Honey or Vinegar: Oneota Interaction in the Central and Northeastern Plains

Abstract Beginning AD 1150 and extending until European contact, the archaeological culture referred to as “Oneota” underwent an explosive spread across the American midcontinent. As Oneota ideas, people, or some combination thereof moved westward, they encountered people from other cultures. Along the western frontier of Oneota culture, evidence suggests that relations between Oneota and Plains indigenes took a variety of forms. To better understand how various environmental and cultural factors may have informed the decision-making process with regard to inter-group interaction, four sites along this western Oneota periphery were selected for analysis: Shea and Sprunk in eastern North Dakota, White Rock in north-central Kansas, and Dixon in northwest Iowa. The evidence for both inter-group contact and site function is evaluated and compared across these four sites, and ultimately synthesized with existing knowledge and theories of Oneota interaction. It is suggested that Oneota social relations may have been partially dependent on whether other groups were in competition for a similar resource base; this process allowed a relationship between Oneota and Psinomani peoples to flourish, while minimizing the possibility of positive relations with Central Plains Tradition peoples. This hypothesis offers directions for future research, including the extent of the relationship between Oneota and Psinomani peoples and the movement of commodities from western Oneota outposts to the Great Lakes and Upper Midwest regions often viewed as the Oneota heartland

cis-Regulation in the Mammalian Rod Photoreceptor

Transcription factors regulate the expression level of target genes by binding to cis-regulatory elements (CREs) present in gene promoters. The goal of my thesis research is to define the sequence components of CREs that determine transcriptional output. In order to accomplish this goal, I developed a method to measure the regulatory activity of thousands of CREs in a single experiment. In this method I insert unique barcodes in the 3\u27UTR of a reporter gene and multiplex expression measurements with RNA sequencing. Using this technique in explanted retinas, I determined the impact of single nucleotide variants in a mammalian promoter by measuring expression controlled by all single nucleotide variants of the Rhodopsin proximal promoter. I found that nearly all (86%) sequence variants drive significantly different activity than the wild-type promoter and that the mechanism of most variants can be interpreted as altered transcription factor binding. In addition, we found that the largest changes in expression resulted from variants located in characterized transcription factor binding site sequences. Next, I explored how combinations of binding sites drive particular levels of gene expression by utilizing a synthetic biology approach. I generated synthetic CREs composed of various combinations of binding sites found in the Rhodopsin promoter and measured the expression driven by these sequences. In this study I found that synthetic CREs containing binding sites for transcriptional activators yielded diverse expression outputs, including both activation and repression of a minimal promoter. Together, these experiments demonstrate that interactions between binding sites and dual regulation of a single binding site can produce diverse gene expression patterns. I conclude that simple cis-regulatory elements can produce complex expression outputs due to interactions between transcriptional activators and detailed quantitative models will be necessary to predict expression from these sequences

Methods for detecting multi-locus genotype-phenotype association

Solutions to the genotype-phenotype problem seek to identify the set of genetic mutations and interactions between them which modify risk for and severity of a trait of interest. I propose association graph reduction (AGR), a novel algorithm to detect such genetic lesions in genome-wide data, particularly in the presence of high-order interactions. I describe several existing methods and evaluate their performance in terms of computational cost and power to detect associations. An objective comparison of the results shows that AGR successfully combines high power with computational efficiency, while providing a detailed account of interactions present in the data. No other known method combines these three properties. When applied to real data, AGR can be used to discover genetic causes of common diseases such as arthritis, hypertension, diabetes, asthma, and many others, which will facilitate the discovery of novel diagnostic tools and treatment protocols

Self-recruitment in anemonefish and the impact of marine ornamental fishery in Spermonde Archipelago, Indonesia: implications for management and conservation

The yearly amount of traded by middlemen on Spermonde Archipelago (Indonesia) is estimated to about 140,000 specimens A. ocellaris and more than 31,000 anemones. Both A. ocellaris and sea anemone densities (p < 0.01) were significantly lower at reefs with a high exploitation than at reefs with a low exploitation. The size of A. ocellaris individuals was significantly smaller in Barrang Lompo than in Samalona (p < 0.01). The private alleles and allelic richness in Samalona were 4% significantly higher than in Barrang Lompo. The allelic richness was positively correlated with the fish density (p < 0.05). Fish stays largely in place despite its pelagic larvae stage (44 % - 60.7 % self-recruitment). The genetic relatedness revealed a close relation between individuals within a group of A. ocellaris in Barrang Lompo. Conversely, unrelated individuals of A. ocellaris in Samalona and of A. perideraion in Barrang Lompo were observed. Altogether, these results provide important insights how marine ornamental fishery has impacted the population of anemonefish and its host in Spermonde Archipelago

