Digitized by Kansas Correctional Industries

This thesis is the study of the role and regulation of the external audit in the UK and US. It argues that the current audit model is fundamentally flawed, which has a negative impact on the quality of financial audits. This research contributes to the debate on external auditing by suggesting a free-standing legal re-conceptualization of the audit model, which aims to improve the quality of audits and reduce opportunity for accounting fraud. The research findings are drawn from qualitative analysis of legal and financial accounting sources. The thesis consists of three main themes. Firstly, it provides a comprehensive analysis of the regulatory framework for auditing by looking at why, how and by whom audit is regulated. Secondly, it analyses numerous flaws inherent in the current audit model. The focus throughout the thesis is on problems relating to auditors' independence, deficiencies in the spheres of legal, professional and social accountability of auditors, and excessive concentration of the audit market. Thirdly, following the analysis of various theoretical and international audit arrangements, the thesis suggests a new audit model. A key function in this model would be played by an external, public body - the Public Auditing Board, which would be in charge of appointment and remuneration of auditors carrying out audits in the public interest. It is submitted that moving audits to the public sector is desirable in order to introduce genuinely autonomous auditors, to provide better quality of audits and to restore public confidence in capital markets.

Sepsis affects hundreds of thousands each year in the United States alone, with an estimated 20-30% mortality rate in spite of current treatment regimens. Sepsis mortality was originally understood to be the caused by overproduction of inflammatory cytokines in response to pathogen detection by the host. However, recent studies suggest that with modern treatments, secondary infection, rather than inflammatory shock, may be of greater concern. In either case, the failure of a large number of anti-inflammatory agents to produce beneficial outcomes in sepsis treatment during clinical trial suggests that the development of a new class of immunomodulatory agents may be required for effective treatment. In experimental models, pre-treatment with sub-lethal doses of lipopolysaccharide (LPS, previously referred to as endotoxin) induces a state of "LPS tolerance" that reduces septic shock lethality. Paradoxically, LPS tolerance also results in increased antimicrobial gene expression and resistance to secondary infection in some models. Further exploration of this process may provide drug targets capable of limiting inflammation without dampening antimicrobial immunity, which could be of great benefit in the treatment of sepsis and chronic inflammatory disease. Many groups have studied signaling changes that occur during LPS tolerance. However, mediators of tolerance that can account for the changes in LPS-induced gene expression that result in increased microbial resistance are not well described. This has prevented proper testing of the physiological effects of tolerance on disease, and it remains unclear if this process could be artificially induced or is of any benefit to sepsis patients. Recent in vitro work suggests that tolerant gene expression patterns are the result of large scale changes in chromatin organization that occur in macrophages after prolonged LPS stimulation. Because microRNAs (miRNAs), a new class of gene regulator, have been found to regulate chromatin modifying complexes in other systems, LPS-induced miRNAs were screened to identify potential mediators of tolerance that could cause changes in gene expression patterns without necessarily impacting LPS signaling itself. Several tolerance-associated miRNAs were identified. One miRNA in particular, miR-222, was found to repress tumor necrosis factor (Tnf) and Brahma-related gene one (Brg1) expression. This attenuates expression of genes dependent on nucleosome remodeling, primarily affecting inflammatory genes. Consequently, miR-222 expression effectively limits septic shock lethality. However, low-level responses, as well as NF-κB signaling and the expression of a subset of antimicrobial and antiviral genes, are left intact. Thus, although miR-222 does not entirely recapitulate the tolerance response, by directing the LPS response into a less damaging expression profile, miR-222 may accelerate the onset of tolerance and be a promising target for therapeutics aiming to treat inflammatory disease without compromising host immunity.

No description available.

