Forest trees are an important pool of biodiversity at the gene, individual and an ecosystem level. This variation is a result of complex environmental interactions, as well as neutral and selective forces acting on populations. Patterns of standing genetic variation are the result of adaption to past and contemporary climate change, but also historical demographic events, and disentangling the role of these forces is a central problem in population genomics. The overall goal of this study is to characterize the relative effects of demography and selection in the genome of Populus trichocarpa, a riparian deciduous tree species of North America. Specifically, I used a variety of methods to summarize patterns of genetic diversity and population structure in P. trichocarpa, and to reconstruct its demographic history. I subsequently incorporated these demographic insights to guide the application of several methods to identify genome-wide targets of natural selection within and among rangewide populations adapted to heterogeneous selection regimes. Results of this study provide insights into the history of divergence and differentiation in P. trichocarpa populations and help us identify the functional genetic variants contributing to phenotypic divergence and fitness of the individuals in it. / Ph. D.

A major challenge to evolutionary developmental biology is to understand the how modifications to gene regulatory networks can lead to biological diversity. Heliconius butterfly wing patterns provide an excellent example of this diversity. In particular, the species H. melpomene and H. erato display wide variation in wing pattern across their ranges in Central and South America, but wherever they co-occur, they have converged on remarkably similar wing patterns due to Müllerian mimicry. Linkage analysis of wing pattern genes has shown that in both species, there are three genomic loci that are responsible for most of the pattern variation, and that these loci are homologous. One locus, containing the transcription factor optix, is responsible for red pattern elements. A set of non-coding sequences linked to some of the red pattern elements have been identified. Another locus, containing the gene WntA, has been linked to the shape of the forewing band elements and is responsible for variation in wing pattern development in several species of lepidoptera. A third locus, responsible for yellow pattern elements, contains multiple candidate genes that may affect wing pattern development, including the gene cortex, which is also linked to the industrial melanism phenotype in the moth Biston betularia, as well as the genes domeless and washout, linked to the Bigeye mutant in Bicyclus anynana. I first investigated modifications to regulatory sequence near the transcription factor optix, detecting a module associated with the band pattern element. I also found that for some pattern regulatory modules at optix, the same sequence has independently evolved the same function in H. melpomene and H. erato, in association with non-coding sequences conserved throughout the Lepidoptera. I then investigated gene expression differences in two morphs from either side of a hybrid zone that vary only in the presence or absence of a yellow pattern element, in order to determine a role for candidate genes at the yellow pattern locus. In H. melpomene the gene cortex was upregulated in the larval wing discs of the black morph, whereas in H. erato it was upregulated in the larval wing discs of the yellow morph. In pupal wings, washout was differentially expressed, again in the opposite pattern in the two species, suggesting the same locus is responsible for convergent pattern modification, but by a different mechanism. Finally, I investigated the spatial transcriptomic landscape across the wings of three different heliconiine butterflies. I identified candidate factors for regulating the expression of wing patterning genes, including genes with a conserved expression profile in all three species, and others, including genes in the Wnt pathway, with markedly different profiles in each of the three species. Each of these studies contributes to our understanding of how gene regulatory networks can be modified to create diversity: first, at the level of cis-regulation, second at the level of gene interaction and expression, and lastly at the level of developmental bias and constraint.

No description available.

