Global ETD Search

1	The role of c-Myb in mammary gland development and tumourigenesis Miao, Yu Rebecca January 2009 (has links) c-Myb/MYB is an established and key player in hematopoietic malignancies but more recently a strong case for c-Myb as an oncogene in breast cancer has emerged. c-Myb and its transcriptional target genes have direct bearing on tumour initiation and progression and thus this has opened new opportunities to the development of therapeutic approaches in a range of cancer types with the aim of treating cancer at its various stages. In this study, the requirement of c-Myb during mammary gland tumourigenesis is being examined. In addition a direct therapeutic approach to targeting c-Myb-driven gene grp78/GRP78 in the context of primary and metastatic breast cancer was assessed. / The first aim of this study is to examine the expression of c-Myb during normal mammary gland development. The expression of c-Myb is extensively characterised in a temporal and spatial fashion. Nuclear staining of c-Myb by immunohistochemisty was found to be most elaborately expressed in the ductal epithelium during early mammary gland development. Mouse mammary gland lacking c-myb showed disorganised ductal structure in virgin mice, but did not affect subsequent pregnancy and lactation. / To extend the view that c-Myb is involved in mammary tumourigenesis c-myb-transduced immortalised mammary epithelial cells and two mammary tumour prone transgenic mouse models were examined. NMuMG cells transduced with c-myb showed enhanced proliferation and reduced Annexin V staining consistent with the protection from apoptosis. This reduced apoptosis is consistent with, and perhaps contingent upon, the elevated expression observed for bcl-2 and grp78. The data assembled by expression studies raised the possibility that c-Myb is essential for the establishment of mammary gland tumor in both MMTV-Neu and MMTV-PyMT spontaneous mammary gland tumor models. Loss of c-Myb expression in these models significantly delayed and in most instances completely abolished the onset of mammary gland tumours in both models. Preliminary evidence also indicated that Stat3 phosphorylation may underpin the elevated c-Myb expression in mouse mammary tumour cells. / The focus of my thesis then shifted to examining ways to exploit elevated c-Myb target gene GRP78 expression on the cell surface of mammary tumour cells. To do this I employed a GRP78 binding pro-apoptotic chimera peptide that specifically binds to GRP78 where I examined its efficacy against primary and metastatic breast cancer models. My results demonstrated the anti-tumour activity of the GRP78-chimera peptide both in vitro and in vivo. More importantly, the peptide is also effective at prolonging disease-free survival in mice with established metastatic disease. / Evidence obtained within these studies suggests that c-Myb plays an important role in mammary gland development and tumourgenesis. Although it may be difficult to directly target c-Myb in malignant disease, alternative anti-tumoural therapy may be developed against c-Myb-regulated target genes that are also implicated in mammary tumours. Collectively my thesis studies have advanced our understanding of c-Myb in mammary cancer initiation, progression and as a direct or indirect therapeutic target. mammary gland, c-Myb, tumourigenesis
2	The effects of 1,4-benzoquinone on c-Myb and topoisomerase II in K-562 cells Singh, Roopam 11 January 2008 (has links) Exposure to benzene, a ubiquitous environmental pollutant, has been linked to leukemogenesis, although the mechanism of benzene initiated carcinogenesis remains unclear. It has been proposed that benzene can be bioactivated to toxic metabolites such as 1,4 benzoquinone (BQ), which can alter signalling pathways and affect chromosomal integrity. BQ has been shown to increase the activity of c-Myb, which is an important transcription factor involved in hematopoiesis, cell proliferation, and cell differentiation. The c-Myb protein also increases topoisomerase IIα (topo IIα) promoter activity specifically in cell lines with hematopoietic origin. Topo II is a critical nuclear enzyme that removes torsional strain by cleaving, untangling and religating double-stranded DNA. Since topo II mediates DNA strand breaks, aberrant topo II activity or increased protein levels may increase the formation of DNA strand breaks, leaving the cell susceptible to mutational events. I hypothesize that BQ increases c-Myb activity, which in turn increases topo IIα promoter activity resulting in increased DNA strand breaks. Using luciferase reporter assays in K-562 cells (human chronic myeloid leukemic cells) I confirmed that BQ exposure (25 and 37 µM) caused an increase in c-Myb activity after 24 hours. Contradictory to previous findings, overexpression of exogenous c-Myb or a polypeptide consisting of c-Myb’s DNA binding domain (DBD), which competitively inhibits the binding of endogenous c-Myb to DNA, did not affect topo IIα promoter activity. However, BQ exposure (37 µM for 24 hours) caused a significant increase in topo IIα promoter activity, which could be blocked by the overexpression of the DBD polypeptide. Western immunoblotting analysis did not show any significant increases in topo IIα protein levels in cells exposed to 37 µM BQ for 24 hours. Overall, this study suggests that BQ exposure increases topo IIα promoter activity through the c-Myb signalling pathway and furthers our understanding of BQ-mediated toxicity. / Thesis (Master, Pharmacology & Toxicology) -- Queen's University, 2008-01-02 14:09:00.011 Benzene c-Myb Topoisomerase II Benzoquinone