This thesis considers the applicability of 'new governance' techniques within the field of emerging biotechnologies. Through three contrasting case studies I construct an argument in favour of new governance, contending that the qualities of this regulatory trend (flexibility, reflexivity, nuance, open discourse, and participation - 'regulatory desirables' ) have much to offer the regulation of emerging biotechnologies. The first case study examines the existing European and international regulatory frameworks for genetically modified organisms (GMOs). This case study explores the role of (bio)ethics within the regulatory process through each progressive stage: design, operation, and assessment. The regime's failure to provide adequate space for ethical reflection, and the limited role of ethics throughout the regulatory process prompts a proposal for an alternative approach that recognizes the multiple contexts in which regulation operates, and is able to accommodate the socio-ethical nuances of the GMO products being assessed. This case study analyses a traditionally structured regulatory framework. It exemplifies a number of qualities that I consider undesirable in the context of regulating biotechnologies: inflexibility, lack of reflexivity, lack of nuance within the regime, absence of ethical discussion, absence of participation from all interested/affected parties. In the second and third case studies I show how these 'regulatory undesirables' can be addressed through new governance techniques. The second case study focuses on the international regulation of stem cell research; I propose developing a polycentric, principles-based regulation (PBR) regime. The third case study centres on the international governance of the gene synthesis industry; here I recommend adopting a risk-based regulation (RBR) approach. In both these fields, voluntary, interdisciplinary, international organisations have collaborated to produce guidelines, codes, protocols, standards, and statements addressing matters of practice. I argue that these 'soft law' documents form the ideal starting point for the development of more sophisticated regulatory regimes in both fields. Furthermore, I argue that the informal organisations producing these documents are, in certain instances, best placed to step into the role of 'regulator' due to their in-depth, inside knowledge of the field, and network. Thus, I collapse the regulator-regulatee distinction held in traditional, 'command and control' style systems, as these organisations typically include those who would traditionally be seen as the 'regulatee'. Each case study considers the nuances of context vis-à-vis the regulatory approach advocated. I conclude by engaging in a comparative analysis of these three case studies, drawing out the qualities, characteristics and considerations that I regard as essential to the construction of responsible, facilitative governance frameworks across the field of emerging biotechnologies. I conclude that new governance is best suited to achieving these (aforementioned) 'regulatory desirables'.

Moloney Murine Leukemia Virus (M-MLV) proviral DNA is transcriptionally silenced in mouse embryonic cells by a repressor complex containing tripartite-motif-containing 28 (Trim28). Trim28 depends on post-translational modifications, such as sumoylation and phosphorylation, and its interactions with several co-repressor proteins to regulate its repressive activity. YY1 is one such Trim28 co-repressor protein, recently found to tether the Trim28 silencing complex to the M-MLV promoter. Here, we investigated the biochemical interaction of Trim28 and YY1, and the role of sumoylation and phosphorylation of Trim28 in mediating M-MLV silencing. Experiments probing the binding of YY1 and Trim28 in vitro suggested that their interaction occurs indirectly. Mutational studies demonstrated that the RBCC domain of Trim28 is sufficient for interaction with YY1 while the acidic region 1 and zinc fingers of YY1 were necessary and sufficient for its interaction with Trim28. Additionally, we found that the K779 residue was critical for Trim28-mediated silencing of M-MLV in embryonic cells. The repressor complex that silences M-MLV is very large and likely consists of many protein subunits. A few proteins contained in the repressor complex have been identified, including Trim28, but the identity of most of the components forming the repressor complex are unknown. We detected a new form of the complex that is of even high molecular weight and likely contains additional associated cofactors. We reported an approach for purifying this larger repressor complex and identified new candidates for cofactors that may potentially function in the silencing of M-MLV. We also examined the regulation of sumoylation in embryonic cells. Sumoylation conjugation is a post-translational modification that affects a diverse range of processes and is important for embryo survival. Overall inhibition of the SUMO pathways results in embryonic lethality demonstrating the importance of the SUMO pathways for embryonic viability; however, our understanding of SUMO function in embryos at the cellular and molecular level is still greatly lacking. We demonstrated that SUMO1 cannot be overexpressed in embryonic carcinoma and embryonic stem cells and that SUMO1 overexpression is prevented at a post-transcriptional level. This occurred specifically for SUMO1 and not for SUMO2 overexpression. Furthermore, blocking conjugation or increasing the deconjugation of SUMO1 to substrates significantly improved SUMO1 overexpression. The results indicate that the overexpression of SUMO1 protein, in itself, is tolerated in embryonic cells, but the accumulation of substrate(s) modified by SUMO1 appears to be strongly prevented by an embryonic-specific post-transcriptional mechanism.

No description available.

