Polyploidy, or whole genome duplication (WGD), is a fundamental evolutionary mechanism for diverse organisms including many plants and some animals. Duplicate genes from WGD are a major source of expression and functional diversity. However, the biological and evolutionary mechanisms for gene expression changes within and between species following WGD are poorly understood. Using genome-wide gene expression microarrays and high-throughput sequencing technology, I studied the genetic and evolutionary mechanisms for gene expression changes in synthetic and natural allopolyploids that are derived from hybridization between closely related species. To investigate evolutionary fate of duplicate genes, I tested how duplicate genes respond to developmental and environmental changes within species and how ancient duplicate genes contribute to gene expression diversity in resynthesized allopolyploids. We found that expression divergence between gene duplicates was significantly higher in response to environmental stress than to developmental process. Furthermore, duplicate genes related to external stresses showed higher expression divergence between two closely related species and in resynthesized and natural allotetraploids than single-copy genes. A slow rate of expression divergence of duplicate genes during development may offer dosage-dependent selective advantage, whereas a high rate of expression divergence between gene duplicates in response to external changes may enhance adaptation. To investigate molecular mechanisms of expression diversity among allopolyploids, I analyzed high-throughput sequencing data of small RNAs in allopolyploids and their progenitors. Small interfering RNAs (siRNAs) induce epigenetic modification and gene silencing of repeats, while microRNAs (miRNAs) and trans-acting siRNAs (ta-siRNAs) induce expression modulation of protein coding genes. Our data showed that siRNA populations in progenitors were highly maintained in allopolyploids, and alteration of miRNA abundance in allopolyploids was significantly correlated with expression changes of miRNA target genes. These results suggest that stable inheritance of parental siRNAs in allopolyploids helps maintain genome stability in response to genome duplication, whereas expression diversity of miRNAs leads to interspecies variation in gene expression, growth, and development. Results from these research objectives show that genome-wide analysis of high throughput gene expression and small RNAs provides new insights into molecular and evolutionary mechanisms for gene expression diversity and phenotypic variation between closely related species and in the new allopolyploids. / text

Genome sequencing technologies are providing huge quantities of data for phylogenetic inference. However, most phylogenomic studies exclude gene families, because many have a complicated history of gene duplication/loss and structural change by domain shuffling, especially in deep phylogenies. Gene tree parsimony (GTP) methods, which seek the species tree that minimizes the cost of gene duplication, have been successfully applied to gene families with frequent duplication history. Their utility and performance in the context of gene families with complex histories of gene duplication and domain reshuffling remains unclear. In this study, we analyzed 4389 gene families from six angiosperm genomes encompassing a wide range of duplication rates, and a broad diversity of domain architecture. Overall species tree inference accuracy increased monotonically with the inclusion of more gene trees, and high accuracy was achieved with 50-100 gene trees. The rate of gene duplication strongly influences species tree inference accuracy, with the highest accuracy at either very low or very high rates of duplication and lowest accuracy centered around one duplication per branch in the unrooted species tree. This is the opposite of the relationship between substitution rates on tree construction accuracy, in which intermediate rates have highest accuracy. Accuracy is generally higher in gene families with high domain architecture diversity but has high variance in families with relatively low domain architecture diversity. The latter is probably due to the high variation of gene duplication number for those gene families. We close with some discussion of potential impacts of domain evolution on phylogenomic reconstruction protocols in general, including its effect on alignment.

