The primary function of erythrocytes is transport of oxygen from lungs to various tissues of the body. Red blood cell mass, due to this important role, must be controlled at precise levels. The number of erythrocytes is primarilly increased by the glycoprotein hormon erythropoietin, which expression is controlled by HIF (hypoxia inducible factor). Transcriptional factor HIF consists of the two subunits, HIFα and HIFβ. Under normoxic conditions, alfa subunit of HIF is hydroxylated by PHD protein. This hydroxylation provides a recognition motif for the VHL protein, a part of an E3 ubiquitin ligase complex that targets hydroxylated HIF for proteasomal degradation. Under hypoxic conditions, the degradation is inhibited. The alfa subunit is translocated to the nucleus, where binds the beta subunit and regulates gene expression. HIF pathway regulates a broad spectrum of cellular functions - energy metabolism, angiogenesis, apoptosis and many others. This diploma thesis is focused on HIF2α and its role in erythropoiesis. In this present study, we used CRISPR/Cas9 technology and created HEL (human erythroleukemia) cell line with knock-out of the gene for HIF2α (EPAS1). To reveal the role of HIF2α, we used specific HIF2α inhibitor in order to block its function in HEL cell line. We also tested this...

O objetivo do presente estudo foi o de investigar, pela primeira vez: os efeitos da administração prolongada de sementes Senna occidentalis (S. occidentalis) em órgãos hematopoiéticos de ratos, utilizando metodologias que poderão ser sugeridas para estudo pré-clínico de medicamentos. Avaliou-se nestes animais: consumo de ração e água; peso médio semanal; parâmetros hematológicos e bioquímicos; padrões histopatológicos; estoque de ferro e ensaios clonogênicos. O estudo foi realizado em ratos Wistar machos de 90 dias de idade, os quais foram expostos a diferentes concentrações de S. occidentalis na ração, a saber: 0% (controle), 0,5% (So0,5%), 1% (So1%) e 2% (So2%) durante 90 dias. Foi importante incluir um grupo pair-fed (PF), o qual recebeu a mesma quantidade de ração consumida pelo grupo So2%, porém, isenta da planta, uma vez que a S. occidentalis é sabidamente anorexígena. O presente estudo demonstrou que os ratos do grupo So2% apresentaram diminuição no número de leucócitos totais bem como alterações nos parâmetros referentes a série vermelha, tais como diminuição do VCM e HCM, caracterizando uma anemia microcítica hipocrômica. Os dados do hemograma corroboram as alterações observadas no mielograma destes animais, uma vez que foi constatada a redução significante da relação Mielóide/Eritróide (M/E) e, portanto, um possível efeito tóxico da S. occidentalis sobre a medula óssea durante o tratamento de 90 dias. A redução significante da relação M/E nos animais pertencentes aos diferentes grupos experimentais ocorreu devido ao aumento progressivo de eritroblastos jovens e policromáticos na medula óssea destes animais. Índices baixos da relação M/E podem estar associados a uma anemia regenerativa em função de hemólise extravascular ou, ainda, à eritropoiese ineficaz. A partir do mielograma observou-se, também, redução significante de células blásticas e de todos os tipos celulares, especialmente, da linhagem granulocítica, com predomínio da linhagem linfocitária. No entanto, uma vez que os linfócitos estão, continuadamente, recirculando, o aumento ou a diminuição destas células não reflete, necessariamente, alteração na linfopoiese. A análise anatomopatológica da medula óssea de animais dos grupos experimentais apontou um aumento progressivo da celularidade, caracterizando hiperplasia medular. Estes resultados se correlacionam aos dados do mielograma. Na anemia regenerativa ocorre diminuição da sobrevida das hemácias na circulação, resultante de hemólise extravascular realizada por macrófagos, especialmente, do tecido esplênico. Assim, o tratamento prolongado com S. occidentalis poderia estar associado ao um processo hemolítico, haja vista a observação de aumento do peso relativo deste órgão; acúmulo de hemossiderina no baço e ainda, reticulocitose em animais tratados com