Pax6 is a highly conserved developmental regulator with a complex temporal, spatial and quantitative expression pattern, that is crucial for correct development of the central nervous system, the eye, and pancreas. Accordingly, the Pax6 gene resides in a complex genomic locus containing a large array of long-range tissue-specific cis-regulatory elements primarily identified through multispecies sequence conservation and reporter studies. I have set out to understand how the chromatin architecture of the locus contributes to the mechanism and specificity of cis-regulatory interactions. As well as addressing whether the DNA looping model for regulatory interactions applies to the mouse Pax6 locus, I will identify which elements facilitate such interactions and if they vary between cell types. Utilising ChIP-array technology the distribution and variability of key regulatory histone modifications and factors were assessed in a set of Pax6 expressing and non-expressing mouse cell lines, acting as models for different regulatory states of the locus. Work in other loci suggests a key role for CTCF and cohesin (subunit Rad21) in chromatin organisation and long distance regulatory interactions. ChIP-chip for CTCF/Rad21 across the Pax6 locus identified numerous sites within the gene and at distal regulatory locations. The majority of these sites are cell type invariable. The active enhancer modification H3K27ac identified both known and several novel putative enhancer elements distributed through the locus that are highly cell type specific. A subset of CTCF/Rad21 sites also acquire the active enhancer modification H3K27ac in a cell type dependent manor, suggesting that CTCF/Rad21 may facilitate looping to the target gene from these sites. Using reporter based assays, putative regulatory elements marked by the looping factors and active histone modifications showed a diverse range of functional activities. Unexpectedly only 3 of the 7 CTCF sites tested showed classical insulator activity in an enhancer blocking reporter assay. Surprisingly the strongest insulator tested resided within intron 7 the Pax6 gene. Other CTCF/Rad21 sites were neutral or enhancers in the insulator assay. This reveals the disparity between predicting regulatory properties using ChIP binding profiles alone and the actual outcome of functional reporter experiments. A novel element, CTCF6 showed a ChIP signature of CTCF/Rad21/H3K27ac in all Pax6 expressing tissues, and functioned as a strong enhancer in transient transfection and stable LacZ reporter assays. CTCF6 recapitulated a broad range of Pax6 expression patterns, at multiple embryonic stages, including the brain, neural tube and pancreas. A second novel element, E-120 identified in the pancreatic derived cell line, drove stable embryonic reporter expression in the embryonic pancreas and sub set of brain regions. Together this has expanded the repertoire and size of Pax6’s regulatory landscape particular in the upstream region. Chromatin conformation capture (3C) was used to characterise the dynamic chromatin architecture of the locus and identify the interaction profiles from three CTCF/Rad21 binding regulatory locations within the Pax6 locus. This revealed a core set of regulatory interactions with the Pax6 gene, while individual elements showed a more variable set of cell type specific interactions. The CTCF6 enhancer showed highly cell type specific promoter interactions throughout the Pax6 gene, indicative of enhancer-promoter looping not detected in the non-expressing cells. While the downstream site CTCF5 at the edge of a cluster of regulatory elements known as the DRR (differentially regulated region), interacted with both the gene and an upstream element CTCF7 300 kb away only in the Pax6 expressing locus. Together these results reveal Pax6 has a chromatin hub structure with regulatory loops from upstream and downstream bringing distant yet variable active elements in to the vicinity of the Pax6 promoters where they can act. This work has revealed new roles for CTCF/cohesin sites in transcriptional regulation of Pax6 and how the cis-regulatory activity and structure of the locus varies across different cell types.