Regulation of gene expression in macrophage immune response

Gene expression quantitative trait loci (eQTL) mapping studies can provide mechanistic insights into the functions of disease-associated variants. However, many eQTLs are cell type and context specific. This is particularly relevant for immune cells, whose cellular function and behaviour can be substantially altered by external cues. Furthermore, understanding mechanisms behind eQTLs is hindered by the difficulty of identifying causal variants. We differentiated macrophages from induced pluripotent stem cells from 86 unrelated, healthy individuals derived as part of the Human Induced Pluripotent Stem Cells Initiative. We generated RNA-seq data from these cells in four experimental conditions: naïve, interferon- gamma (IFNɣ) treatment (18h), Salmonella infection (5h), and IFNγ treatment followed by Salmonella infection. We also measured chromatin accessibility with ATAC-seq in 31-42 individuals in the same four conditions. We detected gene expression QTLs (eQTLs) for 4326 genes, over 900 of which were condition-specific. We also detected a similar number of transcript ratio QTLs (trQTLs) that influenced mRNA processing and alternative splicing. Macrophage eQTLs and trQTLs were enriched for variants associated with Alzheimer’s disease, multiple autoimmune disorders and lipid traits. We also detected chromatin accessibility QTLs (caQTLs) for 14,602 accessible regions, including hundreds of long-range interactions. Joint analysis of eQTLs with caQTLs allowed us to greatly reduce the set of credible causal variants, often pinpointing to a single most likely variant. We found that caQTLs were less condition- specific than eQTLs and ~50% of the stimulation-specific eQTLs manifested on the chromatin level already in the naive cells. These observations might help to explain the discrepancy between strong enrichment of diseases associations in regulatory elements but only modest overlap with current eQTL studies, suggesting that many regulatory elements are in a ‘primed’ state waiting for an appropriate environmental signal before regulating gene expression.Wellcome Trust scholarship for the PhD programme in Mathematical Genomics and Medicin