esta planta. O tratamento de 90 dias com S. occidentalis promoveu, também, redução significante na celularidade do baço bem como alterações anatomopatológicas, incluindo espessamento de cápsula e rarefação celular. Os ensaios clonogênicos padronizados neste estudo apontaram redução da porcentagem de colônias BFU-E e CFU-E em animais do grupo So2%, indicando uma provável ação tóxica da S.occidentalis sobre os progenitores medulares. A partir dos dados apresentados no presente estudo, pode-se concluir que o tratamento por 90 dias com S. occidentalis na ração é hematotóxico. Além disto, os resultados aqui obtidos bem como as metodologias empregadas poderão contribuir para o estabelecimento de um protocolo de hematotoxicidade, haja vista que informações relacionadas aso mecanismos de hematopoiese; metabolismo de ferro; características histológicas e citológicas de órgãos hematopoiéticos (baço e medula óssea) de ratos foram levadas em consideração para a realização deste estudo. / The purpose of this study was to investigate, for the first time: the effects of long term administration of Senna occidentalis (S. occidentalis) seeds in hematopoietic organs of rats, using methodology that can be suggested for pre-clinical studies. We evaluated some parameters in these animals: feed intake and water; weight body; hematological and biochemical parameters; histopathology; iron stores and clonogenic assays. The study was conducted in 90-day-old male Wistar rats, which were exposed to different concentrations of S. occidentalis in feed, ie: 0% (control), 0.5% (So0.5%), 1% (So1%) and 2 % (So2%) for 90 days. It was important to include a pair-fed group (PF), which received the same amount of feed consumed by So2% group, however, without the plant, once S. occidentalis is known anorectic. This study showed that rats from So2% group presented a decrease in the number of total leukocytes as well as changes in the parameters for the red series, such as decrease in MCV and MCH values, featuring a hypochromic anemia. The blood count results corroborate the findings from bone marrow smears of these animals, since it was found the significant reduction ratio Myeloid / Erythroid (M / L) and, therefore, a possible toxic effect of the S. occidentalis on the bone marrow during the treatment of 90 days. The significant reduction in the ratio M / E in animals belonging to different experimental groups occurred due to the progressive increase of young and polychromatic erythrocytes in the bone marrow of these animals. Low levels of M / E ratio may be associated with a regenerative anemia in extravascular hemolysis function or even the ineffective erythropoiesis. From the myelogram there was also a significant reduction of blast cells and all cell types, especially of granulocytic lineage, with a predominance of lymphoid lineage. However, since lymphocytes are, continuously by recirculating the increase or decrease in these cells do not necessarily reflect changes in lymphopoiesis. The pathological examination of the bone marrow of the experimental group animals showed a progressive increase in cellularity, featuring medullary hyperplasia. These results correlate to myelogram. In the regenerative anemia occurs a decreased survival of red blood cells in the circulation, resulting from extravascular hemolysis performed by macrophages, especially in the splenic tissue. Thus, the long term administration of S. occidentalis seeds could be associated with hemolytic process, due to the observation of an increase in the relative weight of this body; hemosiderin accumulation in the spleen and also increased reticulocytes in animals treated with this plant. The treatment with S. occidentalis promoted also significant reduction in cellularity of the spleen and pathological changes, including capsule and cell thinning thickening. Clonogenic assays showed decreased percentage of BFU-E and CFU-E colonies in animals from So2% group, indicating a probable toxic action of S.occidentalis on medullary progenitors. Therefore, the treatment for 90 days with S. occidentalis in feed is promotes blood toxicity.

