Regulation of gene expression is vital for proper growth and prevention of disease states. In eukaryotes this regulation occurs in the context of chromatin which creates an inherent barrier for the binding of trans-acting factors, such as transcription factors and RNA polymerase. This dissertation focuses on the role of transcriptional activators and chromatin remodeling coactivators in the regulation of the repressible acid phosphatase gene PHO5. Our studies show that histone methylation at lysine 4 of histone H3 is required for the full repression of PHO5and GAL1-10. We show that bromodomains, a domain conserved in chromatin remodeling coactivators, may function to stabilize binding. Finally, we present a strategy using DNA methyltransferases as in vivo probes to detect DNA-protein interactions and examine chromatin structure. We extend this strategy to zinc-finger proteins which can be engineered to bind to any desired DNA sequence as a means of targeting methylation with potential use in epigenetic silencing.

Television advertising is a billion-dollar industry in the United States. Currently, advertisers place their messages in television programs that are estimated to have a high proportion of their target demographic viewers. The advertising spots are usually purchased months in advance at set list prices or at negotiated prices. Technologies that can place advertisements at the cable box level, instead of the program level, will provide advertisers with the ability to target any demographic group directly and in real-time. This thesis explores the new decision-making required by this new technology and how the television advertisement space can be sold more effectively. In particular, it compares a list price system to a number of new auction models. The structure of the auctions for the new targeted television advertising system is unique and has not been previously studied in the literature. This thesis explores new auction models that can capture these unique features and lead to desirable results for the seller of the advertisement space. A simplified analytical model shows how these features impact advertisers’ bidding behavior and how a list price system compares to the auction models in the ability to raise revenue for the seller of the advertising space. These issues are then explored under various market settings with differing numbers of advertisers and value distributions that these advertisers have for the advertising space. Since sequential first price auctions have undesirable consequences such as strong price fluctuations, this work focuses on second price auctions. The Vickrey-Clarke-Groves (VCG) mechanism is customized for this problem by developing an optimization formulation that determines the best set of advertisers for a particular advertisement space. Because execution time may be an issue, other auction models are developed that lead to similar outcomes as the VCG mechanism but require less computational effort. This thesis provides guidance on when a list price system will lead to higher expected revenue than an auction model and vice versa in a targeted television advertising system. It also demonstrates why some of the standard auction models cannot be applied to this problem and what type of new models are required to lead to desirable advertising outcomes.

Television advertising is a billion-dollar industry in the United States. Currently, advertisers place their messages in television programs that are estimated to have a high proportion of their target demographic viewers. The advertising spots are usually purchased months in advance at set list prices or at negotiated prices. Technologies that can place advertisements at the cable box level, instead of the program level, will provide advertisers with the ability to target any demographic group directly and in real-time. This thesis explores the new decision-making required by this new technology and how the television advertisement space can be sold more effectively. In particular, it compares a list price system to a number of new auction models. The structure of the auctions for the new targeted television advertising system is unique and has not been previously studied in the literature. This thesis explores new auction models that can capture these unique features and lead to desirable results for the seller of the advertisement space. A simplified analytical model shows how these features impact advertisers’ bidding behavior and how a list price system compares to the auction models in the ability to raise revenue for the seller of the advertising space. These issues are then explored under various market settings with differing numbers of advertisers and value distributions that these advertisers have for the advertising space. Since sequential first price auctions have undesirable consequences such as strong price fluctuations, this work focuses on second price auctions. The Vickrey-Clarke-Groves (VCG) mechanism is customized for this problem by developing an optimization formulation that determines the best set of advertisers for a particular advertisement space. Because execution time may be an issue, other auction models are developed that lead to similar outcomes as the VCG mechanism but require less computational effort. This thesis provides guidance on when a list price system will lead to higher expected revenue than an auction model and vice versa in a targeted television advertising system. It also demonstrates why some of the standard auction models cannot be applied to this problem and what type of new models are required to lead to desirable advertising outcomes.

Regulation of gene expression is vital for proper growth and prevention of disease states. In eukaryotes this regulation occurs in the context of chromatin which creates an inherent barrier for the binding of trans-acting factors, such as transcription factors and RNA polymerase. This dissertation focuses on the role of transcriptional activators and chromatin remodeling coactivators in the regulation of the repressible acid phosphatase gene PHO5. Our studies show that histone methylation at lysine 4 of histone H3 is required for the full repression of PHO5and GAL1-10. We show that bromodomains, a domain conserved in chromatin remodeling coactivators, may function to stabilize binding. Finally, we present a strategy using DNA methyltransferases as in vivo probes to detect DNA-protein interactions and examine chromatin structure. We extend this strategy to zinc-finger proteins which can be engineered to bind to any desired DNA sequence as a means of targeting methylation with potential use in epigenetic silencing.