The role of integrins in flavivirus infection

The Flavivirus genus (Flaviviridae family) comprises the most important arboviruses in the world such as dengue virus, West Nile virus (WNV), Zika virus (ZIKV), Japanese encephalitis virus and yellow fever virus (YFV). Every year, several outbreaks caused by flaviviruses are reported worldwide (i.e.: ZIKV and YFV outbreaks in South America) with a huge impact on economy and public health. In the last few decades, many aspects of the flavivirus biology and the interaction of flaviviruses with host cells have been elucidated. However, many underlying mechanisms concerning receptor usage, entry process and viral interaction with host cell factors are still not completely understood. Integrins, the major class of cell adhesion molecules have been implicated in the infectious cycle of different viruses including flaviviruses. A previous report proposed that a particular integrin, the αVβ3 integrin, might act as a cellular receptor for WNV. However, this hypothesis was not confirmed by other groups. In the present study, murine cell lines lacking the expression of one or more integrin subunits were used to evaluate the involvement of different integrins in the flavivirus infection cycle. Mouse fibroblasts lacking the expression of β1 integrin (MKF-β1-/-) or β3 integrin (MEF-β3-/-) subunits or αVβ3 integrin (MEF-αVβ3-/-) as well as their corresponding wild-type cells were utilized. A second model using Chinese hamster ovary cells (CHO-K1), a cell line that has been described to be refractory to some flaviviruses, were modified to express either αV (CHO-αV+/+) or β3 (CHO-β3+/+) integrin subunits. All cell lines were first characterized by confocal laser microscopy, flow cytometry and functional assays prior to infection to assess their integrin expression. The cell lines were then inoculated with different flaviviruses of public health relevance: WNV, YFV-17D, Usutu virus (USUV), Langat virus (LGTV) and ZIKV. Infection assays were designed in order to evaluate whether integrins influence i) cell susceptibility; ii) binding; iii) internalization and iv) replication of the investigated flaviviruses. Our findings clearly demonstrate that β1, β3 and αVβ3 integrins do not act as flavivirus cellular receptor or attachment factor since their ablation does not completely abrogate flavivirus infection in the investigated cell lines. Flavivirus binding to the cell surface of MEFs, MKFs and CHO cells was not disturbed by the genomic deletion of the above-mentioned integrins. The deletion of β1 and β3 integrin subunit did not affect internalization of any of the flaviviruses tested. In contrast to that, loss of αVβ3 integrin in the MEF-αVβ3-/- cells showed a statistically significant decrease in WNV and USUV internalization while ZIKV, YFV-17D and LGTV internalization remained unaffected suggesting that αVβ3 integrin might be involved in the internalization process of at least some flaviviruses. On the other hand, flavivirus replication was substantially impaired in the integrin-deficient cell lines in comparison to their corresponding wild-type cells. Both, MEF-β3-/- and MKF-β1-/- cells showed a statistically significant reduction on viral load for all flaviviruses tested in comparison to their respective wild-type cells. The MEF-αVβ3-/- cells in particular, showed a strong inhibition of flavivirus replication with a reduction of up to 99% on viral loads for all flaviviruses tested. Levels of flavivirus negative-strand RNA were substantially decreased in MEF-αVβ3-/- cells indicating that integrins might influence flavivirus RNA replication. The ectopic expression of either αV or β3 integrin subunits in CHO cells slightly increased the replication of all flaviviruses tested. Taken together, this is the first report highlighting the involvement of integrins in ZIKV, USUV, LGTV and YFV infection. The results strongly indicate that the investigated integrins play an important role in flavivirus infection and might represent a novel host cell factor that enhances flavivirus replication. Although the exact mechanism of interaction between integrins and flaviviruses is currently unknown, the results provided in this study deepen our insight into flavivirus - host cell interactions and open doors for further investigations.Die Gattung der Flaviviren (Familie Flaviviridae) beinhaltet einige der wichtigsten Arboviren weltweit, beispielsweise das Dengue Virus, das West-Nil Virus (WNV), das Zika Virus (ZIKV), das Japanische-Enzephalitis Virus sowie das Gelbfieber Virus (YFV). Jedes Jahr kommt es zu zahlreichen, durch