John E. Pool, Ph.D. - US grants

[unreadable] DESCRIPTION (provided by applicant): The proposed research concerns the inference of historical patterns of migration. Traditional population genetic models of migration assume that populations have been exchanging migrants at a constant rate over long periods of time. For many species, however, this assumption may not be appropriate. Therefore, the development of a computational method to test for recent changes in migration rate and to estimate the relevant demographic parameters is proposed. While most methods of demographic inference assume that all of the genetic markers being studied are independent (unlinked), this approach will take advantage of the patterns of linkage along a recombining chromosome. By considering this linkage information (specifically, the lengths of DNA segments that inferred to have migrant origin), one can go beyond estimating how much migration has occurred between two populations, and say something about when, historically, this migration occurred. During the first phase of this project, the effect of various population histories on the length distribution of migrant DNA segments will be investigated, making use of an existing simulation program (ms) and inference method (structure 2.0). Next, the new inference method described above will be developed, using Markov chain Monte Carlo methodology in a maximum likelihood or Bayesian framework. Finally, this method will be applied to existing human polymorphism data sets (both SNP and microsatellite) in order to test the null hypothesis that migration among human populations has been constant since their divergence. This analysis will permit the estimation of demographic parameters for admixed human populations, and will therefore aid in the selection of populations for admixture mapping studies of disease association. [unreadable] [unreadable] Relevance to public health: The goal of the proposed research is to test for historical changes in the rate of migration between populations, and to estimate quantities such as the time since a migration rate change and the magnitude of such a change. The computational method developed will have a variety of applications, including the estimation of demographic parameters in human populations with a history of recent admixture (ancestry from multiple sources), such as African-American, Hispanic, Central Asian and Northern African populations. That information will be relevant in assessing the utility of such populations for admixture mapping studies, which aim to identify genetic variants associated with complex diseases that occur at different frequencies in different populations. [unreadable] [unreadable] [unreadable]

This project will investigate the genetic basis of dark body pigmentation (melanism) in high altitude African populations of the fruit fly Drosophila melanogaster. Pigmentation genes identified in Cameroonian and Ethiopian populations will be compared to one already identified in a Ugandan sample to test whether melanism has evolved via distinct genetic pathways. High-throughput DNA sequence data will be collected from fly "introgression lines" (where dark pigmentation has been crossed into lightly pigmented fly stocks) to identify genomic regions containing pigmentation loci. Similar data from wild-collected fly stocks will allow specific genes within these regions to be tested for associations with pigmentation and evidence for natural selection.

This research will significantly improve scientific understanding of how natural selection operates at the genetic level. It will also lay the groundwork for studies that utilize the genetic resources of this model species to pursue the precise mutations underlying melanic evolution. Broader impacts supported by this grant will include (1) the development of instructional materials for the college evolution curriculum, (2) the involvement of a diverse group of undergraduates in research, and (3) the creation of a Virtual Stock Center for Drosophila Population Samples to allow researchers to more efficiently manage living fly stocks.