3	Characterization of Secondary DNA Structures Formed in the c-myb and hTERT Promoters and Their Potential Role in the Regulation of Transcription Palumbo, SunMi Lee January 2009 (has links) In this dissertation, the formation of unusual G-quadruplexes in the critical regions of the c-myb and hTERT promoters for control of promoter activity was investigated.The c-myb promoter contains three copies of an almost perfect (GGA)4 sequence. We demonstrate that the each (GGA)4 repeat forms a tetrad:heptad G-quadruplex and any two of the three can intramolecularly dimerize to form T:H:H:T G-quadruplexes. The three T:H:H:T G-quadruplex combinations are of differing degrees of stability and can be further stabilized by G-quadruplex interactive compounds. We also demonstrate that the c-myb G-quadruplex forming region is a critical transcriptional regulatory element and interacts with various nuclear proteins including MAZ (Myc Associated Zinc finger protein). The data from luciferase reporter assay show that the c-myb GGA repeat region plays dual roles as a transcriptional activator and an inhibitor by serving as binding sites for the activators and by forming G-quadruplex structures in the region, respectively. Furthermore, we show that MAZ is a transcriptional repressor of the c-myb promoter and binds to both the double-stranded and T:H:H:T G-quadruplex-folded conformations of the GGA repeat region of the c-myb promoter.The hTERT core promoter contains a G-rich region of 12 consecutive G-tracts, which includes three critical Sp1 binding sites. Although this G-rich region has the potential to form multiple G-quadruplexes, our investigation on the full-length G-rich sequence demonstrate that the G-rich region forms a unique G-quadruplex structure in which two tandem intramolecular G-quadruplex structures are present, consisted of one G-quadruplex formed by the G-tracts 1-4 and the other formed by the G-tracts 5, 6, 11, and 12. We also demonstrate that the latter unusual structure contains a 26-base middle loop that likely forms a hairpin structure and is more stable than the other conventional G-quadruplex. Significantly, the formation of this unusual tandem G-quadruplex structure in the full-length will disable all three critical Sp1 binding sites, which will dramatically downregulate hTERT expression. G-quadruplex formation in the hTERT promoter suggests that the effect of G-quadruplex interactive ligands on telomerase inhibition and telomere shortening may be exerted by the direct interaction between the hTERT G-quadruplex structure and the ligands. c-myb G-quadruplex hTERT secondary DNA structure transcription
4	Critical roles for the transcription factor c-Myb in early B cell development Greig, K. T. January 2009 (has links) B cell development is a carefully orchestrated process involving many transcription factors acting in concert with cytokine signals, particularly IL-7. The transcription factor c-Myb has long been implicated in B cell development, however surprisingly little is known about the function of c-Myb in B cell progenitors. I have used several mouse models of c-Myb deficiency to investigate the role of c-Myb in the B cell lineage. Conditional deletion of c-Myb in early B cell progenitors using mb-1Cre (c MybΔmb1/Δmb1) leads to a striking lack of B cells from the pre-pro-B cell stage onwards, demonstrating that c-Myb is absolutely required for B cell development. Mice homozygous for a hypomorphic allele of c-Myb (c MybPlt4/Plt4) also display a severe reduction in B cells; in these mice, defects in lymphoid development can be detected within the multipotent progenitor compartment of bone marrow. c-Myb activates transcription via coactivator proteins, particularly CBP and p300. Mice bearing a point mutation in p300 (p300Plt6/Plt6) that inhibits the interaction of p300 with c Myb display a partial block in B cell development, highlighting the importance of the c Myb-p300 complex for B cell development. Together, these mice demonstrate that c-Myb regulates B cell development by functioning both in multipotent progenitor cells and directly in B cell progenitors. In addition, I show that the B-lymphopenia in c-Myb deficient mice is related to a profound defect in IL-7 signalling. IL-7 normally stimulates the proliferation, survival and differentiation of B cell progenitors, however pro-B cells from c-MybPlt4/Plt4 and c MybΔmb1/Δmb1 mice fail to respond to IL 7. Expression of the IL-7Rα chain is reduced on pro-B cells from c MybPlt4/Plt4 and c-MybΔmb1/Δmb1 mice, suggesting that Il7r may be a c-Myb target gene in B cells. Reporter gene assays show that c-Myb can activate the Il7r promoter in synergy with the transcription factor Pu.1. Overall, this work demonstrates that c-Myb is essential for early B cell development and plays a critical role in linking cytokine signals to the transcription factor networks in B cell progenitors.