Flaviviren verursachten Ausbrüchen (u.a. Zika und Gelbfieber Virus Ausbrüche in Südamerika), die mit immensen Auswirkungen auf die Ökonomie und das öffentliche Gesundheitswesen einhergehen. Obwohl die Interaktion von Flaviviren mit verschiedenen Wirtszellen in den letzten Jahrzehnten intensiv untersucht wurde und wichtige Fragen in der Flavivirus Biologie bereits geklärt werden konnten, sind viele zugrundeliegende Mechanismen, u.a. die virale Rezeptornutzung, der Eintrittsprozess sowie die Interaktion mit verschiedenen Wirtszellfaktoren nicht vollständig verstanden. Integrine, eine der wichtigsten Klasse von Zelladhäsionsmolekülen, wurden bereits in der Literatur beschrieben, eine Rolle im Infektionszyklus verschiedener Viren, u.a. auch der Flaviviren, zu spielen. Es gibt zudem Hinweise, dass ein bestimmtes Integrin, das αVβ3 Integrin, als Zellrezeptor für WNV fungieren kann, wobei diese Hypothese bislang nicht weiter bestätigt werden konnte. In dieser Arbeit wurde der Einfluss von bestimmten Integrinen auf die Flavivirusinfektion in verschiedenen, genetisch modifizierten Maus- und Hamsterzelllinien untersucht. Hierfür wurden zum einen Mausfibroblasten verwendet, die für die Expression von β1 oder β3 Integrin Untereinheiten oder für das αVβ3 Integrin deletiert sind (MKF-β1-/-; MEF-β3-/- und MEF-αVβ3-/-), um diese in Infektionsexperimenten mit den entsprechenden Wildtypzellen zu vergleichen. Zum anderen wurde die Chinese Hamster Ovary (CHO) Zelllinie genutzt, welche in der Literatur als refraktär gegenüber bestimmten Flaviviren beschrieben wurde. Diese Zelllinie wurde im Rahmen der Studie genetisch so modifiziert, dass entweder die αV (CHO-αV+/+) oder die β3 (CHO-β3+/+) Integrin Untereinheit exprimiert wurde. Alle rekombinanten Zelllinien sowie deren Wildtyp wurden mittels Konfokalmikroskopie, Durchflusszytometrie und funktionalen Assays bezüglich der Integrinexpression charakterisiert. Anschließend wurden die Zellen mit den folgenden, Public Health relevanten Flaviviren inokuliert: WNV, YFV, ZIKV, Usutu Virus (USUV) und Langat Virus (LGTV). In diesen Experimenten wurde der Einfluss der beschriebenen Integrine auf i) zelluläre Empfänglichkeit; ii) Bindung; iii) Internalisierung und iv) Replikation der verwendeten Flaviviren untersucht. Die Ergebnisse der Studie zeigen, dass die untersuchten Integrine in den verwendeten Maus- und Hamsterzelllinien weder als Zellrezeptor noch als Attachment-Faktor dienen. Die fehlende Expression der Integrine verhindert in keinem Fall die Infektion der Zellen. Unabhängig von der Integrinexpression können alle untersuchten Flaviviren an die entsprechenden Zellen binden und internalisiert werden. Die Deletion der β1 und β3 Integrin Untereinheiten zeigt keinen Effekt auf die Internalisierung der untersuchten Flaviviren. Das Fehlen des αVβ3 Integrins in den MEF-αVβ3-/- Zellen hingegen resultiert in einem statistisch signifikanten Unterschied in der Internalisierung von WNV und USUV im Vergleich zu den entsprechenden Wildtypzellen während die Internalisierung von ZIKV, YFV-17D und LGTV unbeeinträchtigt bleibt. Diese Ergebnisse deuten darauf hin, dass αVβ3 Integrin in die Internalisierung bestimmter Flaviviren involviert sein könnte. Die Flavivirusreplikation zeigt sich in den Integrin-defizienten Zellen in dieser Studie stark beeinträchtigt im Vergleich zu den Wildtypzellen. Die Deletion der β1 und β3 Untereinheiten resultiert in einer statistisch signifikant verminderten Replikation in den entsprechenden Mausfibroblasten. Eine noch deutlichere Beeinträchtigung der Replikation aller untersuchter Flaviviren mit einer Reduktion der Viruslast um bis zu 99% wird zudem in den MEF-αVβ3-/- Zellen beobachtet. Diese Ergebnisse werden zusätzlich durch deutlich reduzierte Mengen an detektierbarer Negativstrang-RNA in den MEF-αVβ3-/- Zellen unterstützt, was auf einen Einfluss der Integrine auf die Flavivirusreplikation hinweist. Die ektopische Expression der beschriebenen Integrine in CHO Zellen resultiert ebenfalls in einem leichten Anstieg der Flavivirusreplikation. Insgesamt ist dies der erste Bericht, der die Beteiligung von Integrinen in ZIKV, USUV, LGTV und YFV Infektionen beschreibt. Die Ergebnisse dieser Studie deuten stark darauf hin, dass bestimmte Integrine eine entscheidende Rolle in der Flavivirusinfektion spielen und möglicherweise einen neuen Wirtszellfaktor für Flaviviren darstellen. Auch wenn ein eindeutiger Mechanismus für die Interaktion von Integrinen mit Flaviviren bislang nicht bekannt ist, können die gewonnenen Ergebnisse dieser Studie den Anstoß für weiterführende Untersuchungen geben