L’érythropoïèse est le processus aboutissant à la production des hématies à partir d’une cellule souche hématopoïétique. La différenciation érythroïde implique des changements morphologiques en partie liés à la perte d’expression membranaire du récepteur à activité tyrosine kinase de classe III, c-Kit. En réponse à son ligand, le SCF, c-Kit est activé puis internalisé et dégradé par la voie du protéasome, via l’ubiquitine E3-ligase c-Cbl, ou par la voie lysosomale suite à une endocytose. Dans la première partie de ce travail, nous avons pu mettre en évidence qu’en absence de SCF et en réponse à un inhibiteur de tyrosine kinase, l’imatinib, les érythroblastes cultivés ex vivo perdent l’expression membranaire de c-Kit et accélèrent leur entrée en différenciation terminale. Au vu de ces observations, nous avons cherché à comprendre les mécanismes impliqués. Sur un modèle de cellules érytholeucémiques dépendantes de l’érythropoïétine, mais exprimant de manière endogène c-Kit, nous avons montré que l’imatinib induit une internalisation du récepteur ainsi que sa dégradation par la voie lysosomale et de manière indépendant de c-Cbl. De plus, nous avons montré que cet effet est réversible et que l’imatinib ne bloque pas la réexpression de c-Kit après son internalisation en réponse au SCF. Des marquages métaboliques ont permis de montrer que l’imatinib ne modifie ni la synthèse ni la maturation de c-Kit et que le profil phospho-tyrosine des cellules traitées à l’imatinib est globalement inchangé. Enfin, nous avons montré que la fixation de l’imatinib à la poche catalytique de c-Kit est indispensable à son internalisation, et par conséquent à sa dégradation. Il apparait donc que l’imatinib lève l’auto-inhibition de c-Kit, qui semble nécessaire pour son maintien à la membrane. Dans la seconde partie de ce travail, nous nous sommes intéressés aux changements morphologiques subis par les nucléoles, lieu de la biogenèse des ribosomes, au cours de différenciation des érythroblastes. L’étude de la taille et du potentiel prolifératif des cellules, ainsi que l’analyse morphologique des nucléoles, nous a permis de confirmer que la réduction de taille des cellules est contemporaine d’un ralentissement de leur prolifération ainsi que de la réduction du volume et de la surface du composé granulaire (CG), « matrice » du nucléole. En microscopie électronique, nous montrons la persistance des CG en fin de maturation. Enfin, nous avons également étudié l’évolution des nucléoles dans un contexte pathologique de syndromes myélodysplasiques de faible risque, qui se caractérisent par une hématopoïèse inefficace. Nous observons que les cellules pathologiques immatures ont des CG plus volumineux que les cellules normales immatures, et qu’au cours de la différenciation, la morphologie des nucléoles est identique entre les cellules normales et pathologiques. En conclusion, ce travail a permis de décrire 1) le mécanisme d’internalisation d’un récepteur à activité tyrosine kinase de classe III, c-Kit par l’imatinib et 2) la morphologie du nucléole au cours de la différenciation érythroïde normale et pathologique des syndromes myélodysplasiques de faible risque. / Erythropoiesis is the process leading to the production of red blood cells from hematopoietic stem cell. The erythroid differentiation involves morphological cell changes, in part related to the loss of membrane expression of the type III receptor tyrosine kinase, c-Kit. In response to its ligand SCF, c-Kit is activated, then internalized and degraded by the proteasome pathway via the E3 ubiquitin ligase c-Cbl, or by the lysosomal pathway, after endocytosis. In the first part of this work, we demonstrated that in the absence of SCF and in response to tyrosine kinase inhibitor, imatinib, erythroblasts cultured ex vivo, lose membrane expression of c-Kit and accelerate their terminal differentiation. In view of these observations, we sought to understand the mechanisms involved. On an erythropoietin dependent cell line expressing c-Kit at the membrane, we showed that imatinib induces receptor internalization and degradation by the lysosomal pathway, independently of c -Cbl. Furthermore, we showed that this effect is reversible and that imatinib does not block the c-Kit re-expression after its internalization, in response to SCF. Metabolic labelling showed that imatinib does not alter synthesis or maturation of c -Kit and that the phospho-tyrosine profile of cells treated with imatinib is generally unchanged. Finally, we showed that the binding of imatinib to the catalytic pocket of c-Kit is essential for its internalization, and therefore its degradation. So, it appears that imatinib removes c-Kit self-inhibition, which seems necessary to its retention at the membrane. In the second part of this work, we studied the morphological changes of nucleoli, the site of ribosome biogenesis, during erythroid differentiation. We showed that the reduction of cell size takes place at the same time than reduction of cell proliferation and reduction of surface and volume of the Granular Compound (GC), the “matrix” of the nucleolus. Moreover, we showed by electronic microscopy, the persistence of GC at the end of maturation. Finally, we also studied the evolution of nucleoli in a pathological context of low risk myelodysplastic syndromes, which are characterized by ineffective hematopoiesis. We observed that immature pathological cells have larger GC than immature normal cells, but that during differentiation, the morphology of nucleoli is identical between normal and pathological cells. In conclusion, this work has allowed us to describe 1) the mechanisms of internalization of a class III receptor tyrosine kinase, c-Kit by imatinib and 2) the morphology of the nucleolus during normal and pathological low risk myelodysplastic syndromes of erythroid differentiation.