Centrosomes are organelles that promote microtubule growth. Normally, a single centrosome duplicates once each cell cycle to guide assembly of a bipolar mitotic spindle, ensuring that each daughter cell inherits an equal complement of the genome and a single centrosome. Centrosomes are composed of a pair of ‘mother-daughter’ centrioles and, during duplication, each mother centriole assembles one daughter at a single site. However, mother centrioles can inappropriately assemble multiple daughters, thereby generating centriole amplification (or overduplication), resulting in multipolar spindle assembly and, consequently, chromosome missegration - a driving force for chromosomal instability/aneuploidy which induces birth defects, miscarriage, and tumorigenesis. We have elucidated how the cell cycle control program regulates the centriole duplication machinery to limit centriole duplication to one event per cell cycle via the cell cycle-dependent regulation of Ana2/STIL and PLK4 degradation. In the case of the centrosome licensing factor Plk4, we found that autophosphorylation promotes its own destruction during interphase, which is then counteracted by the Protein Phosphatase 2A (PP2A) in complex with its Twins (tws) regulatory subunit during mitosis. This promotes stabilization of Plk4 and thus allows for the licensing of the mother centriole, making it competent to duplicate during the proceeding S-phase. While PP2Atws plays a positive role in regulating Plk4 to promote centriole duplication, we found that PP2A complexed with the Well-rounded (wrd) and Widerborst (wdb) regulatory subunits negatively regulates Ana2 by promoting its degradation to limit centriole duplication. PP2Awrd/wdb dephosphorylates numerous serine/threonine residues residing in Ana2, including several CDK phosphorylation consensus motifs. We found that CDK1/cycA and CDK2/cycE phosphorylate these residues to promote Ana2 stabilization from S-phase, the start of centriole duplication, to M-phase, the start of centriole duplication licensing. Interestingly, we found that the tumorigenic SV40 virus protein Small Tumor Antigen (ST) amplifies centrioles by targeting the PP2A complex to stabilize Plk4 as well as Ana2, underscoring the oncogenic importance of these newly discovered centriole duplication pathways. Finally, we shed insight into the mechanism for centriole amplification upon Ana2 stabilization by showing that Ana2 associates with Plk4 to promote Plk4 kinase activity as well as Plk4 stabilization.

The buildup of salt in soils is a major abiotic stress that affects agricultural productivity, limiting the growth and yield of most crop species which cannot tolerate even modest levels of salinity (glycophytes). Genetic variability for salt tolerance exists as some plants (halophytes) have adapted to environments with high levels of salt. Understanding how salt tolerance has been acquired in halophytic species will be an important part of strategies to improve the ability of crops to grow in saline soils. The CALCINEURIN B-LIKE10 (AtCBL10) calcium sensor was identified as a component of salt signaling in the glycophyte Arabidopsis thaliana (A. thaliana) based on hypersensitivity of the Atcbl10 mutant to salt. When A. thaliana is grown in the presence of salt, AtCBL10 interacts with the AtSOS2 protein kinase to activate the AtSOS1 sodium/proton exchanger, resulting in the removal of sodium ions from the cytosol. Eutrema salsugineum (E. salsugineum), a halophytic relative of A. thaliana, has two CBL10 genes (EsCBL10a and EsCBL10b). In this research, the duplication of CBL10 in E. salsugineum was characterized and the functions of EsCBL10a and EsCBL10b in salt tolerance were determined. My analyses indicate that the coding sequences of EsCBL10a and EsCBL10b are highly conserved, as they share 85% nucleotide identity. An analysis of transcript structure indicates transcripts from EsCBL10a and EsCBL10b loci are alternatively spliced, but in distinct ways. My results suggest that EsCBL10a and AtCBL10 likely share the ancestral genomic position, while EsCBL10b might have moved to a different genomic region, and that the duplication took place prior to the divergence of expanded Lineage II species. The expression patterns of EsCBL10a and EsCBL10b are different; EsCBL10b transcript is high in shoots and low in roots while EsCBL10a transcript is detectable in both tissues. Preliminary analysis of E. salsugineum lines with reduced expression of EsCBL10a and EsCBL10b suggest that both genes might play a role during growth in the presence of salt, but that these roles are distinct.