From condition-specific interactions towards the differential complexome of proteins

While capturing the transcriptomic state of a cell is a comparably simple effort with modern sequencing techniques, mapping protein interactomes and complexomes in a sample-specific manner is currently not feasible on a large scale. To understand crucial biological processes, however, knowledge on the physical interplay between proteins can be more interesting than just their mere expression. In this thesis, we present and demonstrate four software tools that unlock the cellular wiring in a condition-specific manner and promise a deeper understanding of what happens upon cell fate transitions. PPIXpress allows to exploit the abundance of existing expression data to generate specific interactomes, which can even consider alternative splicing events when protein isoforms can be related to the presence of causative protein domain interactions of an underlying model. As an addition to this work, we developed the convenient differential analysis tool PPICompare to determine rewiring events and their causes within the inferred interaction networks between grouped samples. Furthermore, we present a new implementation of the combinatorial protein complex prediction algorithm DACO that features a significantly reduced runtime. This improvement facilitates an application of the method for a large number of samples and the resulting sample-specific complexes can ultimately be assessed quantitatively with our novel differential protein complex analysis tool CompleXChange.Das Transkriptom einer Zelle ist mit modernen Sequenzierungstechniken vergleichsweise einfach zu erfassen. Die Ermittlung von Proteininteraktionen und -komplexen wiederum ist in großem Maßstab derzeit nicht möglich. Um wichtige biologische Prozesse zu verstehen, kann das Zusammenspiel von Proteinen jedoch erheblich interessanter sein als deren reine Expression. In dieser Arbeit stellen wir vier Software-Tools vor, die es ermöglichen solche Interaktionen zustandsbezogen zu betrachten und damit ein tieferes Verständnis darüber versprechen, was in der Zelle bei Veränderungen passiert. PPIXpress ermöglicht es vorhandene Expressionsdaten zu nutzen, um die aktiven Interaktionen in einem biologischen Kontext zu ermitteln. Wenn Proteinvarianten mit Interaktionen von Proteindomänen in Verbindung gebracht werden können, kann hierbei sogar alternatives Spleißen berücksichtigen werden. Als Ergänzung dazu haben wir das komfortable Differenzialanalyse-Tool PPICompare entwickelt, welches Veränderungen des Interaktoms und deren Ursachen zwischen gruppierten Proben bestimmen kann. Darüber hinaus stellen wir eine neue Implementierung des Proteinkomplex-Vorhersagealgorithmus DACO vor, die eine deutlich reduzierte Laufzeit aufweist. Diese Verbesserung ermöglicht die Anwendung der Methode auf eine große Anzahl von Proben. Die damit bestimmten probenspezifischen Komplexe können schließlich mit unserem neuartigen Differenzialanalyse-Tool CompleXChange quantitativ bewertet werden

Die Anwendung von Mikrosatelliten zum Studium der Sozialstruktur bei Großsäugetieren : am Beispiel von italienischem Wolf und Damwild

Scandura M. The use of microsatellites in the study of social structure in large mammals : Italian wolf and fallow deer as case studies. Bielefeld (Germany): Bielefeld University; 2004.The content of the present PhD thesis deals with the application of microsatellite analysis to the study of two species of large mammals, referring to some aspects of their social and mating systems. The Italian wolf (Canis lupus) and the fallow deer (Dama dama) were chosen as case studies, since genetic investigations on their populations result, for different reasons, problematic. The wolf in Italy is a particularly protected species, recovering throughout the peninsula from the effects of a recent bottleneck. Sampling wolves may not rely on capturing or killing them, therefore an alternative, non-invasive, approach was adopted in my study. Scats, shed hairs and blood drops collected on the snow represented the main source of DNA for the analysis. Methodological cares were necessary to obtain reliable wolf genotypes. A set of ten canine microsatellites was employed to achieve unique multilocus genotypes in the population. Fifty-two individuals were typed in the period 1998 - 2003. In some cases, pack composition was determined, confirming that familiar bonds are at the basis of wolf social units. An unpredictable high local differentiation was found among geographic areas. Early dispersal seem to be common in the population, but its effects on the gene flow are not detectable, at least at my study scale. I proposed that most of this dispersal may be unsuccessful or over long distances. The study population, indeed, seem to have reached a high level of local saturation, with clumped pack territories and high reproductive rates, and thus possibly represents a source, from which wolves disperse toward sink areas. The second study involves an enclosed population of fallow deer. Lekking is seldom observed in mammals, and among them, it is more common in ungulate species. Fallow deer is one of the most studied lekking ungulate and this particular population is object of long-term researches on male mating strategies. Mature bucks in the population join into leks during the breeding season: a costly strategy, which apparently does not guarantee high direct benefits (fitness). In this study, for the first time, I used a molecular approach to verify the existence of a genetic basis to lek formation. The recourse to microsatellites enabled to overcome the strong limitation due to the extremely monomorphism of the species, documented in several studies. Though the low variability even at microsatellite loci, the hypotheses of kin selection (territorial bucks in a lek are relatives) and of heterozygosity advantage (territorial bucks in a lek have an overall high heterozygosity) were tested and resulted not supported by data. Hence, future studies should be addressed towards phenotypic variation and consider in detail physiological and ecological factors, in order to clarify the reasons why lekking takes place in fallow deer