5	Rapid induction of B-cell lymphomas : Insertional activation of thec-myb proto-oncogene / Kanter, Madge Ruth. January 1989 (has links) Thesis (Ph. D.)--Cornell University, 1989. / Vita. Includes bibliographical references.
6	Studies on human ribonuclease H1 and its action on 2'-fluoroarabinose oligonucleotide hybrid substrates Alla, Nageswara Rao January 2012 (has links) Ribonuclease H1 is a conserved enzyme that is localized in the nuclear and mitochondrial compartments of eukaryotic cells, and functions in DNA replication, repair, recombination and transcription. (Arunachandran et al., 2000; Cerritelli et al., 2003) Oligonucleotide binding to a complementary RNA sequence can provide a substrate for RNase H1, and provides the mechanistic basis for antisense oligonucleotide (AON)-directed gene silencing in cells (Opalinska et al., 2002). Effective evaluation of the therapeutic efficacy of next-generation AONs with novel structures requires an in vitro system involving, purified, highly active RNase H1 of human cells, and a full understanding of the catalytic mechanism of the enzyme. The goal of project 1 described in chapter 3, was to determine the involvement of a conserved Histidine (H264) in the catalytic mechanism of human RNase H1. Based on this analysis I was able to conclude that H264 has a dual role in phosphodiester hydrolysis and in product release. The goal of project 2 (Chapter 4) was to examine the reactivities towards human RNase H1 of model hybrid substrates containing specific types of 2'-FANA substitutions (abbreviated as `F', with 2'-deoxyribose abbreviated as `D'), either at the "wings" of the molecule ("7-gapmer"; each wing=7 nt: FFFFFFF-DDDDDDD-FFFFFFF), or with 3 nt alternations ("3-altimer": FFF-DDD-FFF-DDD-FFF-DDD-FFF). The results of this study strongly support the continued examination of the potential therapeutic utility of the 2'-FANA modification in AONs. The highly efficient and selective inhibition of protein expression is a primary basis of action of most antisense therapeutic strategies. These data suggest that the 2'-FANA modification supports sustained silencing after a single administration, either by mRNA cleavage or by a translational block, and at substantially lower concentrations compared to the unmodified AON. The results of this project underscore the proposal that 2'-FANA-modified AONs will be important additions to the repertoire of rational antisense strategies for the effective treatment of disease. / Chemistry Chemistry Biochemistry 2'-fana Antisense C-myb Rnase H1
7	Regulace transkripce proteiny rodin Early growth response a Myb / Regulation of transcription by proteins of the Early growth response and Myb families Čermák, Vladimír January 2013 (has links) The regulation of transcription of tens of thousands of genes in a vertebrate organism is an enormously complex phenomenon which entails the participation of thousands of various regulatory proteins. The largest functional category of these regulators is accounted for by sequence-specific DNA-binding proteins known as transcription factors. Proteins of the EGR and Myb families of transcription factors are long-studied regulators of a variety of physiological processes including cellular proliferation and differentiation. The structural and physical aspects of their function have been well characterized. Their cell-type specific participation in complex gene-regulatory networks, on the other hand, is still incompletely understood and represents a major challenge in the respective research areas. Preliminary analysis of gene expression data from metastasizing PR9692 and non- metastasizing PR9692-E9 chicken sarcoma cell lines revealed that the transcription factor EGR1 is expressed at a higher level in metastasizing cells and can thus take part in the regulatory processes that underlie the differences between the two cell lines. Further investigation demonstrated that the introduction of exogenous EGR1 into PR9692-E9 cells restored their metastatic potential to a level indistinguishable from PR9692...