It has become increasingly apparent that the misregulation of histone modification actively contributes to cancer. The histone H3 lysine 79 (H3K79) methyltransferase Dot1l has been implicated in the development of leukemias bearing translocations of the Mixed Lineage Leukemia (MLL) gene. We studied the global epigenetic profile for H3K79 dimethylation and found abnormal H3K79 dimethylation profiles exist not only in leukemias driven by MLL-fusion proteins with nuclear partners like AF9, but also in leukemia with MLL-fusions containing cytoplasmic partners like AF6. Genetic inactivation of Dot1l led to downregulation of fusion target genes and impaired both in vitro bone marrow transformation and in vivo leukemia development by MLL-AF10, CALM-AF10 as well as MLL-AF6, suggesting that aberrant H3K79 methylation by DOT1L sustains fusion-target gene expression in MLL rearranged leukemias and CALM-AF10 rearranged leukemias. Pharmacological inhibition of DOT1L selectively killed MLL-AF10 and MLL-AF6 transformed cells but not Hox9/Meis1 transformed cells, pointing to DOT1L as a potential therapeutic target in MLL-rearranged leukemia. We further characterized the DOT1L complex under physiological conditions from human leukemia cells and identified AF10 as a key DOT1L complex component. Given the importance of H3K79 methylation in MLL-rearranged leukemia, we sought to study the role of DOT1L complex component AF10 in H3K79 methylation and MLL leukemia. We generated conditional knockout mice in which the Dot1l-interacting octapeptide-motif leucine-zipper (OM-LZ) domain of Af10 was flanked by LoxP sites. Cre induced deletion of \(Af10^{OM-LZ}\) is predicted to abrogate the Af10-Dot1l interaction. Our histone mass spectrometry data demonstrated that deletion of the endogenous \(Af10^{OM-LZ}\) domain abrogated global H3K79 dimethylation but retained H3K79 monomethylation. Interestingly, bone marrow transformation by MLLAF6 and MLL-AF9 is abrogated by induced deletion of endogenous \(Af10^{OM-LZ}\), while bone marrow transformation by MLL-AF10 and CALM-AF10 is not affected by deletion of endogenous \(Af10^{OM-LZ}\), confirming the importance of Af10-Dot1l interaction in MLL- or CALM fusion-leukemias. Moreover, we showed \(Af10^{OM-LZ}\) deletion prolonged survival of MLL-AF9 leukemia in vivo and led to chromotin compaction and downregulation of MLL fusion targets in MLL-AF9 leukemia. Therefore our results demonstrate a role for Af10 in the conversion of H3K79 monomethylation to dimethylation and reveal the AF10-DOT1L interaction as an attractive therapeutic target in MLL-rearranged leukemias.

Hemangioma is a type of tumor commonly found in infants that is characterized by heavy vascularization and a disfiguring appearance. Hemangioma, though benign, can sometimes proliferate and be threatening to infants. Current treatments for infantile hemangioma include surgical removal as well as the use of topical and oral medication. However, current therapies are often ineffective at treating lesions and are commonly accompanied by dangerous side effects, creating the need for a new, safer treatment. This study targets the Yes-Associated Protein-1 (YAP-1), which has been described as an oncogene, by use of an interfering RNA technique in attempts to mediate tumor growth and progression. Western blotting of treatment and control BEND3 murine cells reveals that YAP-1 is knocked-down in treatment groups which have been infected with shYAP-1 siRNA genes. By successfully knocking down the YAP-1 protein, the potential for developing a novel targeted therapy for infantile hemangioma has been established.

This study investigated teacher targeted bullying in primary schools to examine whether students serve as an unsafe work stressor for teachers, because teacher targeted bullying is on the rise. This study looked at teacher targeted bullying as an antecedent of stress and the mechanisms, such as mediators and moderators, which affected this bullying-stress relationship. 113 primary school teachers completed a self-report questionnaire which measured bullying, self-efficacy, safety climate perceptions, social support and stress. The results provided evidence that social support from others outside work moderated the mow level bullying-stress, and severe bullying-stress relationships. Most importantly, unsafe job perceptions mediated the relationship between low level bullying and stress. The results of this research imply that student bullying can influence teachers to perceive their job as unsafe and lead to stress. These findings can be used to develop effective strategies to not only prevent and manage bullying, but create safer schools for teachers and pupils.

The United States use of Unmanned Combat Aerial Vehicles (UCAV) or drones for targeted killings of terrorists has been on the rise in recent years and the method has become the core element of president Obamas strategy in the war against terror. This study examines the deterrent effect of targeted killings on terrorism using UCAV/drones as a method of counterterrorism. Building on the literature on counterterrorism, UCAV, targeted killings, deterrence theory and statistics on terrorism the study provides a case study of the CIA drone operations in Pakistan between the years 2004-2010. The goal has been to analyze drone operations and the extent of terrorism from the beginning of the drone campaign until 2010. This as a means of identifying possible trends in terrorism activity due to the occurrence of drone strikes. Findings suggest that the possibility of a deterrent effect is far from evident and that there are some indications of increased terrorism.

Freie Univ., Diss.--Berlin, 1997.

Univ., Diss.--Münster, 1993.