different life history traits such as generation times. We showed that fast-X evolution is lineage-specific and provided evidences that fast-X evolution is more evident in species with relatively weak slow-X evolution. We discussed potential contribution of various degrees of slow-X evolution on the conflicting population genetic inferences about human demography. Research advance 2: human-specific traits have long been considered to reside in the genome. There has been a surge of interest to identify genomic regions with accelerated evolution rate in the human genome. However, these studies either rely on a priori knowledge or sliding windows of arbitrary sizes. My research provided an unbiased approach based on previously developed “maximal segment” algorithm to identify genomic segments with accelerated lineage-specific substitution rate. Under this framework, we identified a large number of human genomic segments with clustered human-specific substitutions (named “maximal segments” after the algorithm). Our identified human maximal segments cover a significant amount of previously identified human accelerated regions and overlap with genes enriched in developmental processes. We demonstrated that the underlying evolutionary forces driving the maximal segments included regionally increased mutation rate, biased gene conversion and positive selection. Research advance 3: DNA methylation is one of the most common epigenetic modifications and plays a significant role in gene regulation. How DNA methylation status varies on the evolutionary timescale is not well understood. In this study, we investigated the role of genetic changes in shaping DNA methylation divergence between humans and chimpanzees in their sperm and brain, separately. We find that for orthologous promoter regions, CpG dinucleotide content difference is negatively correlated with DNA methylation level difference in the sperm but not in the brain, which may be explained by the fact that CpG depleting mutations better reflect germline DNA methylation levels. For the aligned sites of orthologous promoter regions, sequence divergence is positively correlated with methylation divergence for both tissues. We showed that the evolution of DNA methylation can be affected by various genetic factors including transposable element insertions, CpG depleting mutations and CpG generating mutations.

En aquest treball es pretén utilitzar la informació que deixa la recombinació al nostres genomes per fer inferències sobre la història evolutiva recent de les poblacions humanes. Per fer-ho, s’ha desenvolupat un mètode novedós, anomenat IRiS, que permet la detecció de recombinacions antigues específiques en un conjunt de seqüències. Hem validat extensivament IRiS i l'hem sotmès a diferents escenaris per tal d’avaluar-ne l’ eficàcia. Un cop els events de recombinació són detectats, es poden utilitzar com a marcadors genètics per estudiar els patrons de diversitat de les poblacions humanes. Finalment, hem aplicat aquesta innovadora aproximació a un conjunt de poblacions humanes del Vell Món, que varen ser genotipades específicament amb aquesta finalitat, aportant nous coneixements en la història evolutiva recent dels humans / The aim of this work is to use the information left by recombination in our genomes to make inferences on the recent evolutionary history of human populations. For that, a novel method called IRiS has been developed that allows detecting specific past recombination events in a set of extant sequences. IRiS is extensively validated and studied in whole set of different scenarios in order to assess its performance. Once recombination events are detected, they can be used as genetic markers to study the recombinational diversity patterns of human populations. We apply this innovative approach to a whole set of different human populations within the Old World that were specifically genotyped for this end and we provide new insights in the recent human evolutionary history of our species.

Orientador: Maricilda Palandi de Mello / Tese (doutorado) - Universidade Estadual de Campinas, Insituto de Campinas / Made available in DSpace on 2018-08-13T00:56:19Z (GMT). No. of bitstreams: 1 Coeli_FernandaBorchers_D.pdf: 7777269 bytes, checksum: 053a66d2bc3df6ad0d5de7c2038f3246 (MD5) Previous issue date: 2009 / Resumo: A hiperplasia congênita da adrenal é causada pela deficiência de uma das cinco enzimas responsáveis pela síntese do cortisol na esteroidogênese, sendo que mais de 90% dos casos ocorrem devido à deficiência de 21-hidroxilase (21-OH). O genoma haplóide humano possui duas cópias em tandem do gene que codifica para a 21-OH, denominados CYP21A2 e o CYP21A1P. Embora as duas cópias, CYP21A1P e CYP21A2, tenham aproximadamente 98% de homologia, CYP21A1P é classificado como um pseudogene, devido a algumas alterações deletérias em sua seqüência. Foram mapeados no braço curto do cromossomo 6, assim como os genes RP, C4 e TNX também duplicados em tandem. Este loco é denominado modulo RCCX, onde cada letra representa um gene. Uma conseqüência esperada deste tipo de organização é que esta favorece eventos de crossingover desigual, produzindo cromátides irmãs assimétricas e pares de gametas com um número variável de unidades completas. O crossover desigual não gera somente um tipo definido de deleção (alelos monomodulares), de duplicação (alelo trimodular) ou