alpha-actinin is a ubiquitous protein found in most eukaryotic organisms. The ability to form dimers allows alpha-actinin to cross-link actin in different structures. In muscle cells alpha-actinin is found at the Z-disk of sarcomeres. In non-muscle cells alpha-actinin is found in zonula adherens or focal adhesion sites where it can bind actin to the plasma membrane. alpha-actinin is the shortest member of the spectrin superfamily of proteins which also includes spectrin, dystrophin and utrophin. Several hypotheses suggest that alpha-actinin is the ancestor of this superfamily. The structure of alpha-actinin in higher organisms has been well characterized consisting of three main domains: an N-terminal actin-binding domain with two calponin homology domains, a central rod domain with four spectrin repeats and a C-terminal calcium-binding domain. Data mining of genomes from diverse organisms has made possible the discovery of new and atypical alpha-actinin isoforms that have not been characterized yet. Invertebrates contain a single alpha-actinin isoform, whereas most of the vertebrates contain four. These four isoforms can be broadly classified in two groups, muscle isoforms and non-muscle isoforms. Muscle isoforms bind actin in a calcium independent manner whereas non-muscle isoforms bind actin in a calcium-dependent manner. Some of the protozoa and fungi isoforms are atypical in that they contain fewer spectrin repeats in the rod domain. We have purified and characterized two ancestral alpha-actinins from the parasite Entamoeba histolytica. Our results show that despite the shorter rod domain they conserve the most important functions of modern alpha-actinin such as actin-bundling formation and calcium-binding regulation. Therefore it is suggested that they are genuine alpha-actinins. The phylogenetic tree of alpha-actinin shows that the four different alpha-actinin isoforms appeared after the vertebrate-invertebrate split as a result of two rounds of genome duplication. The atypical alpha-actinin isoforms are placed as the most divergent isoforms suggesting that they are ancestral isoforms. We also propose that the most ancestral alpha-actinin contained a single repeat in its rod domain. After a first intragene duplication alpha-actinin with two spectrin repeats were created and a second intragene duplication gave rise to modern alpha-actinins with four spectrin repeats.

Duplicated genes are present in the teleost fish lineage owing to a whole-genome duplication (WGD) event that occured ~ 230-400 million years ago. In the duplication-degeneration-complementation (DDC) model, partitioning of ancestral functions (subfunctionalization) and acquisition of novel functions (neofunctionalization) have been proposed as principal processes for the retention of duplicated genes in the genome. The DDC model was tested by analyzing the differential tissue-specific distribution of transcripts for the duplicated fatty acid-binding protein 10 (fabp10) genes in embryos, larvae and adult zebrafish (Danio rerio). The distribution of zebrafish fabp10a and fabp10b transcripts show a strikingly different tissue-specific pattern leading us to suggest that the zebrafish fabp10 duplicates had been retained in the genome owing to neofunctionalization. In another experiment to test the DDC model, transcriptional regulation of duplicated fabp genes was analyzed in zebrafish fed clofibrate, a peroxisome proliferator-activated receptor (PPAR) agonist. Clofibrate increased the steady-state level of both the duplicated copies of fabp1a/fabp1b.1, and fabp7a/fabp7b mRNA and heteronuclear RNA (hnRNA), but in different tissues of zebrafish. The steady-state level of fabp10a and fabp11a mRNA and hnRNA was elevated in liver of zebrafish, but not for fabp10b and fabp11b. We also investigated the effect of dietary fatty acids (FAs) and clofibrate on the transcriptional regulation of single copy fabp genes, fabp2, fabp3 and fabp6 in zebrafish. The steady-state level of fabp2 transcripts increased in intestine, while fabp3 mRNA increased in liver of zebrafish fed diets differing in FA content. In zebrafish fed clofibrate, fabp3 mRNA in intestine, and fabp6 mRNA in intestine and heart, was elevated. Whether the regulation of fabp gene transcription by clofibrate is controlled either directly or indirectly, the regulatory elements in the zebrafish fabp genes have diverged markedly since the WGD event, thereby supporting the DDC model.

