G protein-coupled receptors (GPCRs) are transmembrane proteins that allow cells to respond to extracellular stimuli. GPCR activation occurs when a ligand binds to the extracellular portion of the receptor. The ligand-bound receptor undergoes a conformational change that allows the intracellular domain of the receptor to engage and activate signaling effectors. The protease-activated receptors (PARs) are a family of GPCRs that are activated by proteases such as thrombin. Unlike traditional GPCRs, which can return to their inactive state after signaling, PARs are irreversibly activated. Therefore, the eventual fate of an activated PAR is degradation. The internalization and endocytic sorting of PARs play key roles in their signal regulation.

Trafficking has been well-characterized for PAR1, the prototypical thrombin receptor. PAR4 was the last of the four thrombin receptor to be discovered, and has in large part been relatively under-studied. Recent studies have shown that PAR4 plays distinct roles that differ from those of PAR1, and as such may represent a potential drug target. Thus, understanding the mechanisms that regulate PAR4 signal regulation is important.

In this dissertation, I characterized the role that trafficking and heterodimerization play in regulating PAR4 signaling. I found that adaptor- protein complex-2 (AP-2) is a key mediator of PAR4 agonist-induced internalization, and that AP-2 binds to intracellular loop 3 (ICL3) of PAR4. Disruption of PAR4 internalization resulted in enhanced and prolonged ERK1/2 activation, suggesting that endocytosis may mediate PAR4 signal attenuation.

I also characterized the role of heterodimerization with the puringeric receptor P2Y12 in modulating PAR4 signaling. Previous studies had demonstrated that PAR4 and P2Y12 coordinate β-arrestin-dependent Akt activation. My work strongly suggests that PAR4 and P2Y12 physically associate basally and co-internalize in response to activation of PAR4. I also demonstrate that β-arrestin is recruited to the co-internalized PAR4-P2Y12 complex. In contrast to the effect on ERK1/2 activation, disruption of PAR4 internalization diminished Akt activation. Taken together, the studies summarized in this dissertation highlight the importance of PAR4 internalization in modulating activity of functionally and spatially distinct signaling pathways.

Monkeypox virus (MPXV), a zoonotic DNA virus, is now considered the most important orthopoxvirus infection in human populations since the eradication of smallpox. The role of reservoir host species is highly important in this zoonotic disease due to the large percentage of human infections attributed to wildlife origins. As a disease that infects multiple hosts, limited knowledge of MPXV prevalence across host species and the relative importance of each host species to disease maintenance makes it difficult to predict human risk and target disease management efforts. In this study, we screened over 1000 museum skin specimens collected from 1899 to 1993 for viral DNA to understand historical MPXV infection in various Funisciurus species across the Congo Basin. MPX viral DNA was found in 93 of 1038 (9.0%) specimens from as early as 1899 and in five different Funisciurus species: Funisciurus anerythrus, Funisciurus carruthersi, Funisciurus congicus, Funisciurus lemniscatus, and Funisciurus pyrropus. We identify two new potential host species, gain insight into the relative prevalence rates of infection within museum skins between various Funisciurus species, and examine the spatial and temporal distribution of monkeypox virus in these possible host species.

Unraveling the unique ways anthropogenic effects impact the genomes of populations, and applying such genomic knowledge to improve conservation policy, is paramount for effective preservation and management of biodiversity in our changing world. The central African country of Cameroon, widely recognized as a global conservation priority, is a prime landscape to study these issues. Here, I explore genetic and epigenetic variation between natural and human-altered habitats in the Cameroonian sunbird Cyanomitra olivacea. I use SNP data and methylation data to contrast patterns of genetic and epigenetic variation, finding that methylation frequency levels shift with changes in environment, and are higher overall in human-altered habitats. Then, I develop a decision-making framework to enable more effective research-to-policy translation, and apply it to a particular case study: preserving adaptive genetic variation in Cameroon. This study is one of the first to explore genetic and epigenetic variation in concert, and reveals the potential ways organisms adapt to anthropogenic change. It is also an attempt to bridge the much-maligned science-policy gap and develop policy tools to bolster conservation.