How do bacteria grow and replicate rapidly? How do viruses and phages adapt to their host environments? Bacteria require efficient translation to grow and replicate rapidly, and translation is often rate-limited by initiation. A feature that is conserved across bacterial lineages is the Shine-Dalgarno (SD) sequence at the mRNA 5’ UTR, which pairs with the anti-SD sequence located at the 3’ end of mature 16S rRNA. Nonetheless, much about this interaction remains unclear. Chapter 2 reveals evolutionary differences between Cyanobacteria and chloroplast translation initiation using a new model (DtoStart) that better define optimal SD sequence and an RNA-Seq-based approach that reliably characterize the 3’ end of mature 16S rRNAs. Efficacy of translation elongation depends much on tRNA-mediated codon adaptation. In Escherichia coli, selection favours major codons because they are rapidly decoded by abundantly available cognate tRNAs. Nonetheless, the degree codon bias correlates with tRNA availability is unclear in many bacterial species because tRNA abundance is often inadequately approximated by gene copy numbers. To better understand tRNA-mediated codon bias, Chapter 3 describes an RNA-Seq-based approach to robustly quantify tRNA abundance. Finally, Chapter 4 evaluates the degree optimal translation initiation and elongation signals affect ribosome dynamics. The emergence of COVID-19 pandemic poses a serious global health emergency. To establish infection during cell entry, the coronavirus Spike protein binds to the host ACE2 receptor, and a high binding potential between these two players is key to infectivity. While SARS-CoV-2 transmits efficiently in humans, it is less clear which other mammals are at risk of being infected. Chapter 5 investigates the host range of SARS-CoV-2 through comparative sequence analyses at the ACE2 receptors and the Spike proteins. As obligate parasites, coronaviruses regularly infect host tissues that express antiviral proteins (AVPs) in abundance and must evade or adapt to the host cellular environments post-entry. Two AVPs that shape viral genomes are ZAP that binds to CpG dinucleotides to facilitate viral transcript degradation, and APOBEC3 which deaminates C into U leading to dysfunctional transcripts. Chapter 6 shows that coronavirus genomes are CpG deficient to evade ZAP and are subjected to constant C to U deamination by APOBEC3. This thesis examines two key concepts of microbial genome evolution: 1) coevolution between gene features and the translation machinery in bacteria, and 2) adaptation of viruses to the hosts they infect. Chapters 2, 3, and 4 are aimed at improving our understanding in bacterial gene expression in the applications of transgenic biosynthesis and phage therapy. Chapters 5 and 6 are aimed at improving our understanding in the origin and evolution of SARS-CoV-2 and our ability to control the spread of infection.

Gene duplications can facilitate genetic innovation, reduce pleiotropy and catalyze reproductive incompatibilities and speciation. Therefore, the molecular and transcriptional fate of duplicate genes plays an important role in the evolutionary trajectory of entire genomes and transcriptomes. Using the polyploid African clawed frog Xenopus, I have investigated mechanisms that promote the retained expression of duplicate genes (paralogs) after whole genome duplication. The studies herein estimated molecular evolution and characterized expression divergence of thousands of duplicate genes and a singleton ortholog from a diploid outgroup. In this thesis, I have discussed the multiple mechanisms for the retention of duplicate genes in a polyploid genome and examined the potential effects that gene characteristics before duplication have on the odds of duplicate gene persistence. I have also explored the use of microarrays for comparative transcriptomics between duplicate genes, and between diverged genomes. The main objectives of my thesis were to better understand the genetic mechanisms that promote the retained expression of gene duplicates. My research utilized the duplicated genome from the allopolyploid clawed frog Xenopus. Genome duplication in clawed frogs offers a compelling opportunity to study factors that influence the genetic fates of gene duplicates because many paralogs in these frogs are of the same age, permitting one to control for the influence of time when evaluating the impact of duplication. My work has major impacts on several biological fronts including evolutionary genomics and comparative transcriptomics, and also on technical aspects of using microarrays. I have provided among the most comprehensive studies of its kind, in terms of examining molecular and regulatory aspects of thousands of expressed duplicates of the same age, and exploring various alternative hypotheses to explain how these genes are retained. / Thesis / Doctor of Philosophy (PhD)

The ocean plays essential roles in Earth's biochemistry. Most of the nutrient transformations that fuel trophic webs in the ocean are mediated by microorganisms. The extent of phylogenetic and metabolic diversity of key culture and uncultured marine microbial clades started to be revealed due to progress in sequencing technologies, however we still lack a comprehensive understanding of the evolutionary processes that led to the microbial diversity we see in the ocean today. In this dissertation, I apply phylogenomic and comparative genomic methods to explore the evolutionary genomics of bacterial and archaeal clades that are relevant due to their abundance and biogeochemical activities in the ocean. In Chapter 1, I review relevant literature regarding the evolutionary genomics of marine bacteria and archaea, with emphasis on the origins of marine microbial diversity and the evolution of genome architecture. In Chapter 2, I use a comparative framework to get insights into the evolutionary forces driving genome streamlining in the Ca. Marinimicrobia, a clade widely distributed in the ocean. This