de conversão (alelo bimodular), mas pode, dependendo da sua exata localização, produzir um grande número de alelos diferentes, com significados funcionais variáveis. O objetivo deste trabalho foi investigar a variabilidade dos genes híbridos CYP21A21P/CYP21A2 quanto à região de recombinação nos alelos monomodulares, bimodulares e trimodulares de indivíduos com deficiência de 21- hidroxilase. Foram incluídos 55 pacientes com deficiência de 21 - hidroxilase, que foram avaliados por Southern blot, Multiplex ligation-dependent probe amplification (MLPA), PCR - Alelo especifico (ASO-PCR) e seqüenciamento. Na triagem por Southern blot foram identificados 26 alelos mono-, 26 bi- e 5 trimodulares com prováveis genes híbridos. Foi identificado um alelo monomodular novo portador da variante C4A [6,4 kb] que se mostrou único inclusive quanto à formação híbrida CYP21A21P/CYP21A2. Com a técnica de MLPA foi possível mapear três regiões principais de recombinação dos genes híbridos CYP21A21P/CYP21A2 nas três configurações alélicas. Além disso, foram identificados possíveis híbridos dos genes C4A e B, tanto nas configurações mono quanto nas bimodulares. Assim, ficaram definidos 5 haplótipos monomodulares, 7 bimodulares e 3 trimodulares. As técnicas de ASO-PCR e sequenciamento para análise dos híbridos CYP21A21P/CYP21A2 e dos CYP21A21P nos alelos bi e trimodulares refinaram a caracterização subdividindo estes haplótipos em 10 mono, 15 bi e 5 trimodulares. Dado o alto grau de variabilidade encontrado não foi possível se identificar efeito fundador de nenhum haplótipo específico para a deficiência de 21-hidroxilase. Por outro lado, um haplótipo novo correspondendo a cerca de 15% dos monomodulares foi caracterizado como portador das mutações p.P34L e p.H62L e um haplótipo igualmente não descrito portador da p.H62L foi encontrado entre os bimodulares. SNPs no terminal 5'UTR, no íntron 2 e no éxon 7 responderam pela diferenciação principal entre os híbridos tanto nos haplótipos de mesmo grupo como na comparação entre os de grupos diferentes. Este trabalho indica que a combinação de quatro técnicas e o estudo de segregação nas famílias foram fundamentais para o esclarecimento dos genótipos dos pacientes. Os genes híbridos podem estar relacionados às formas clínicas perdedora de sal, não perdedora de sal e não clássica dependendo da região onde ocorre a recombinação para sua formação. / Abstract: Congenital adrenal hyperplasia is caused by deficiency of one of the five enzymes responsible for cortisol synthesis in the steroidogenesis. More than 90% of the cases occur due to deficiency of 21-hidroxilase (21-OH). The haploid human genome bears two copies in tandem of 21-OH coding gene, CYP21A2 and CYP21A1P. Although the two copies are approximately 98% homologous, CYP21A1P is a pseudogene, due to some deleterious mutations. They map to the short arm of chromosome 6, as well as RP, C4 and TNX genes which are also duplicated in tandem. This locus is called RCCX module, each letter representing one gene. An expected consequence of such organization is that it favors events of unequal crossing-overs, producing pairs of gametes with different number of complete units. The aim of this investigation was to estimate the variability of CYP21A21P/CYP21A2 chimeric genes based on the region of recombination in the monomodular, bimodular and trimodular alleles in patients with 21-hydroxylase deficiency. Fifty-five patients were included for Southern blot, Multiplex ligationdependent probe amplification (MLPA), Allele-specific PCR (ASO-PCR) and sequencing analyses. Southern blot identified alleles which were: mono (n = 26), bi (n = 26) and trimodular (n = 5) with chimeric genes. A novel monomodular allele was identified that carry C4A [6,4 kb] variant and also bore an unique CYP21A21P/CYP21A2 formation. MLPA technique mapped three main recombination regions in CYP21A21P/CYP21A2 chimerical genes in the three RCCX configurations. Moreover, it indicated possible chimeric C4A and B genes in both mono- and bimodular configurations. Therefore, five mono-, seven bi- and three trimodular haplotypes had been defined. Both ASO-PCR and sequencing techniques for CYP21A21P/CYP21A2 and CYP21A21P analysis had refined the characterization subdividing these haplotypes in ten mono-, fifteen bi- and five trimodular. Considering the high degree of variability observed it was not possible to identify a founder effect of any specific haplotype for the deficiency of 21-hidroxilase. Conversely, a novel haplotype corresponding to about 15% of the monomodular alleles was characterized as carrying the mutations p.P34L and p.H62L and, similarly one haplotype carrying the p.H62L was found among bimodular alleles. SNPs in the 5 ' UTR, intron 2 and exon 7 were responsible for the main differentiation among chimerical genes within a group as well as upon comparison between different groups. The results presented here indicate that the combination of four different techniques and the study of segregation in the families had been essential for defining the genotypes of the patients. It is also shown that CYP21A21P/CYP21A2 chimeric genes can be related to different clinical forms: salt losing, non-salt losing and non-classical depending on the region where the recombination for its formation occurs. / Doutorado / Genetica Animal e Evolução / Doutor em Genetica e Biologia Molecular