DESCRIPTION (provided by applicant): This research investigates the genetic basis, cellular mechanisms, and developmental consequences of body and wing size evolution in a newly discovered high altitude population of Drosophila melanogaster which represents the largest known flies of this species. This trait offers the unique opportunity to relate a biologically important phenotypic difference to its underlying cellular mechanisms and to specific genes and mutations responsible for this change. The Drosophila system offers a range of advantages to study adaptive evolution at the genetic level, from population genomic data to transgenic tools. Results will increase our knowledge of the polygenicity of adaptation and the properties of causative variants (e.g. coding vs. regulatory; new mutations vs. standing genetic variation). This research integrates biological subdisciplines to reveal cell-level mechanisms (e.g. cell proliferation and somatic ploidy) responsible for phenotypic evolution. It also takes a rare look a the potential influence of adaptive evolution on developmental stability. This work will reveal how evolution has altered the function of genes involved in insulin signaling and the cell cycle without harmful consequences to the organism - findings that may ultimately prove relevant for research on cancer, diabetes, and other medical conditions. 1. Conduct a genome-wide search for genes that play a role in body size evolution. Confirm their effects and test causative mutations using transgenesis. A novel QTL mapping method will localize causative genes to the ~100kb scale. A new population genetic statistic will reveal genes within QTL intervals that show evidence of population-specific selection in the highland sample (initial outliers include insulin signaling genes). Genes identified will be functionally tested for influence on body and wing size using a newly developed transgenic approach; specific mutations will be tested in the same way. 2. Reveal cellular mechanisms underlying body size evolution. Research will test whether changes in both cell size and number may give rise to the strikingly large wings of Ethiopian D. melanogaster. Research will also confirm whether the observed enlargement of larval muscles (polynucleate cells which strongly influence adult body size) is due to increases in the number of nuclei or their ploidy. Transgenic constructs will allow the cellular influence of specific adaptive mutations to be assessed. 3. Test whether wing size evolution disrupted developmental canalization. Ethiopian inbred lines show large wings but also very high frequencies of wing vein abnormalities. The hypothesis that phenotypic evolution has destabilized a more developmentally buffered ancestral wing state will be directly tested using a mutagenesis test of genetic perturbility. Artificial selection to recapitulate wing size evolution n the lab will further probe the generality of a link between adaptation and decanalization.

DESCRIPTION (provided by applicant): This research investigates the genetic basis, cellular mechanisms, and developmental consequences of body and wing size evolution in a newly discovered high altitude population of Drosophila melanogaster which represents the largest known flies of this species. This trait offers the unique opportunity to relate a biologically important phenotypic difference to its underlying cellular mechanisms and to specific genes and mutations responsible for this change. The Drosophila system offers a range of advantages to study adaptive evolution at the genetic level, from population genomic data to transgenic tools. Results will increase our knowledge of the polygenicity of adaptation and the properties of causative variants (e.g. coding vs. regulatory; new mutations vs. standing genetic variation). This research integrates biological subdisciplines to reveal cell-level mechanisms (e.g. cell proliferation and somatic ploidy) responsible for phenotypic evolution. It also takes a rare look a the potential influence of adaptive evolution on developmental stability. This work will reveal how evolution has altered the function of genes involved in insulin signaling and the cell cycle without harmful consequences to the organism - findings that may ultimately prove relevant for research on cancer, diabetes, and other medical conditions. 1. Conduct a genome-wide search for genes that play a role in body size evolution. Confirm their effects and test causative mutations using transgenesis. A novel QTL mapping method will localize causative genes to the ~100kb scale. A new population genetic statistic will reveal genes within QTL intervals that show evidence of population-specific selection in the highland sample (initial outliers include insulin signaling genes). Genes identified will be functionally tested for influence on body and wing size using a newly developed transgenic approach; specific mutations will be tested in the same way. 2. Reveal cellular mechanisms underlying body size evolution. Research will test whether changes in both cell size and number may give rise to the strikingly large wings of Ethiopian D. melanogaster. Research will also confirm whether the observed enlargement of larval muscles (polynucleate cells which strongly influence adult body size) is due to increases in the number of nuclei or their ploidy. Transgenic constructs will allow the cellular influence of specific adaptive mutations to be assessed. 3. Test whether wing size evolution disrupted developmental canalization. Ethiopian inbred lines show large wings but also very high frequencies of wing vein abnormalities. The hypothesis that phenotypic evolution has destabilized a more developmentally buffered ancestral wing state will be directly tested using a mutagenesis test of genetic perturbility. Artificial selection to recapitulate wing size evolution n the lab will further probe the generality of a link between adaptation and decanalization.