project shows that differences in the environmental conditions found along the water column led to contrasting mechanisms of evolution and ultimately genome architectures. In Chapter 3, I assess the phylogenetic signal and congruence of marker genes commonly used for phylogenetic studies of bacteria and archaea and propose a pipeline and a set of genes that provide a robust phylogenetic signal for the reconstruction of multi-domain phylogenies. In Chapter 4, I apply a phylogeny-based statistical approach to evaluate how tightly genome size in bacteria and archaea is linked to evolutionary ii history, including marine clades. I present evidence suggesting that phylogenetic history and environmental complexity are strong drivers of genome size in prokaryotes. Lastly, in Chapter 5, I estimate the emergence time of marine bacterial and archaeal clades in the context of the Prokaryotic Tree of Life and demonstrate that the diversification of these groups is linked to the three main oxygenation periods occurring throughout Earth's history. I also identify the metabolic novelties that likely led to the colonization of marine realms. Here I present methodological frameworks in the fields of comparative genomics and phylogenomics to study the evolution of marine microbial diversity and show evidence suggesting that the main evolutionary processes leading to the extant diversity seen in the ocean today are intimately linked to geological and biological innovations occurring throughout Earth's history. / Doctor of Philosophy / The ocean plays essential roles in the functioning of our planet. Many of the nutrient's transformation happening in marine environments are mediated by microorganisms, whose metabolic activities underpin higher trophic levels. The identity of the most prevalent marine microbial groups has been reveled during the last two decades through sequencing technologies. Despite having a great progress in our understanding of the functions that these microorganisms have in the ocean; we still lack information about the evolutionary processes that allowed their diversification and colonization into marine realms. In this work, I developed and applied computational strategies to disentangle the evolutionary genomics of marine microorganisms. One particularity about most these marine groups is that they have very small genomes. To explore the evolutionary forces driving their genome reduction, I analyzed a broad set of genomes of Marinimicrobia, a bacterial group widely distributed in the ocean. This analysis shows that genome reduction in Marinimicrobia is driven by negative selection, an evolutionary force that allows the deletion of non-essential genes, subsequently leading to genome reduction. Moreover, I developed a benchmarked pipeline for the reconstruction of phylogenetic trees to study the evolutionary relationships of microorganisms. This pipeline allowed me to link the diversification of the main marine groups and the geological periods in which they first emerged. I discovered that the colonization of these groups happened during three different periods, which are coincident with the main oxygenation events occurring across Earth's history. Moreover, the diversification of vi marine microbial groups was associated with the acquisition of genes to exploit the newly created niches that followed the oxygenation of the atmosphere and the ocean. Overall, my work shows that the diversification of the marine microbial clades that are essential for the functioning of the ocean is intimately linked to the redox state of the ocean and the atmosphere throughout Earth's history.

Sexual selection has long been a working explanation for the elaboration of appreciable traits in plants and animals, but the idea that it is an equally potent agent of change at the level of individual molecules is relatively recent. Indications that genes associated with reproductive biology evolve especially rapidly planted this notion, but many details about the genomics of sex remain elusive. Numerous studies have characterized rapid sequence and expression divergence of sex-related molecules, but few if any have demonstrated convincingly that these patterns exist as a result of sexual selection. This dissertation describes several genome-scale studies related to reproduction and the sexes in teleost fishes, a group of animals underexploited in regard to this topic. Using commercial microarrays I measured the extent of sexually dimorphic gene expression in the zebrafish, Danio rerio. Sex-biased patterns of gene expression in this species are similar to those described in other animals. A number of genes expressed at high levels in ovaries and testes relative to the body were identified as a product of the study, and these data may be useful for future studies of reproductive genes in Danio fishes. In a second study, the recent advent of high throughput cDNA pyrosequencing was leveraged to characterize the relationships between tissue-, sex-, and species-specific expression patterns of genes and rates of sequence evolution in swordtail fishes (Xiphophorus). I discovered ample evidence for expression biases of all three types, and a generally positive but idiosyncratic relationship between the magnitude of expression bias and rates of protein-coding sequence evolution. Pyrosequencing of cDNA was also used to explore the possibility that postcopulatory sexual selection drives the rapid evolution of male pregnancy genes, a novel class of reproductive molecules unique to syngnathid fishes (seahorses and pipefishes). Genes differentially expressed in the male brooding tissues as a function of pregnancy status evolve more rapidly at the amino acid level than genes exhibiting static expression. Brooding tissue genes expressed during male pregnancy have evolved especially rapidly in polyandrous lineages, a finding that supports the hypothesized relationship between postcopulatory sexual selection and the adaptive evolution of reproductive molecules.