O objetivo do presente estudo foi o de investigar, pela primeira vez: os efeitos da administração prolongada de sementes Senna occidentalis (S. occidentalis) em órgãos hematopoiéticos de ratos, utilizando metodologias que poderão ser sugeridas para estudo pré-clínico de medicamentos. Avaliou-se nestes animais: consumo de ração e água; peso médio semanal; parâmetros hematológicos e bioquímicos; padrões histopatológicos; estoque de ferro e ensaios clonogênicos. O estudo foi realizado em ratos Wistar machos de 90 dias de idade, os quais foram expostos a diferentes concentrações de S. occidentalis na ração, a saber: 0% (controle), 0,5% (So0,5%), 1% (So1%) e 2% (So2%) durante 90 dias. Foi importante incluir um grupo pair-fed (PF), o qual recebeu a mesma quantidade de ração consumida pelo grupo So2%, porém, isenta da planta, uma vez que a S. occidentalis é sabidamente anorexígena. O presente estudo demonstrou que os ratos do grupo So2% apresentaram diminuição no número de leucócitos totais bem como alterações nos parâmetros referentes a série vermelha, tais como diminuição do VCM e HCM, caracterizando uma anemia microcítica hipocrômica. Os dados do hemograma corroboram as alterações observadas no mielograma destes animais, uma vez que foi constatada a redução significante da relação Mielóide/Eritróide (M/E) e, portanto, um possível efeito tóxico da S. occidentalis sobre a medula óssea durante o tratamento de 90 dias. A redução significante da relação M/E nos animais pertencentes aos diferentes grupos experimentais ocorreu devido ao aumento progressivo de eritroblastos jovens e policromáticos na medula óssea destes animais. Índices baixos da relação M/E podem estar associados a uma anemia regenerativa em função de hemólise extravascular ou, ainda, à eritropoiese ineficaz. A partir do mielograma observou-se, também, redução significante de células blásticas e de todos os tipos celulares, especialmente, da linhagem granulocítica, com predomínio da linhagem linfocitária. No entanto, uma vez que os linfócitos estão, continuadamente, recirculando, o aumento ou a diminuição destas células não reflete, necessariamente, alteração na linfopoiese. A análise anatomopatológica da medula óssea de animais dos grupos experimentais apontou um aumento progressivo da celularidade, caracterizando hiperplasia medular. Estes resultados se correlacionam aos dados do mielograma. Na anemia regenerativa ocorre diminuição da sobrevida das hemácias na circulação, resultante de hemólise extravascular realizada por macrófagos, especialmente, do tecido esplênico. Assim, o tratamento prolongado com S. occidentalis poderia estar associado ao um processo hemolítico, haja vista a observação de aumento do peso relativo deste órgão; acúmulo de hemossiderina no baço e ainda, reticulocitose em animais tratados com esta planta. O tratamento de 90 dias com S. occidentalis promoveu, também, redução significante na celularidade do baço bem como alterações anatomopatológicas, incluindo espessamento de cápsula e rarefação celular. Os ensaios clonogênicos padronizados neste estudo apontaram redução da porcentagem de colônias BFU-E e CFU-E em animais do grupo So2%, indicando uma provável ação tóxica da S.occidentalis sobre os progenitores medulares. A partir dos dados apresentados no presente estudo, pode-se concluir que o tratamento por 90 dias com S. occidentalis na ração é hematotóxico. Além disto, os resultados aqui obtidos bem como as metodologias empregadas poderão contribuir para o estabelecimento de um protocolo de hematotoxicidade, haja vista que informações relacionadas aso mecanismos de hematopoiese; metabolismo de ferro; características histológicas e citológicas de órgãos hematopoiéticos (baço e medula óssea) de ratos foram levadas em consideração para a realização deste estudo. / The purpose of this study was to investigate, for the first time: the effects of long term administration of Senna occidentalis (S. occidentalis) seeds in hematopoietic organs of rats, using methodology that can be suggested for pre-clinical studies. We evaluated some parameters in these animals: feed intake and water; weight body; hematological and biochemical parameters; histopathology; iron stores and clonogenic assays. The study was conducted in 90-day-old male Wistar rats, which were exposed to different concentrations of S. occidentalis in feed, ie: 0% (control), 0.5% (So0.5%), 1% (So1%) and 2 % (So2%) for 90 days. It was important to include a pair-fed group (PF), which received the same amount of feed consumed by So2% group, however, without the plant, once S. occidentalis is known anorectic. This study showed that rats from So2% group presented a decrease in the number of total leukocytes as well as changes in the parameters for the red series, such as decrease in MCV and