Mountain yellow-legged frogs (Rana muscosa and Rana sierrae) of California's Sierra Nevada have been extirpated from over 90% of their historic range, initially by introduced trout predators, and more recently by the emergence of the lethal amphibian chytrid fungus (Batrachochytrium dendrobatidis). Formerly widespread and abundant, mountain yellow-legged frogs are grazers and highly connected high level predators, living in low diversity, low productivity high elevation lakes. Here, I examine the extent to which frog extinctions impact Sierra Nevada lake communities. First, in a study that combined surveys of benthic macroinvertebrates in 22 lakes over 5 years with a reanalysis of benthic macroinvertebrate community data from over 150 lakes, I found no strong differences in benthic macroinvertebrate community diversity or composition between lakes with frogs vs. lakes without frogs. I also conducted experiments to evaluate the impact of tadpoles as grazers on benthic algae, and found that tadpoles do reduce algal biomass in artificial habitats, but that the effect can be small and may not outweigh the influence that abiotic variability has on algal biomass in lakes. Lastly, while I documented that large and periodic tadpole aggregations create biogeochemical hotspots of dissolved nitrogen, the phenomena may not be widespread enough to drive differences in diatom diversity in lakes with vs. without tadpoles. Within lakes, I found little indication that diatom community diversity and composition were responding to tadpole generated nitrogen hotspots as a fluctuation-dependent mechanism with potential to enhance diatom coexistence. While none of the ecological effects of mountain yellow-legged frogs and tadpoles that I observed suggests that their extinctions are having large impacts on Sierra Nevada lake communities, they should not be discounted as expendable, because there are many other ways in which these amphibians could be important in their communities.

Species across the globe are faced with unprecedented pressures due to human impacts. In what is now recognized as the Sixth Mass Extinction, habitat loss and climate change have driven many populations to and over the brink of extinction. Drawing from recent advances in genomic sequencing and analyses, I aimed to facilitate the conservation and recovery of two declining species, the Tricolored Blackbird (Agelaius tricolor) and the Western Burrowing Owl (Athene cunicularius hypugaea). I analyzed population structure and genetic diversity in both species, the former with both reduced representation and whole genome sequencing and the latter with low coverage, whole genome data. For the Tricolored Blackbird, I found no indications of population structure in the species, an important finding for conservation as this indicates there is neither a need for individual management units for the species nor a concern about barriers to gene flow. Burrowing Owls are more complicated in having both resident and migratory breeding groups. I found genetic structure was linked to the migratory phenotype, with residents being highly structured and migrants having no structure. Furthermore, I found indications of high inbreeding in resident breeding populations. Using numerous metrics of environmental variation, I found significant genetic differences between residents and migrants. Among those regions of the genome most differentiated and correlating with environmental differences, genes associated with the processing of fats, or lipophagy, were found to be significantly more represented. Revisiting Tricolored Blackbirds with whole genome data, I found indications that perhaps the population is not in complete panmixia as previously indicated by reduced representation data analyses, but that any restrictions to gene flow are indeed low. Further, the higher resolution dataset detected that the recent known decline of the species is apparent in changes in effective population size over time. My work provided essential information for conservation efforts for both species and should be a foundation for continuing genomics research on these and other species of conservation concern as well.

How new species evolve is one of the most fundamental questions in biology. In particular, there has been considerable debate about the relative roles of genetic drift and natural selection in speciation. A salient example is the controversy surrounding the origins of Amazonian biodiversity. The Pleistocene has long considered one of the most important periods of diversification in the Amazon. According to the Pleistocene refugia hypothesis rainforest contracted into small refugia and genetic drift played the dominate role in driving speciation. However, the centers of diversity that would have provided evidence for these supposed refugia were never found. In addition, phylogenies and fossil records show that most lineages are older than the Pleistocene, refuting this simple allopatric model of speciation. Although multiple mechanisms such as geographical barriers, marine incursions, and ecological factors have been hypothesized to explain the Amazon's hyperdiversity there have been relatively few tests of these hypotheses.

Here I examined the importance of ecological factors in driving population divergence in the lesser tree frog (D. minutus) between the Amazon and Cerrado and along the gradient between these two biomes. Substantial temperature and precipitation differences between these biomes raise the possibility that environmental factors drive population differentiation, potentially resulting in speciation.

In chapter one, I examined the importance of environmental variables in driving reproductive divergence in the lesser tree frog. Water availability and temperature emerged as important drivers of body size variation, with body size found to be negatively correlated with vocal traits. For frogs, vocalization is one of the most important traits for mating success. Thus, these results suggest that the contrasting selective pressures on body size between the Amazon and Cerrado influence vocalization, which may result in reproductive divergence and suggests how reproductive isolation may evolve as a result of divergent natural selection pressures in these distinct habitats.

In chapter two, I evaluated the relative roles of genetic drift and natural selection in driving divergence in the lesser tree frog along the Amazon-Cerrado gradient. I showed that environmental variables are more important than geographical distance for driving divergence in reproductive and morphological traits. This suggests that ecological differences along the gradient result in adaptive divergence. Such divergence in phenotypic traits indicates the dominant role of natural selection over neutral processes for speciation events in this region.