This thesis focuses on the genomic study of symbionts of two different groups of hymenopterans: bees and ants. Both groups of insects have major ecological impact, and investigating their microbiomes increases our understanding of their health, diversity and evolution. The study of the bee gut microbiome, including members of Lactobacillus and Bifidobacterium, revealed genomic processes related to the adaptation to the gut environment, such as the expansion of genes for carbohydrate metabolism and the acquisition of genes for interaction with the host. A broader genomic study of these genera demonstrated that some lineages evolve under strong and opposite substitution biases, leading to extreme GC content values. A comparison of codon usage patterns in these groups revealed ongoing shifts of optimal codons. In a separate study we analysed the genomes of several strains of Lactobacillus kunkeei, which inhabits the honey stomach of bees but is not found in their gut. We observed signatures of genome reduction and suggested candidate genes for host-interaction processes. We discovered a novel type of genome architecture where genes for metabolic functions are located in one half of the genome, whereas genes for information processes are located in the other half. This genome organization was also found in other Lactobacillus species, indicating that it was an ancestral feature that has since been retained. We suggest mechanisms and selective forces that may cause the observed organization, and describe processes leading to its loss in several lineages independently. We also studied the genome of a species of Rhizobiales bacteria found in ants. We discuss its metabolic capabilities and suggest scenarios for how it may affect the ants’ lifestyle. This genome contained a region with homology to the Bartonella gene transfer agent (GTA), which is a domesticated bacteriophage used to transfer bacterial DNA between cells. We propose that its unique behaviour as a specialist GTA, preferentially transferring host-interaction factors, originated from a generalist GTA that transferred random segments of chromosomal DNA. These bioinformatic analyses of previously uncharacterized bacterial lineages have increased our understanding of their physiology and evolution and provided answers to old and new questions in fundamental microbiology.

Abstract A central goal in evolutionary biology is to identify the ecological and genetic mechanisms that give rise to adaptation and speciation. Importantly, a large body of theoretical work has modelled the adaptive evolution under selection, migration and drift. Yet to test these predictions on an empirical level has proven a challenging task. The aim of my thesis is to explore outstanding questions in local adaptation and reproductive isolation using natural populations of Arabidopsis lyrata: How does differential selection lead to adaptive divergence in the face of gene flow and drift? What traits underlie both short- and large-scale adaptive differentiation? And what reproductive barriers are involved in incipient speciation? By combining whole-genome based demography simulations with a multi-year reciprocal transplant experiment, I confirmed that alpine and lowland populations of A. lyrata are adapted to their local environments despite high gene flow and strong drift. Patterns of trait differentiation, supported by analysis of phenotypic selection, further suggested that flowering traits have contributed to the adaptive divergence. Selection patterns at the sequence level confirmed that the genetic architecture underlying the local adaptation conforms to theory: populations under higher levels of gene flow had