O objetivo deste trabalho é apresentar uma análise dos aspectos regulatórios e de comercialização sob a visão da sustentabilidade, com foco na questão de como a participação crescente das fontes consideradas incentivadas na matriz elétrica brasileira pode proporcionar benefícios energéticos e de redução nas emissões de gases de efeito estufa, contemplando possíveis aperfeiçoamentos regulatórios que possam vir a ser definidos pelas instituições do setor elétrico que detém essa competência. Destaca-se a seqüência de fatos que contribuíram para a ampliação da oferta das fontes alternativas (FA\'s), sublinhando: (i) a criação do Programa de Incentivo às FA\'s (PROINFA); (ii) o regramento para a comercialização de Energia Incentivada; (iii) o Mecanismo de Realocação de Energia (MRE); (iv) facilidades para compra de Geração Distribuída pelas distribuidoras, mitigar o risco de variação de mercado; (v) Leilões de Fontes Alternativas e (vii) Leilões de Energia de Reserva (LER). Nesse âmbito de análise, inclui-se uma aferição de risco de comercialização da Energia Incentivada e de como esse risco pode ser mitigado através de hedge contratual entre fontes incentivadas. Para avaliar e qualificar a sinergia entre energia eólica e da biomassa, que tem perfil de produção complementar às Pequenas Centrais Hidrelétricas (PCH\'s), foram realizadas simulações de análise de complementariedade de geração para identificar a melhor estratégia de contratação de energia, a fim de buscar a máxima receita líquida possível, atendendo restrições de risco. Finalmente, são apresentadas propostas de alterações regulatórias e comerciais que estão sendo discutidas no setor e que ainda não estão aprovadas, porém são consideradas relevantes e impactantes no que diz respeito à energia incentivada, destacando-se a possibilidade de cessão de energia excedente pelos consumidores livres e especiais; a criação de penalidades por alavancagem; a proposta de criação do Comercializador varejista; e dos certificados de energia (CEE\'s). / This work is aiming at to present an analysis of the regulatory aspects and commercialization, under sustainability point of view, focusing with special emphazis the question on how the increasing of incentivized sources participation in the Brazilian energy matrix can provide energy benefits and reduction of greenhouse emissions, considering possible regulatory improvements that may be defined by the adequate institutions of the Brazilian electrical sector (BES). Highlighting the sequence of events that contributed to increase the expansion of alternative sources (FA\'s), it should be worthwhile to stress: (i) the creation of the Incentive Program for FA\'s (PROINFA); (ii) the rules for the commercialization of the Energy with Incentive; (iii) the Energy Reallocation Mechanism (MRE); (iv) facilities for the purchase of Distributed Generation by distributors having the purpose of mitigating the market risk represented by the exposure to the short term market price (Market Cleasing Price); (v) Energy Auctions of Alternative sources (vii) Energy Auctions Reserve (LER). The context of the analysis carried out includes the assessment of commercialization risk affecting FA\'s and how this risk can be mitigated through of \"hedge\" contracts between different FA\'s. To evaluating and qualify the synergy between Wind and biomass plants, which has production with complementary profile to the Small Hydro Power (PCH\'s), simulations were performed to identify the best strategy to energy contracting, considering the objective of maximum profit under riskconstraints. Finally, some proposals of commercial and regulatory changes are presented, being important to emphasize that the quoted proposals are now in a discussion process in the BES, meaning that they are not approved yet. However, it is relevant to consider these news possibilities, as they impact the economic feasibility of incentivized energy, highlighting the possibility of to sell eventual surplus in the short term marketby free consumer; the creation of penalties for leverage; the creation of an agent focused on small free consumers (special consumers) commercialization, and energy certificates (CEE\'s).