8	The Role of Cell Cycle Machinery in Ischemic Neuronal Death Iyirhiaro, Grace O. 09 October 2013 (has links) Ischemic stroke occurs as a result of a lack or severe reduction of blood supply to the brain. Presently therapeutic interventions are limited and there is a need to develop new and efficacious stroke treatments. To this end, a great deal of research effort has been devoted to studying the potential molecular mechanisms involved in ischemic neuronal death. Correlative evidence demonstrated a paradoxical activation of the cell cycle machinery in ischemic neurons. The levels and activity of key cell cycle regulators including cyclin D1, Cdk2 and Cdk4 are upregulated following ischemic insults. However, the functional relevance of these various signals following ischemic injury was unclear. Accordingly, the research described in this thesis address the functional relevance of the activation of the cell cycle machinery in ischemic neuronal death. The data indicate that the inhibition of Cdk4 protects neurons from ischemia-induced delayed death, whereas abrogation of Cdk5 activity prevents excitotoxicity-induced damage in vitro and in vivo. Examination of upstream activators of mitotic-Cdks showed that Cdc25A is a critical mediator of delayed ischemic neuronal death. Investigation of the potential molecular mechanism by which cell cycle regulators induced neuronal death revealed perturbations in the levels and activity of key downstream targets of Cdk4. The retinoblastoma protein family members, pRb and p130 are increasingly phosphorylated following ischemic stresses. Importantly, p130 and E2F4 proteins are drastically reduced following ischemic insults. Additionally, E2F1 association with promoters of pro-apoptotic genes are induced while that of E2F4 is reduced. These changes appear to be important determinants in ischemic neuronal death. Cumulatively, the data supports the activation of the cell cycle machinery as a pathogenic signal contributing to ischemic neuronal death. The development of neuroprotectant strategies for stroke has been hampered in part by its complex pathophysiology. Previous research indicated that flavopiridol, a general CDK-inhibitor, is unable to provide sustained neuroprotection beyond one week following cerebral ischemia. The potential benefit of combining flavopiridol with another neuroprotectant, minocycline, was explored. The data indicate that while this approach provided histological protection 10 weeks after insult, the protected neurons are not functional due to progressive dendritic degeneration. This evidence indicates that targeting cell cycle pathways in stroke while important must be combined with other therapeutic modalities to fully treat stroke-induced damage. Cell Cycle Cell death Neuronal death Stroke Cerebral Ischemia Cdks Cdc25 E2F4 p130 pRb Neuroprotection Inflammation Flavopiridol Minocycline C-Myb B-Myb Excitotoxicity Cdk4 Cyclin D1 Cdk5
9	The Role of Cell Cycle Machinery in Ischemic Neuronal Death Iyirhiaro, Grace O. January 2013 (has links) Ischemic stroke occurs as a result of a lack or severe reduction of blood supply to the brain. Presently therapeutic interventions are limited and there is a need to develop new and efficacious stroke treatments. To this end, a great deal of research effort has been devoted to studying the potential molecular mechanisms involved in ischemic neuronal death. Correlative evidence demonstrated a paradoxical activation of the cell cycle machinery in ischemic neurons. The levels and activity of key cell cycle regulators including cyclin D1, Cdk2 and Cdk4 are upregulated following ischemic insults. However, the functional relevance of these various signals following ischemic injury was unclear. Accordingly, the research described in this thesis address the functional relevance of the activation of the cell cycle machinery in ischemic neuronal death. The data indicate that the inhibition of Cdk4 protects neurons from ischemia-induced delayed death, whereas abrogation of Cdk5 activity prevents excitotoxicity-induced damage in vitro and in vivo. Examination of upstream activators of mitotic-Cdks showed that Cdc25A is a critical mediator of delayed ischemic neuronal death. Investigation of the potential molecular mechanism by which cell cycle regulators induced neuronal death revealed perturbations in the levels and