Chez les organismes sexués, des suppressions de recombinaison peuvent évoluer dans certaines régions génomiques pour conserver des combinaisons d’allèles bénéfiques, ce qui aboutit à la transmission de plusieurs gènes en un seul locus, alors appelé « supergène ». Les supergènes déterminent des phénotypes complexes, comme l’identité sexuelle chez les organismes qui ont des chromosomes sexuels. Sur certains chromosomes sexuels, la région sans recombinaison s’est étendue plusieurs fois successivement, produisant des « strates évolutives ». Il est communément admis que ces strates évolutives sont issues de liaisons successives de gènes sexuellement antagonistes (qui ont des allèles bénéfiques à un sexe mais délétère à l’autre) à la région qui détermine le sexe, mais peu de preuves empiriques soutiennent cette hypothèse. Les champignons constituent des modèles intéressants pour étudier les causes évolutives des suppressions de recombinaison parce qu’ils peuvent avoir des chromosomes de types sexuels non recombinants sans être associés à des fonctions mâles ou femelles. Dans cette thèse, nous avons étudié l’évolution de la suppression de recombinaison sur les chromosomes de type sexuel chez les champignons castrateurs de plantes du genre Microbotryum. Chez les champignons Microbotryum, les croisements ne sont possibles qu’entre des gamètes qui ont des allèles distincts aux deux locus de types sexuels. Nous avons montré que les suppressions de recombinaison ont évolué plusieurs fois indépendamment pour lier les deux locus de types sexuels, depuis l’état ancestral avec les locus de types sexuels situés sur deux chromosomes différents. La suppression de recombinaison a soit lié les locus de types sexuels à leur centromère respectif, ou a lié les locus de types sexuels entre eux après que des réarrangements chromosomiques, différents dans les différentes espèces, les aient amenés sur le même chromosome. Les deux sortes de suppression de recombinaison sont bénéfiques sous le mode de reproduction par auto-fécondation intra-tétrade de Microbotryum, parce qu’ils augmentent le taux de compatibilité entre gamètes. Les suppressions de recombinaison ont donc évolué plusieurs fois indépendamment via des chemins évolutifs et des changements génomiques différents, ce qui renseigne sur la répétabilité de l’évolution. De plus, nous avons révélé l’existence de strates évolutives sur les chromosomes de type sexuels de plusieurs espèces de Microbotryum, ce qui remet en cause le rôle de l’antagonisme sexuel dans la formation de strates évolutives, les types sexuels n’étant pas associés à des fonctions mâles / femelles. Des études précédentes ont rapporté peu de différences phénotypiques associées aux types sexuels, ce qui rend peu probable qu’une sélection antagoniste existe entre types sexuels sur de nombreux gènes (l’existence de gènes avec des allèles bénéfiques à un type sexuel mais délétère à l’autre). Certains gènes situés dans les régions non-recombinantes des chromosomes de types sexuels étaient différentiellement exprimés entre types sexuels, mais nos analyses suggèrent qu’un tel différentiel d’expression peut être dû à la dégénérescence. En effet, des mutations délétères s’accumulent dans les régions non-recombinantes, ce qui peut modifier l’expression des gènes ou les séquences protéiques. Nous avons donc conclu que la sélection antagoniste ne peut pas expliquer la formation des strates évolutives chez les champignons Microbotryum. Par conséquent, des mécanismes alternatifs doivent être considérés pour expliquer l’extension progressive des régions non-recombinantes, et ces mécanismes pourraient aussi générer des strates évolutives sur les chromosomes sexuels. Ces travaux incitent de futures études à d’une part identifier d’autres strates évolutives qui ne sont pas associées à des fonctions mâles/femelles, et d’autre part à identifier leurs causes évolutives et leurs conséquences en termes de dégénérescence. / In sexual organisms, recombination suppression can evolve in specific genomic regions to protect beneficial allelic combinations, resulting in the transmission of multiple genes as a single locus, which is called a supergene. Supergenes determine complex phenotypes, such as gender in organisms with sex chromosomes. Some sex chromosomes display successive steps of recombination suppression known as “evolutionary strata”, which are commonly thought to result from the successive linkage of sexually antagonistic genes (i.e. alleles beneficial to one sex but detrimental to the other) to the sex-determining region. There has however been little empirical evidence supporting this hypothesis. Fungi constitute interesting models for studying the evolutionary causes of recombination suppression in sex-related chromosomes, as they can display non-recombining mating-type chromosomes not associated with male/female functions. Here, we studied the evolution of recombination suppression on mating-type chromosomes in the Microbotryum plant-castrating fungi using comparative genomic approaches. In Microbotryum fungi, mating occurs between gametes with distinct alleles at the two mating-type loci, as is typical of basidiomycete fungi. We showed that recombination suppression evolved multiple times independently to link the two mating-type loci from an ancestral state with mating-type loci on two distinct chromosomes. Recombination suppression either linked the mating-type genes to their respective centromere or linked mating-type loci after they were brought onto the same chromosome through genomic rearrangements that differed between species. Both types of linkage are beneficial under the intra-tetrad mating system of Microbotryum fungi as they increase the odds of gamete compatibility. Recombination suppression thus evolved multiple times through distinct evolutionary pathways and distinct genomic changes, which give insights about the repeatability and predictability of evolution. We also reported the existence of independent evolutionary strata on the mating-type chromosomes of several Microbotryum species, which questions the role of sexual antagonism in the stepwise extension of non-recombining regions because mating-types are not associated with male/female functions. Previous studies reported little phenotypic differences associated to mating-types, rending unlikely any antagonistic selection between mating types (i.e. “mating-type antagonism”, with genes having alleles beneficial to one mating-type but detrimental to the other). The genes located in non-recombining regions on the mating-type chromosomes can be differentially expressed between mating types, but our analyses indicated that such differential expression was more likely to result from genomic degeneration than from mating-type antagonism. Deleterious mutations are indeed known to accumulate in non-recombining regions resulting in modifications of gene expression or of protein sequence. We concluded that antagonistic selection cannot explain the formation of evolutionary strata in Microbotryum fungi. Alternative mechanisms must be therefore be considered to explain the stepwise expansion of non-recombining regions, and they could also be important on sex chromosomes. This work thus prompts for future studies to identify further evolutionary strata not associated with male/female functions as well as to elucidate their evolutionary causes and consequences in terms of genomic degeneration.