Finally, in the third chapter, I investigated the sources of phenotypic variation in response to environmental variation between the Amazon and Cerrado biomes. The goal was to understand the contributions of genetic and environment to D. minutus phenotypic variation in larval development between these two biomes. I conducted a common garden experiment to test whether phenotypic variation in D. minutus has a genetic component, whether tadpoles from the Cerrado develop faster due to a more seasonal habitat, and whether D. minutus shows plastic responses to different thermal conditions. This experimental work showed that both environment and genetics play a role in phenotypic variation. Moreover, tadpoles from Cerrado populations developed faster than those from the Amazon populations. This is indicative of local adaptation to a more seasonal habitat by the Cerrado populations.

Examining patterns of biodiversity in the Tropical Andes provides insight into ecological and evolutionary processes leading to accumulation of lineages in montane regions. In the Tropical Andes, the dramatic changes in topographic and environmental conditions over short distances constitute an ideal system for studying morphological adaptation and geographic discontinuities. These patterns of biodiversity along elevational gradients in the Tropical Andes remains intriguing because there is no comprehensive theory that explains, species richness, high levels of endemism, and the extraordinary variation in traits that are ecologically relevant, in a region that occupies less than 1% of global surface. In particular, I examine phylogenies, eco morphology and population genetics of birds in the Tropical Andes.

In chapter one, I use phylogenetic-based methods to assess slowdown in evolution of body size, changes among lineages in elevational-range overlap and rate-shifts in cladogenesis. Interspecific phylogenetic comparative analysis of the Core Tanager clade supports an overall density-dependent cladogenesis consistent with tenets of adaptive radiation. In chapter two, I use explicit spatial hypotheses of the distribution of the Andean Purplish-Mantled Tanager for present-day and to the past ice-age 18,000-22,000 years ago to test for concordant patterns between population genetic structure using microsatellites loci and mtDNA sequence variation, and potential effects of climate change. Topographical and environmental determinants explain a large proportion of spatial genetic variation in this tanager, but demographic history does not appear to be responding to the expansion of suitable area since last ice-age. In the third chapter, I use coalescent methods for a multilocus data set to examine demographic history between of The Olive-striped and Streak-necked Flycatcher (the montane Mionectes) and between populations for each of these species. The amount of differentiation between species that barely overlap ranges along elevation is similar to the differentiation observed between populations within the same species across the Andean ranges. Because divergence times broadly overlapped between and within the same species, discordant patterns of differentiation in plumage and vocal traits between populations within the same species as compared with distinctive plumage and vocal phenotypes between species cannot be simply explained by rapid differentiation due to genetic drift alone.

Tropical rainforests are among the most structurally complex environments on earth. In these ecosystems, animals disperse the seeds of up to 90% of tree species, a process that influences vegetation structure in turn. In this dissertation, I use animal tracking and remote sensing to reveal elements of a feedback loop between 3D rainforest structure and movements of seed-dispersing birds and bats in southern Cameroon. In Chapter 1, I synthesize current research in a literature review that describes the feedback between vegetation structure and animal ecological roles. This chapter provides a worked example for applying the feedback loop concept to conservation problems. In Chapter 2, I assess how 3D rainforest structure can shape seed dispersal patterns by influencing movements and habitat selection of black-casqued (Ceratogymna atrata) and white-thighed hornbills (Bycanistes albotibialis). The seed dispersal model in this chapter incorporates hornbill selection for canopy height, vertical complexity, and distance to canopy gaps in a framework that maps spatial probabilities of seed dispersal. In addition, the results show that white-thighed hornbills tend to avoid swamp habitats, while black-casqued increase their preference for swamps during the hottest temperatures—a key niche difference between the species. In Chapter 3, I use recent advances in spaceborne remote sensing to show how 3D vegetation structure influences hammer-headed bat (Hypsignathus monstrosus) habitat selection over their full movement trajectories. Hammer-headed bats prefer swamp habitats, intermediate canopy height, and areas closer to canopy gaps. Chapter 4 combines approaches from Chapters 2 and 3 to show how seed dispersal patterns by hornbills arise from diversity in both movement behavior and landscape-scale habitat selection. Black-casqued and white-thighed hornbills clustered into three “movement syndromes” based on variation in movement distances and home range size. Seed dispersal models involving hornbills from all three movement syndromes yielded the greatest diversity in both distances and directions of seed dispersal routes. The results of this dissertation show the importance of movement behavioral diversity for seed dispersal and provide further evidence for a feedback loop between vegetation structure and animal ecological roles.