activity of key downstream targets of Cdk4. The retinoblastoma protein family members, pRb and p130 are increasingly phosphorylated following ischemic stresses. Importantly, p130 and E2F4 proteins are drastically reduced following ischemic insults. Additionally, E2F1 association with promoters of pro-apoptotic genes are induced while that of E2F4 is reduced. These changes appear to be important determinants in ischemic neuronal death. Cumulatively, the data supports the activation of the cell cycle machinery as a pathogenic signal contributing to ischemic neuronal death. The development of neuroprotectant strategies for stroke has been hampered in part by its complex pathophysiology. Previous research indicated that flavopiridol, a general CDK-inhibitor, is unable to provide sustained neuroprotection beyond one week following cerebral ischemia. The potential benefit of combining flavopiridol with another neuroprotectant, minocycline, was explored. The data indicate that while this approach provided histological protection 10 weeks after insult, the protected neurons are not functional due to progressive dendritic degeneration. This evidence indicates that targeting cell cycle pathways in stroke while important must be combined with other therapeutic modalities to fully treat stroke-induced damage. Cell Cycle Cell death Neuronal death Stroke Cerebral Ischemia Cdks Cdc25 E2F4 p130 pRb Neuroprotection Inflammation Flavopiridol Minocycline C-Myb B-Myb Excitotoxicity Cdk4 Cyclin D1 Cdk5
10	Study of the regulation and signalling of cdk2-Cyclin o complexes during apoptosis Roset i Huguet, Ramon 04 April 2008 (has links) The aim of this thesis is the characterization of a protein involved in apoptosis. Our group has identified an early step common to different forms of intrinsic apoptosis stimuli. This step requires de novo synthesis of a novel Cyclin, Cyclin O, that upon apoptosis induction in lymphoid cells forms active complexes, primarily with Cdk2. Cyclin O expression precedes glucocorticoid and gamma radiation-induced apoptosis in vivo in mouse thymus and its overexpression induces apoptosis in cultured cells. Knocking down the endogenous expression of Cyclin O by shRNA leads to the inhibition of glucocorticoid and DNA damage-induced apoptosis while leaving CD95 death receptor mediated apoptosis intact. This data demonstrates that apoptosis induction in lymphoid cells is one of the physiological roles of Cyclin O and it does not act by perturbing a normal cellular process such as the cell cycle. In addition we have identified c-Myb a substrate of Cdk2-Cyclin O complexes and we show that c-Myb is downregulated during apoptosis of lymphoid cells. / L'objectiu d'aquesta tesi és la caracterització d'una proteïna involucrada en l'apoptosi. El nostre grup ha identificat un pas primerenc comú en diversos estímuls apoptòtics de la ruta intrínseca. Aquest pas requereix la síntesi de novo d'una nova Ciclina, Ciclina O, que quan s'indueix apoptosi en cèl·lules limfoides forma complexes actius majoritàriament amb Cdk2. L'expressió de la Ciclina O és prèvia a l'apoptosi induïda per glucocorticoids i radiació gamma i la seva sobreexpressió indueix apoptosi en cultius cel·lulars. La baixada dels nivells d'expressió de la Ciclina O endògena amb shRNA provoca una inhibició de l'apoptosi induïda per glucocorticoids o agents que danyen el DNA, mentre que l'apoptosi mediada pel receptor CD95 es manté intacta. Aquests resultats demostren que la inducció d'apoptosi en cèl·lules limfoides és una de les funcions fisiològiques de la Ciclina O i que no es deu a una pertorbació de processos cel·lulars normals com ara el cicle cel·lular. A més a més, hem identificat c-Myb com a substrat dels complexes Cdk2-Ciclina O i demostrem que els nivells de c-Myb baixen durant l'apoptosis de cèl·lules limfoides. Doble híbrid en llevat Dany al DNA Ciclina Apoptosis Yeast two Irbid WEHI7.2 Translation Transcription Thymus Thymocytes siRNA Programmed cell death p53 Mitochondria Mouse Embryonic Fibroblasts Lymphocytes Glucocorticoids Gamma radiation Etoposide DNA-damage Cyclin c-Myb Cdk Cdc25 Caspases Bim Bid BH3-only Bcl-2 Apoptosis Etopòsit Fibroblasts Embrionaris de Ratolí Glucocorticoids Limfòcits Mitocondri Mort cel·lular programada Radiació gamma Timòcits Timus Traducció Transcripció 57 61

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