fewer adaptive loci that were also found in areas of reduced recombination. Although most selection outliers were population specific, indicating conditional neutrality, a small proportion showed potential for genetic trade-offs (antagonistic pleiotropy). The analysis also revealed important traits and biological processes linked to alpine and lowland adaptation. The role of seed germination in large-scale adaptation and reproductive isolation was also studied. Populations representing the European and North American subspecies exhibited germination patterns consistent with adaptive differentiation. Comparisons against first- and second-generation hybrids then indicated that genetic incompatibilities impede germination of the hybrid seeds. Furthermore, genetic mapping helped to clarify the genetic basis of these phenotypic traits. Taken together, the three studies in this thesis highlight the value of combining traditional organismal methods with next-generation genomics, by providing novel insights into processes underlying adaptation and speciation. / Tiivistelmä Evoluutiobiologian keskeinen tehtävä on sopeutumiseen ja lajiutumiseen johtavien prosessien selvittäminen. Vaikka evoluutiovoimien – luonnonvalinnan, geenivirran ja geneettisen satunnaisajautumisen – vaikutusta adaptiivisen muuntelun määrään on mallinnettu laajalti, teoreettisten ennusteiden tarkastelu empiirisellä tasolla on usein osoittautunut haastavaksi. Tässä väitöskirjatyössä pyrin vastaamaan eräisiin paikallissopeutumisen ja lajiutumisen kannalta tärkeisiin kysymyksiin, hyödyntäen kasvilaji idänpitkäpalkoa (Arabidopsis lyrata) malliorganismina: Kuinka luonnonvalinta johtaa paikallissopeutumiseen tilanteessa, jossa geenivirta samankaltaistaa eriytyvien populaatioiden perimää? Mitkä ominaisuudet vaikuttavat adaptiiviseen erilaistumiseen eri etäisyyksillä olevien populaatioiden välillä? Sekä millaiset lisääntymisesteet johtavat alkavaan lajiutumiseen? Yhdistämällä genomisekvensointiin perustuvan demografia-analyysin ja monivuotisen siirtoistutuskokeen, selvitin kuinka idänpitkäpalkopopulaatiot ovat sopeutuneet elinympäristöihinsä runsaasta geenivirrasta ja voimakkaasta satunnaisajautumisesta huolimatta. Fenotyyppisen muuntelun ja kelpoisuuden yhteys vahvisti myös, että kukkimisominaisuudet ovat vaikuttaneet populaatioidenväliseen adaptiiviseen erilaistumiseen. Valinnan merkkien etsiminen sekvenssitasolla osoitti, että havaintoni paikallissopeutumisen geneettisestä arkkitehtuurista tukevat teoreettisia ennusteita: populaatioista, joihin kohdistuu voimakasta geenivirtaa, löytyi vähemmän adaptiivisia lokuksia ja ne olivat keskittyneet matalamman rekombinaation alueille. Suurin osa adaptiivisista lokuksista löytyi ainoastaan yhdestä populaatiosta, ollen näin todennäköisesti valinnan alla ainoastaan tietyssä elinympäristössä. Pieni osuus lokuksista vastasi kuitenkin harvoin havaittua tilannetta, jossa hajottava valinta on johtanut eri alleelien runsastumiseen populaatioissa, joita yhdistää geenivirta. Tutkin myös, miten itämisajan muuntelu vaikuttaa sopeutumiseen pitkällä aikavälillä. Eurooppalaista ja pohjoisamerikkalaista alalajia edustavat populaatiot itivät tavalla, joka viittaa adaptiiviseen erilaistumiseen. Ensimmäisen ja toisen hybridisukupolven siementen vertailu paljasti lisäksi, että geneettiset yhteensopimattomuudet haittaavat hybridien itämistä, toimien näin lisääntymisesteenä. Geenikartoitus auttoi myös selventämään näiden itämisominaisuuksien geneettistä taustaa. Tämän väitöskirjan kolme osatyötä korostavat miten perinteisten yhteiskenttäkokeiden ja uuden sukupolven genomimenetelmien yhdistelmä voi tuottaa arvokasta lisätietoa sopeutumisen ja lajiutumisen mekanismeista.