Despite being found in all presently sequenced genomes, the evolution of tandemly repeated sequences has only just begun to be understood. We can represent the duplication history of tandemly repeated sequences with duplication trees. Most phylogenetic techniques need to be modified to be used on duplication trees. Due to gene loss, it is not always possible to reconstruct the duplication history of a tandemly repeated sequence. This thesis addresses this problem by providing a polynomial-time locally optimal algorithm to reconstruct the duplication history of a tandemly repeated sequence in the presence of gene loss. Supertree methods cannot be directly applied to duplication trees. A polynomial-time algorithm that takes a forest of ordered phylogenies and looks for a super duplication tree is presented. If such a super duplication tree is found then the algorithm constructs the super duplication tree. However, the algorithm does not always find a super duplication tree when one exists. The SPR topological rearrangement in its current form cannot be used on duplication trees. The necessary modifications are made to an agreement forest so that the SPR operation can be used on duplication trees. This operation is called the duplication rooted subtree prune and regraft operation (DrSPR). The size of the DrSPR neighbourhood is calculated for simple duplication trees and the tree shapes that maximize and minimize this are given.

No description available.

La duplication de code source a de nombreuses origines : copie et adaptation inter-projets ou clonage au sein d'un même projet. Rechercher des correspondances de code copié permet de le factoriser dans un projet ou de mettre en évidence des situations de plagiat. Nous étudions des méthodes statiques de recherche de similarité sur du code ayant potentiellement subi des opérations d'édition telle que l'insertion, la suppression, la transposition ainsi que la factorisation et le développement de fonctions. Des techniques d'identification de similarité génomique sont examinées et adaptées au contexte de la recherche de clones de code source sous forme lexemisée. Après une discussion sur des procédés d'alignement de lexèmes et de recherche par empreintes de n-grams, est présentée une méthode de factorisation fusionnant les graphes d'appels de fonctions de projets au sein d'un graphe unique avec introduction de fonctions synthétiques exprimant les correspondances imbriquées. Elle utilise des structures d'indexation de suffixes pour la détermination de facteurs répétés. Une autre voie d'exploration permettant de manipuler de grandes bases indexées de code par arbre de syntaxe est abordée avec la recherche de sous-arbres similaires par leur hachage et leur indexation selon des profils d'abstraction variables. Des clones exacts de sous-arbres de forte proximité dans leurs arbres d'extraction peuvent alors être consolidés afin d'obtenir des correspondances approchées et étendues. En amont et en aval de la recherche de correspondances, des métriques de similarité sont définies afin de préselectionner les zones d'examen, affiner la recherche ou mieux représenter les résultats

[Truncated abstract] For the convenience of the reader, detailed figures and tables have been enlarged and compiled in Appendix 2, at the end of this thesis. This thesis is presented as an approach to identify, annotate and detect genomic duplication and polymorphism within large genomic regions. To demonstrate this, I have used as a model, the genomic region known as the Regulators of Complement Activation (RCA). The RCA complex is located on the long arm of chromosome 1 at position 1q32 and is a reservoir of complement regulatory proteins. The genes of the RCA share many similarities implying that all have arisen through multiple complex duplication events. My analysis of this region in the following chapters demonstrates the complexity of this duplication and identifies the many functional units within the RCA. It was my aim at the beginning of these studies to demonstrate an approach that could define the Ancestral Haplotypes (AHs) of the RCA gene cluster. To do this, extensive genomic analysis was required and the ever-increasing availability of genomic sequence has made this thesis possible. Each of the chapters serves to address the following aims set out at the beginning of this thesis: 1. Further characterise the relationship between the genes (Complement Control proteins-CCPs) and domains of the Regulators of Complement Activation (RCA). 2. Identify and examine the duplicated elements within the RCA. - 6 - 3. Examine the effects of retroviruses and other insertions and deletions (indels) in generating the divergence of duplicated genes. 4. Investigate the applicability of the Genomic Matching Technique (GMT) to define AH within the region. 5. Examine association of AHs with CCP implicated diseases. 6. Determine the GMT applicability in non-human species