The Na+-D-glucose cotransporter in small intestine is regulated in response to food composition. Short term regulation of SGLT1 occurs post-transcriptionally in response to changes in luminal glucose. Adaptation to dietary carbohydrate involves long term regulation at the transcriptional level. The intracellular protein RS1 (gene RSC1A1) is involved in transcriptional and post-transcriptional regulation of SGLT1. RS1 contains an N-terminal domain with many putative phosphorylation sites. By Expressing SGLT1 in oocytes of Xenopus laevis it was previously demonstrated that the post-transcriptional down-regulation of SGLT1 by RS1 was dependent on the intracellular glucose concentration and activated by protein kinase C (PKC). The role of RS1 for short term regulation of SGLT1 in mouse small intestine in response to glucose and PKC was investigated comparing effects in RS1-/- mice and wildtype mice. Effects on SGLT1 activity were determined by measuring phlorizin inhibited uptake of α-methylglucoside (AMG). The involvement of RS1 in glucose dependent short term regulation could not be elucidated for technical reasons. However, evidence for RS1 independent short-term downregulation of SGLT1 after stimulation of PKC could be provided. It was shown that this downregulation includes decrease in the amount and/or in turnover of SGLT1 in the brush-border membrane as well as an increase of substrate affinity for AMG transport. Trying to elucidate the role of RS1 in long term regulation of SGLT1 in small intestine in response to glucose and fat content of the diet, wildtype and RS1-/- mice were kept for 2 months on a normo-caloric standard diet with high glucose and low fat content (ND), on a hyper-caloric glucose-galactose reduced diet with high fat content (GGRD) or on a hyper-caloric diet with a high fat and high glucose content (HFHGD). Thereafter the animals were starved overnight and SGLT1 mediated AMG uptake was measured. Independent of diet AMG uptake in ileum was smaller compared to duodenum and jejunum. In jejunum of wildtype and RS1-/- mice kept on the fat rich diets (GGRD and HFHGH) transport activity of SGLT1 was lower compared to mice kept on ND with low fat content. This result suggests an RS1 independent downregulation due to fat content of diet. Different to RS1-/- mice, the duodenum of wildtype mice showed transport activity of SGLT1 smaller in mice kept on glucose galactose reduced diet (GGRD) compared to the glucose galactose rich diets (ND and HFHGG). These data indicate that RS1 is involved in glucose dependent long term regulation in duodenum. / Der Na+-Glukose-Cotransporter SGLT1 im Dünndarm wird in Abhängigkeit zur Nahrungszusammensetzung reguliert. Kurzzeitregulation von SGLT1 tritt posttranskritionell als Antwort zu sich ändernden Glukosekonzentrationen im Darmlumen auf. Anpassung an Nahrungskohlenhydrate beinhaltet die Langzeitregulation auf transkripionellem Level. Das intrazelluläre Protein RS1 (Gen RSC1A1) ist an der transkriptionellen und post-transkriptionellen Regulation von SGLT1 beteiligt. Es enthält eine N-terminale Domäne mit vielen putativen Phosphorylierungsstellen. Bei der Expression von SGLT1 im Xenopus leavis Oocytensystem wurde gezeigt, dass die posttranskriptionelle Herunterregulation von SGLT1 durch RS1 von der intrazelluläre Glukosekonzentration abhängt und durch Proteinkinase C (PKC) aktiviert wird. Die Rolle von RS1 in der Kurzzeitregulation von SGLT1 im Dünndarm der Maus als Antwort auf Glukose und PKC wurde durch vergleichende Studien zwischen RS1- knockout (RS1-/-)- Mäusen und Wildtyp-Mäusen untersucht. Effekte auf die SGLT1-Aktivität wurden durch Messung der durch Phlorizin inhibierbaren Aufnahme des SGLT1-spezifischen Substrats α-Methyl-Glycopyranosid (AMG) bestimmt. Der Einfluss von RS1 in der Glukose-abhängigen Kurzzeitregulation konnte aus technischen Gründen nicht untersucht werden, jedoch gab es Anzeichen für eine von RS1 unabhängige Kurzzeitregulation von SGLT1 durch PKC. Es wurde gezeigt, dass diese Herunterregulation sowohl eine Abnahme der Menge und/oder der Umsatzrate von SGLT1 in der Bürstensaummembran wie auch eine Zunahme der Substrat-Affinität für den AMG-Transport beinhaltet. Um die Rolle von RS1 auf die Langzeitregulation von SGLT1 in Dünndarm als Antwort auf den Glukose- und Fettgehalt der Nahrung zu untersuchen, wurden Wildtyp- und RS1-/- Mäuse für 2 Monate entweder auf einer normalenergetischen Standarddiät mit hohem Glukose- und niedrigem Fettgehalt (ND), auf einer hochenergetischen Diät mit reduziertem Glukose und Galaktose-Gehalt (GGRD) oder auf einer hochenergetischen Diät mit hohem Fett- und Glukosegehalt (HFHGD) gehalten. Danach wurden die Tiere über Nacht gefastet und die durch SGLT1 vermittelte AMG –Aufnahme gemessen. Unabhängig der Diät war die AMG-Aufnahme im Ileum geringer als in Duodenum und Jejunum. Im Jejunum von Wildtyp- und RS1-/- Mäusen die auf einer fettreichen Diät (GGRD und HFHGD) gehalten wurden war die Transportaktivität von SGLT1 geringer verglichen mit der Aktivität von Mäusen auf ND. Dieses Ergebnis lässt eine RS1-unabhängige Herunterregulation die durch den Fettgehalt hervorgerufen wird vermuten. Anders als in RS1-/- Mäusen war die Transportaktivität von SGLT1 im Duodenum von Wildtypmäusen bei der Glukose-Galaktose- reduzierten Diät niedriger verglichen mit den Glukose-Galaktose-reichen Diäten (ND und HFHGD). Diese Daten legend die Vermutung nahen, das RS1 an der Glukose-abhängigen Langzeitregulation im Duodenum beteiligt ist.