MCH values, featuring a hypochromic anemia. The blood count results corroborate the findings from bone marrow smears of these animals, since it was found the significant reduction ratio Myeloid / Erythroid (M / L) and, therefore, a possible toxic effect of the S. occidentalis on the bone marrow during the treatment of 90 days. The significant reduction in the ratio M / E in animals belonging to different experimental groups occurred due to the progressive increase of young and polychromatic erythrocytes in the bone marrow of these animals. Low levels of M / E ratio may be associated with a regenerative anemia in extravascular hemolysis function or even the ineffective erythropoiesis. From the myelogram there was also a significant reduction of blast cells and all cell types, especially of granulocytic lineage, with a predominance of lymphoid lineage. However, since lymphocytes are, continuously by recirculating the increase or decrease in these cells do not necessarily reflect changes in lymphopoiesis. The pathological examination of the bone marrow of the experimental group animals showed a progressive increase in cellularity, featuring medullary hyperplasia. These results correlate to myelogram. In the regenerative anemia occurs a decreased survival of red blood cells in the circulation, resulting from extravascular hemolysis performed by macrophages, especially in the splenic tissue. Thus, the long term administration of S. occidentalis seeds could be associated with hemolytic process, due to the observation of an increase in the relative weight of this body; hemosiderin accumulation in the spleen and also increased reticulocytes in animals treated with this plant. The treatment with S. occidentalis promoted also significant reduction in cellularity of the spleen and pathological changes, including capsule and cell thinning thickening. Clonogenic assays showed decreased percentage of BFU-E and CFU-E colonies in animals from So2% group, indicating a probable toxic action of S.occidentalis on medullary progenitors. Therefore, the treatment for 90 days with S. occidentalis in feed is promotes blood toxicity.

Doctor of Philosophy (PhD) / Gene expression is known to be regulated at the level of transcription. Recently, however, there has been a growing realisation of the importance of gene regulation at the post-transcriptional level, namely at the level of pre-mRNA processing (5’ capping, splicing and polyadenylation), nuclear export, mRNA localisation and translation. Erythroid krüppel-like factor (Eklf) is the founding member of the Krüppel-like factor (Klf) family of transcription factors and plays an important role in erythropoiesis. In addition to its nuclear presence, Eklf was recently found to localise to the cytoplasm and this observation prompted us to examine whether this protein has a role as an RNA-binding protein, in addition to its well-characterised DNA-binding function. In this thesis we demonstrate that Eklf displays RNA-binding activity in an in vitro and in vivo context through the use of its classical zinc finger (ZF) domains. Furthermore, using two independent in vitro assays, we show that Eklf has a preference for A and U RNA homoribopolymers. These results represent the first description of RNA-binding by a member of the Klf family. We developed a dominant negative mutant of Eklf by expressing its ZF region in murine erythroleukaemia (MEL) cells. We used this to investigate the importance of this protein in haematopoietic lineage decisions by examining its effect on the multipotent K562 cell line. We provide evidence that Eklf appears to be critical not only for the promotion of erythropoiesis, but also for the inhibition of megakaryopoiesis.

Controlling fates of stem and progenitor cells is one of the central goals of regenerative medicine. However, conventional cell enumeration methods are unable to distinguish between the effects of cell death, proliferation, and differentiation through molecular interventions on the output of specific cell types. We have devised a strategy to simultaneously obtain proliferation and death rates in cultures of highly purified erythroid progenitors. The approach is based on combining cell-surface marker analysis, cell-division tracking and 7-amino-actinomycin-D staining to monitor cell death. A compartment model of cell proliferation was developed to evaluate cell generation-specific length of cell-division, rates of entry into division, and cell death, from the experimental cell-division tracking data obtained following stimulation with erythropoietin (EPO) and Stem cell factor (SCF). The results indicated that EPO and SCF, either as single factor or in combination, differentially affect the rates of differentiation, length of cell-division and rates of death.