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<p>Sex chromosomes have evolved independently multiple times in plants and animals. Sex chromosome evolution theory predicts the gradual degeneration of the sex-specific sex chromosome due to suppression of recombination, which lowers the efficacy of natural selection. Suppressed recombination also acts to resolve sexual conflict and ensure proper segregation of sex-specific alleles. However, sex chromosome degeneration is not always the case, and evolutionarily old, and young, but homomorphic (nondegenerate) sex chromosomes have been observed. African clawed frogs (<em>Xenopus</em> and <em>Silurana</em>) have homomorphic sex chromosomes due to a recent turnover event. However, occasional recombination between the sex chromosomes may contribute to the maintenance of homomorphic sex chromosomes in African clawed frogs. Mechanisms that prevent divergence of sex chromosomes may be related to polyploidization, which is frequently observed in African clawed frogs. The studies herein construct a phylogenetic framework to test alternative hypotheses for selection on sex-linked and autosomal genes involved in sex determination, map sex chromosomes and compare sex chromosomes across African clawed frogs. I have also explored the relationship between phenomena like recent turnover events, recombination and polyploidization to sex chromosome degeneration (or lack thereof). In this dissertation, I have discussed the potential for multiple mechanisms of sex determination and the unique pseudoautosomal nature of sex chromosomes within this group of frogs. This body of work provides a comprehensive study of sex chromosomes in a group lacking phylogenetic resolution and sheds light on the origin and evolution of sex chromosomes in other organisms.</p> / Doctor of Philosophy (PhD)

<div>Chromosomal mechanisms of sex determination vary greatly in phylogenetically closely related species, indicative of rapid evolutionary rates. Sex chromosome karyotypes are generally conserved within families; however, many species have derived sex chromosome configurations. Insects display a plethora of sex chromosome systems due to rapid diversification caused by changes in evolutionary processes within and between species. A good example of such a system are insects in the blow fly family Calliphoridae. While cytogenetic studies observe that the karyotype in blow flies is highly conserved (five pairs of autosomal chromosomes and one pair sex chromosome), there is variation in sex determining mechanisms and sex chromosome structure within closely related species in blow flies. The evolutionary history of sex chromosomes in blow fly species have not been fully explored. Therefore, the objective of this research was to characterize the sex chromosome structures in four species of blow flies and investigate the selective forces which have played a role in shaping the diverse sex chromosome system observed in blow flies. The blow fly species used in this study are Phormia regina, Lucilia cuprina, Chrysomya rufifacies and Chrysomya albiceps. Phormia regina,and Lucilia cuprina have a heteromorphic sex chromosome system and are amphogenic (females produce both male and female offspring in equal ratio). In contrast, Chrysomya rufifacies and Chrysomya albiceps, have a homomorphic sex chromosome system, are monogenic (females produce unisexual progeny), have two types of females (arrhenogenic females – male producers and thelygenic females – female producers), and sex of the offspring is determined by the maternal genotype. </div><div>To accomplish these tasks, a total of nine male and female individual draft genomes for each of the four species (including three individual draft genomes of Chrysomya rufifacies – male, and the two females) were sequenced and assembled providing genomic data to explore sex chromosome evolution in blow flies. Whole genome analysis was utilized to characterize and identify putative sex chromosomal sequences of the four blow fly species. Genomic evidence confirmed the presence of genetically differentiated sex chromosomes in P. regina and L. cuprina; and genetically undifferentiated sex chromosomes in C. rufifacies and C. albiceps. Furthermore, comparative analysis of the ancestral Dipteran sex chromosome (Muller element F in Drosophila) was determined to be X-linked in P. regina and L. cuprina contributing to sex chromosome differentiation but not sex-linked in C. rufifacies and C. albiceps. Evolutionary pressures are often quantified by the ratio of substitution rates at non-synonymous (dN) and synonymous (dS) sites. Substitution rate ratio analysis (dN/dS) of homologous genes indicated a weaker purifying selection may have contributed to the loss of sex-linked genes in Muller element F genes of the undifferentiated sex chromosome as compared to the differentiated sex chromosome system. Overall, the results presented herein greatly expands our knowledge in sex chromosome evolution within blow flies and will reinforce the study of sex chromosome evolution in other species with diverse sex chromosome systems.</div><div><br></div>