Indiana University-Purdue University Indianapolis (IUPUI) / RNA-binding proteins (RBPs) are vital post-transcriptional regulatory molecules in transcriptome of mammalian species. It necessitates studying their expression dynamics to extract how post-transcriptional networks work in various mammalian tissues. RNA binding proteins (RBPs) play important roles in controlling the post-transcriptional fate of RNA molecules, yet their evolutionary dynamics remains largely unknown. As expression profiles of genes encoding for RBPs can yield insights about their evolutionary trajectories on the post-transcriptional regulatory networks across species, we performed a comparative analyses of RBP expression profiles across 8 tissues (brain, cerebellum, heart, lung, liver, lung, skeletal muscle, testis) in 11 mammals (human, chimpanzee, gorilla, orangutan, macaque, rat, mouse, platypus, opossum, cow) and chicken & frog (evolutionary outgroups). Noticeably, orthologous gene expression profiles suggest a significantly higher expression level for RBPs than their non-RBP gene counterparts, which include other protein-coding and non-coding genes, across all the mammalian tissues studied here. This trend is significant irrespective of the tissue and species being compared, though RBP gene expression distribution patterns were found to be generally diverse in nature. Our analysis also shows that RBPs are expressed at a significantly lower level in human and mouse tissues compared to their expression levels in equivalent tissues in other mammals: chimpanzee, orangutan, rat, etc., which are all likely exposed to diverse natural habitats and ecological settings compared to more stable ecological environment humans and mice might have been exposed, thus reducing the need for complex and extensive post-transcriptional control. Further analysis of the similarity of orthologous RBP expression profiles between all pairs of tissue-mammal combinations clearly showed the grouping of RBP expression profiles across tissues in a given mammal, in contrast to the clustering of expression profiles for non-RBPs, which frequently grouped equivalent tissues across diverse mammalian species together, suggesting a significant evolution of RBPs expression after speciation events. Calculation of species specificity indices (SSIs) for RBPs across various tissues, to identify those that exhibited restricted expression to few mammals, revealed that about 30% of the RBPs are species-specific in at least one tissue studied here, with lung, liver, kidney & testis exhibiting a significantly higher proportion of species specifically expressed RBPs. We conducted a differential expression analysis of RBPs in human, mouse and chicken tissues to study the evolution of expression levels in recently evolved species (i.e., humans and mice) than evolutionarily-distant species (i.e., chickens). We identified more than 50% of the orthologous RBPs to be differentially expressed in at least one tissue, compared between human and mouse, but not so between human and an outgroup chicken, in which RBP expression levels are relatively conserved. Among the studied tissues (brain, liver and kidney) showed a higher fraction of differentially expressed RBPs, which may suggest hyper- regulatory activities by RBPs in these tissues with species evolution. Overall, this study forms a foundation for understanding the evolution of expression levels of RBPs in mammals, facilitating a snapshot of the wiring patterns of post-transcriptional regulatory networks in mammalian genomes. In our second study, we focused on elucidating novel features of post-transcriptional regulatory molecules called as circRNA from LongPolyA RNA-sequence data. The debate over presence of nonlinear exon splicing such as exon-shuffling or formation of circularized forms has finally come to an end as numerous repertoires have shown of their occurrence and presence through transcriptomic analyses. It is evident from previous studies that along with consensus-site splicing non-consensus site splicing is robustly occurring in the cell. Also, in spite of applying different high-throughput approaches (both computational and experimental) to determine their abundance, the signal is consistent and strongly conforming the plausible circularization mechanisms. Earlier studies hypothesized and hence focused on the ribo-minus non-polyA RNA-sequence data to identify circular RNA structures in cell and compared their abundance levels with their linear counterparts. Thus far, the studies show their conserved nature across tissues and species also that they are not translated and preferentially are without poly (A) tail, with one to five exons long. Much of this initial work has been performed using non-polyA sequencing thus probably underestimates the abundance of circular RNAs originating from long poly (A) RNA isoforms. Our hypothesis is if the circular RNA events are not the artifact of random events, but has a structured and defined mechanism for their formation, then there would not be biases on preferential selection / leaving of polyA tails, while forming the circularized isoforms. We have applied an existing computational pipeline from earlier studies by Memczack et. al., on ENCODE cell-lines long poly (A) RNA-sequence data. With the same pipeline, we achieve a significant number of circular RNA isoforms in the data, some of which are overlapping with known circular RNA isoforms from the literature. We identified an approach and worked upon to identify the precise structure of circular RNA, which is not plausible from the existing computational approaches. We aim to study their expression profiles in normal and cancer cell-lines, and see if there exists any pattern and functional significance based on their abundance levels in the cell.