Controlling fates of stem and progenitor cells is one of the central goals of regenerative medicine. However, conventional cell enumeration methods are unable to distinguish between the effects of cell death, proliferation, and differentiation through molecular interventions on the output of specific cell types. We have devised a strategy to simultaneously obtain proliferation and death rates in cultures of highly purified erythroid progenitors. The approach is based on combining cell-surface marker analysis, cell-division tracking and 7-amino-actinomycin-D staining to monitor cell death. A compartment model of cell proliferation was developed to evaluate cell generation-specific length of cell-division, rates of entry into division, and cell death, from the experimental cell-division tracking data obtained following stimulation with erythropoietin (EPO) and Stem cell factor (SCF). The results indicated that EPO and SCF, either as single factor or in combination, differentially affect the rates of differentiation, length of cell-division and rates of death.

Thesis (Ph.D.)--University of Tennessee Health Science Center, 2009. / Title from title page screen (viewed on February 4, 2010). Research advisor: John M. Cunningham, M.D. Document formatted into pages (xiv, 115 p. : ill.). Vita. Abstract. Includes bibliographical references (p. 89-103).

Utilization of iron by hemoglobin-producing cells is highly efficient. The acquisition of iron from plasma requires the binding of diferric transferrin (Tf) to its cognate receptor (Tf-R) on the erythroid cell membrane, followed by internalization of the Tf - Tf-R complexes via receptor-mediated endocytosis. Through a poorly understood mechanism, iron is targeted to mitochondria, the site of heme biosynthesis. We believe that a direct interaction between iron-containing endosomes and mitochondria is essential for iron transfer to mitochondria and its efficient incorporation into heme. / In order to illustrate the interaction between endosomes and mitochondria, we have employed flow cytometry. Flow cytometry analysis of reticulocytes (erythrocyte precursors which still synthesize hemoglobin) stained with fluorescent dyes specific to mitochondria and endosomes revealed three distinct populations: mitochondria, endosomes and a population labeled with both dyes. This double-labeled population suggests a population composed of endosomes associated with mitochondria. Using non-fluorescent diferric-Tf, we were able to remove the double population, leaving only the endosomal and the mitochondrial population. This finding has confirmed that the double population is the result of the interaction between the two organelles. / Additionally, we established a cell-free assay consisting of fluorescent mitochondria and endosomes isolated from erythroid cells. Using confocal microscopy, we demonstrated a colocalization between the two organelles. We repeated the assay using fluorescent mitochondria and endosomes isolated from HeLa spinner cells. Using the mitochondrial uncoupler CCCP, we were able to significantly reduce the colocalization between the two organelles, indicating that the interaction between the organelles is specific and that the mitochondrial potential is a requirement for organellar interaction. / Based on our results from flow cytometry and confocal microscopy, we conclude that a specific and direct interaction exists between the two organelles.

There is a strong association between estrogen exposure and breast cancer risk. Estrogen can activate estrogen receptor alpha (ERalpha) to increase cell proliferation. Estrogen can also be metabolized into genotoxic compounds to induce DNA damage and mutations. Activation of the aryl hydrocarbon receptor (AHR) and nuclear factor erythroid-2 like 2 (NFE2L2; NRF2) can alter the production of genotoxic estrogen. The present thesis investigated the signalling mechanisms of ERalpha, AHR, and NRF2 and how their interaction might modulate breast cancer risk. In Chapter 2, genome-wide, but promoter-focused analysis of ERalpha binding sites in T-47D breast cancer cells identified potential cell line specific differences in estrogen signalling between T-47D and the commonly used MCF-7 breast cancer cells. CYP2B6 was identified to be an ERalpha target gene in T-47D cells but not MCF-7 cells, supporting cell line dependent effect in estrogen signalling. In Chapter 3 and 4, genome-wide analyses of AHR binding sites were performed to investigate the molecular criteria governing genomic AHR transactivation in vivo in mouse and in vitro in MCF-7 breast cancer cells. Our analysis identified 1) the previously established aryl hydrocarbon response element to be an important, but not an absolute requirement in AHR transactivation and 2) key epigenetic modifications that modulate AHR-dependent gene regulation. Lastly, in Chapter 5, interaction among ERalpha, AHR, and NRF2 was presented at the regulatory region of two NRF2 target genes, NADPH Quinone Oxidoreductase 1 (NQO1) and Heme Oxygenase 1 (HMOX1). ERalpha repressed, whereas AHR enhanced NRF2-dependent NQO1 and HMOX1 mRNA expression through altered p300 recruitment and Histone H3 Lysine 9 acetylation. Collectively, this thesis examined novel molecular mechanisms that might alter breast cancer development/progression by modulating ER, AHR, and NRF2 activity.