Project Summary This research aims for a deeper and more nuanced understanding of the genetics of adaptation than has been possible to date. While many trait-associated variants have now been detected by genome-wide association studies, very few of these SNPs have been directly connected to adaptive phenotypes, and the genetic interactions that govern whether their effects are visible to selection. Such knowledge is crucial to composing realistic and testable models for how widespread standing genetic variation within populations is funneled through the sieve of natural selection. The evolution of melanism in high altitude Drosophila melanogaster populations offers several critical advantages for this endeavor. First, the species offers key functional genetic and population genomic resources, along with a well-annotated genome. Second, prior molecular and evolutionary studies have provided strong background knowledge on the trait, including a compelling set of candidate genes. Third, the study of recent adaptive evolution between populations of the same species maximizes the utility of genetic mapping, population genetics, and functional comparisons of alleles. These features will allow the dissection of this model adaptive trait in unparalleled detail, yielding insights regarding: 1. the functional nature of causative variants, 2. genetic variability of the adaptive response, 3. the prevalence and molecular logic of epistasis among adaptive variants, 4. roles of cryptic variation in adaptive change, 5. the importance of standing genetic variation in trait evolution. Results of this research will advance basic understanding of the adaptive evolutionary process. It will also inform on the importance of genetic background in assessing the phenotypic impact of genetic variants, a key step in understanding the genetic architecture of complex traits including human disease. Investigation of these critical topics will be bolstered by a profoundly integrative research plan that leverages the investigators' complementary backgrounds to fuse novel molecular experiments, genomic analysis, and statistical inference. This research will identify genetic variants underlying melanic adaptation in Ethiopian D. melanogaster, fusing genomic mapping and variation analysis with transgenic tests to pinpoint causative changes (Aim 1). It will also advance beyond that goal to reveal the complex interactions that modulate the phenotypic impact of causative variants (Aim 2), examining tissue- and population-specific gene regulation, and non-additive interactions among melanic variants. These investigations will provide a critical case study that will clarify the complexity of adaptive trait evolution at molecular and genetic levels.

Project Summary: Genomic Diversity and the Architectures of Adaptation and Incompatibility This research program addresses fundamental yet unresolved questions at the interface between genetic variation and evolutionary processes. In particular, it focuses on three interconnected research themes: (1) The Genetic Complexity of Adaptive Trait Evolution, (2) The Genetic Basis of Early Stage Reproductive Isolation, and (3) The Determinants of Genomic Diversity and Adaptive Potential. It fuses ambitious but cost-effective Drosophila experiments with novel approaches to the analysis of large genomic data sets. D. melanogaster offers critical advantages for this work. The global expansion of this species enables the study of adaptive trait differences, and partial reproductive isolation, between populations that diverged in the last ~10,000 years. Due to this recent time-scale, adaptive differences may be detected from genetic variation. The experimental efficiency of Drosophila allows the study of large lab populations across many generations. Its compact genome allows economical sequencing. Once relevant genes are found, functional confirmation and study are aided by a well-annotated genome and a wealth of genetic resources and transgenic tools. This research will bolster understanding of The Genetic Complexity of Adaptive Trait Evolution. Initial findings have suggested a portrait of adaptation that often begins with standing genetic variation, includes variants with larger effect sizes, and features variable genetic architectures among individuals. Proposed work will solidify and extend these inferences in multiple respects, including by launching scaled-up mapping studies for a wider range of adaptive traits, and pursuing previous hints of epistasis involving adaptive variants. Proposed work will also open up a promising new system for studying The Genetic Basis of Early Stage Reproductive Isolation. African and European D. melanogaster show evidence of incompatibilities impacting viability and reproduction, but these have received no genetic study. This research will deploy a combination of incompatibility mapping and population genomics to identify specific incompatibility genes for further study, and it will advance understanding of the genetic architecture of the earliest stages of reproductive isolation. Finally, this research will clarify The Determinants of Genomic Diversity and Adaptive Potential. This work will feature experimental and computational studies on the relative roles of neutral and adaptive genetic diversity in future adaptation, and the most relevant ways to quantify each. It will also investigate the relative importance of different types of natural selection in shaping genomic diversity, including by probing the utility of genetic differentiation between populations to separate the signals of positive and negative selection. Collectively, this research is highly consequential for basic evolutionary genetics. It also holds strong medical relevance in terms of the relevance of local adaptation and epistasis for the genetic basis of human disease and infertility, and for understanding the evolutionary dynamics of insect disease vectors.