Thesis (MScMedSc (Obstetrics and Gynaecology))--University of Stellenbosch, 2005. / CHAPTER 1 presents comprehensive background information regarding all aspects addressed in this thesis. Special attention was given to literature on paternal influences on embryonic development, the role of sperm RNA, sperm chromatin and sperm functional aspects i.e. morphology and acrosomal status and size. The experimental design and all relevant methods used during the study as well as the material that were used are presented in CHAPTER 2. The results of the different techniques and evaluations are provided in CHAPTER 3. It was found that 70% of the embryos that showed no developmental potential were Y-chromosome bearing embryos. The sperm selection process for ICSI based on the approach of choosing the “best looking“ spermatozoon in the ejaculate seem to provide cells that can be classified as normal based on the length width ratio set by the WHO for normal cells. The chromatin packaging quality of the sperm correlated significantly and negatively with the percentage normal cells in the ejaculates. CHAPTER 4 comprises of a general discussion of the results and short summary of the major findings during the project. The discussion section focused on the paternal influence on the embryonic development and provided a suggestion for future research that can possibly lead to the use of X-chromosome bearing sperm in case of severe male factor cases. CHAPTER 5 contains the bibliographical information of the study.

Sex chromosomes experience distinct evolutionary environments, due to their unusual pattern of inheritance, and studies of sex chromosome evolution can shed light on the fundamental evolutionary forces acting across the genome as a whole. Here, I combine genomic and transcriptomic data across a wide range of avian species to explore the evolutionary processes governing sex chromosome evolution. Birds are female heterogametic and therefore it is possible, via comparisons with male heterogametic species, to identify the fundamental factors driving sex chromosome evolution, versus those associated with sex. In this thesis, I uncover a complex mosaic of recombination suppression between the Z and W chromosomes, characterized by repeated and independent divergence of gametologs, together with ongoing genetic exchange. Additionally, I highlight the role of mating system, and interplay between evolutionary forces, in driving coding and expression evolution on the Z and W chromosomes. My findings indicate that although the Z chromosome is masculinized for male-specific effects, the magnitude of genetic drift acting on Z-linked genes is elevated in promiscuous relative to monogamous mating systems. In contrast, evolution of the female-limited W chromosome is governed predominately by purifying selection. Together, my results suggest that the role of the Z chromosome in encoding sexual dimorphisms may be limited, but that W-linked genes play a significant role in female-specific fitness. In conclusion, my findings reveal the power of mating system in shaping broad patterns of genome evolution.

The effect of 17α-methyltestosterone a 17β-estradiol on sex of livebearer fish Poecilia reticulata and P. wingei was studied. Solution of 2.5 mg testosterone was added in the aquarium the first and fifteenth day after delivery of new fry. Estrogen was administered in food to gravid females (400 mg per 1 kg food). Sex of all individuals was determined in one, two and three months. Administration of neither testosterone nor estrogen caused sex reversal; in the second case reproduction of almost all females was stopped.

Les mécanismes de détermination génétique du sexe, qui requièrent la présence de régions chromosomiques non recombinantes ou bien de chromosomes sexuels, ont émergé de manière indépendante et répétée au sein de plusieurs lignées d'eucaryotes. La plupart des connaissances acquises dans ce domaine portent sur un nombre limité de groupes d'eucaryotes. La disponibilité d'une espèce modèle pour le groupe des algues brunes, Ectocarpus siliculosus, dont le génome a été séquencé, permet de disposer des outils nécessaires pour étudier ces mécanismes au sein d'une lignée phylogénétiquement éloignée des modèles classiquement étudiés. L'un des premiers défis a été d'identifier les chromosomes sexuels dans le génome d'E. siliculosus et de réaliser l'analyse comparative de ces structures. Par la suite, l'analyse de l'expression des gènes entre individus mâles et femelles à différents stades du cycle de vie a permis d'identifier les gènes différentiellement exprimés, de caractériser leurs fonctions et d'analyser leur évolution moléculaire. Les nombreuses données générées afin de réaliser ces différentes analyses ont permis de proposer une nouvelle version de l'assemblage du génome et de l'annotation structurale et fonctionnelle de l'ensemble des gènes codants et non-codants d'E. siliculosus. Ces différents travaux ont permis d'apporter une importante contribution sur les connaissances dans le domaine de l'analyse fonctionnelle et évolutive du déterminisme sexuel chez les algues brunes ainsi qu'une importante actualisation des ressources génomiques du modèle Ectocarpus. / Genetically determined sex determination mechanisms, which are controlled by non-recombinant chromosome regions or sex chromosomes, have emerged independently and repeatedly across several eukaryotic lineages. Most of the knowledge acquired in this area has been obtained for a limited number of eukaryotic groups. The availability of a model organism for the brown algae, Ectocarpus, whose genome has been sequenced, allows the development of tools to study these mechanisms in a lineage that is phylogenetically distant from classically studied models. One of the first challenges was to identify the sex chromosomes in Ectocarpus and to carry out a comparative analysis of these genomic structures. Analysis of gene expression in males and females at different stages of the life cycle then allowed the identification of differentially expressed genes. The functions and molecular evolution of these sex-biased genes was then studied. The large amount of data generated during the course of these analyses allowed the establishment of a new version of the genome assembly and refined structural and functional annotation of both coding and non-coding genes in Ectocarpus. This work helped made a significant contribution to knowledge in the field of functional and evolutionary analysis of sex determination in brown algae and a significantly updated the genomic resources available for the model organism Ectocarpus.

Le déterminisme génétique du sexe nécessite souvent l’évolution d’une région non-recombinante (NR) formant ainsi paire de chromosomes sexuels. Bien que la reproduction sexuée ait une origine commune à tous les eucaryotes, l’évolution des chromosomes sexuels s’est quant à elle effectuée de manière répétée et indépendante. Les chromosomes du sexe ont été particulièrement étudiés dans les systèmes diploïdes (chromosomes sexuels XY et ZW) des plantes et animaux. Le récent séquençage du génome d’Ectocarpus, modèle d’étude des algues brunes, donne non seulement une chance unique d’analyser les chromosomes sexuels dans un groupe phylogénétiquement distant des opisthocontes et de la lignée verte ; mais il donne aussi l’opportunité d’examiner un système haploïde de chromosomes sexuels (système UV). Chez Ectocarpus l’expression du sexe a lieu pendant la phase haploïde du cycle de vie, avec les chromosomes U et V, respectivement spécifiques aux femelles et aux mâles. L’analyse des chromosomes sexuels chez Ectocarpus a montré que la taille de la région NR est restée modeste pour un système vieux de plus de 70 millions d’années. Une analyse des dimorphismes sexuels a été effectuée ainsi que l’étude comparative des transcriptomes mâle et femelle d’Ectocarpus. Le développement parthénogénétique est, dans certaines populations d’Ectocarpus, un dimorphisme sexuel. Le lien génétique entre parthénogenèse et sexe a été analysé et suggère qu’un locus contrôlant la parthénogenèse est localisé au niveau de la partie recombinante du chromosome sexuel d’Ectocarpus. De plus, une analyse de fitness indique que le locus de la parthénogenèse est soumis à une sélection antagoniste entre les deux sexes. / Genetic sex determination is usually controlled by sex chromosomes carrying a non-recombining sex-determining region (SDR). Despite the common origin of sex (meiosis) in Eukaryotes, the evolution of sex chromosomes has evolved repeatedly and independently. Our knowledge in sex chromosomes comes mainly from the analysis of diploid systems (XY and ZW sex chromosomes) in animals and land plants. However the recent genome sequencing of the brown alga Ectocarpus, not only opens up the possibility of studying sex chromosomes in a phylogenetic distant group but also of analysing a haploid sex chromosome system (UV sex chromosomes). Indeed in Ectocarpus sex is expressed during the haploid phase of the life cycle, where U and V sex chromosomes are restricted to female and male, respectively. The Ectocarpus sex chromosomes have some unusual evolutionary features such as the size of the non-recombining region, which is surprisingly small for a 70 million year old system. Also the evolutionary aspect of sexual dimorphism was studied by analyzing male and female transcriptomes and by identifying several subtle sexual dimorphic traits. Parthenogenetic capacity is a sexual dimorphic trait in some populations of Ectocarpus. The genetic link between parthenogenesis and sex was analysed and a locus that controls parthenogenetic was located to the Ectocarpus sex chromosome, in the recombining pseudoautosomal region. Fitness analysis strongly suggested that the parthenogenetic locus is a sexual antagonistic locus