No description available.

No description available.

No description available.

<p>The Ras proteins, composed of H, N, and KRas, are a family of small GTPases that normally transmit extracellular cues to the cell in a regulated manner. However, Ras is commonly mutated to be inappropriately activated in human cancers, promoting a vast array of tumor phenotypes. Activation of the Raf, PI3K, and RalGEF Ras effector pathways is required to promote Ras-mediated tumorigenesis, leading not only to cell autonomous tumor phenotypes, but also the establishment of a tumor microenvironment. However, following tumor initiation, the requirement upon oncogenic Ras signaling is reduced to activation of PI3K, most likely due to a contribution of the tumor microenvironment. In order to further delineate the requirements for oncogenic Ras signaling pathways during tumorigenesis, I sought to 1) identify PI3K-independent factors necessary for tumor initiation, and 2) determine how PI3K activation maintains tumor growth in the absence of oncogenic Ras. Using cell-based assays and tumorigenesis assays in mice, I have shown that interleukin-6 (IL-6) is secreted upon induction of Ras expression, is required for Ras-mediated tumor initiation, and promotes tumorigenesis in a paracrine manner by fostering angiogenesis. Additionally, I have shown that eNOS, a downstream target of the PI3K pathway, is required for Ras-induced tumor initiation and maintenance, and, moreover, that eNOS-mediated S-nitrosylation and activation of wildtype Ras proteins is required throughout tumorigenesis. Pancreatic cancer is the cancer most highly associated with oncogenic Ras mutations, and I have shown that both IL-6 and eNOS are required for the tumorigenic growth of pancreatic cancer cell lines in mice. I therefore suggest that these proteins, perhaps in combination with other Ras inhibitors, may provide potential anti-cancer targets for oncogenic-Ras driven cancers in the clinic.</p> / Dissertation

<p>Cerebral cavernous malformations (CCM) are vascular anomalies of the central nervous system comprised of grossly-dilated blood-filled capillaries. CCM leisons may occur sporadically or by inheritance of a mutation in one of three genes, <italic>CCM1</italic>, <italic>CCM2</italic>, or <italic>CCM3</italic>. Prior to the identification of the genes involved in pathogenesis, sporadic and inherited cases could be distinguished by lesion burden where sporadic cases exclusively showed single lesions, and patients with inherited disease developed multiple lesions. This observation lead us to hypothesize that CCM lesion genesis may follow a two-hit genetic mechanism. To investigate this hypothesis and determine the molecular mechanism underlying CCM pathogeneis, we used resected human lesion samples to identify biallelic somatic and germline mutations that are specific to the lesion endothelium. Additionaly, we created mouse models in which heterozygosity of <italic>Ccm1</italic> or <italic>Ccm2</italic> in conjunction with deficiency for either the p53 or Msh2 genes, recapitulates the genetic and phenotypic properties of the human condition. In conclusion, we have provided evidence that CCM lesion genesis requires inactivation of both allelic copies for <italic>CCM1</italic>, <italic>CCM2</italic>, or <italic>CCM3</italic>within a subset of vascular endothelium.</p> / Dissertation

<p>Characterizing how genomic sequence interacts with trans-acting regulatory factors to implement a program of gene expression in eukaryotic organisms is critical to our understanding of genome function. One means by which patterns of gene expression are achieved is through the differential packaging of DNA into distinct types of chromatin. While chromatin state exerts a major influence on gene expression, the extent to which cis-acting DNA sequences contribute to the specification of chromatin state remains incompletely understood. To address this, we have used a fission yeast sequence element (L5), known to be sufficient to nucleate heterochromatin, to establish <italic>de novo</italic> heterochromatin domains in the <italic>S. pombe</italic> genome to address the role of DNA sequence in shaping the spatial and temporal propagation of heterochromatin. In this thesis, I describe a major effect of genomic sequences in determining spatial propagation of such <italic>de novo</italic> heterochromatin domains. I demonstrate that the sequence content of a genomic region plays a significant role in shaping its response to encroaching heterochromatin and suggest a role of DNA sequence in specifying chromatin state. Despite the role of DNA sequence in the spatial propagation of chromatin domains, I demonstrate that heterochromatin, once assembled, can propagate by an epigenetic signal, entirely independent of the original nucleating sequences. While the epigenetic signal is sufficient for maintenance and transmission of the heterochromatic state, it is insufficient for reestablishment of heterochromatin following its loss. Thus, these data demonstrate uncoupling of genomic and epigenetic signals necessary for the establishment, spatial propagation, and temporal propagation of chromatin states.</p> / Dissertation

Thesis (Ph. D.)--University OF MICHIGAN.

Some mutations at the gene coding for glucose-6-phosphate dehydrogenase (G6PD) cause a clinical condition of G6PD deficiency, however these alleles have also been shown to confer resistance to malaria based on geographical allele distributions that coincide with malaria, epidemiology, and in vitro studies. A detailed study of nucleotide variability at G6PD can shed light on the signature of selection on the human genome and can also provide insight into the natural history of human malaria. In Appendix A, patterns of nucleotide variability are described in a worldwide panel of humans at G6PD and at a neighboring locus ( L1CAM). Patterns at G6PD do not significantly differ from patterns found at other loci. Nonetheless, significant long-range linkage disequilibrium (LD) is associated with a selected G6PD deficiency allele from Africa (G6PD A-), implying that the allele is young, and that malaria is a recent agent of selection in humans (within the past 10,000 years). In Appendix B, patterns of nucleotide variability are examined in a panel from sub-Saharan Africa at G6PD and at nine loci located around G6PD. LD and reduced nucleotide variability are observed spanning a region of >1 Mb around G6PD, confirming a young age for G6PD A- and implying that the allele has been under strong selection (0.05 &lt; s &lt; 0.20). In Appendix C, a comparable survey of nucleotide variability was conducted in a Eurasian panel with respect to G6PD med, an independently arisen selected G6PD deficiency allele. Patterns of LD provide strong evidence for independent origins of G6PD med in two geographic regions, and that resistance to malaria arose at approximately the same time in Africa and Eurasia. In Appendix D, a panel of Kurdish Jews was studied at G6PD, at neighboring loci, and at 2 unlinked loci to determine if the remarkably high frequency of G6PDmed in this population is attributable primarily to selection or to a severe bottleneck. Patterns of nucleotide variability that are consistent with selection are observed around G6PD, while nucleotide variability at unlinked loci is typical of other non-African populations, suggesting that natural selection is largely responsible for the high frequency of G6PDmed among Kurdish Jews.

Bronchopulmonary dysplasia (BPD), a disease characterized by an arrest in alveolar development, remains one of the leading causes of morbidity in very low birth weight newborns [1, 2]. Insight into the mechanisms and genes involved in alveolarization could be used to better understand the lung pathophysiology of this disease and help with the development of new treatment interventions. Lgl1, is a developmentally regulated, glucocorticoid induced, secreted glycoprotein in the lung [3]. In O2 toxicity animal models of BPD, Lgl1 levels are reduced and return to normal during recovery in air [4]. To determine if a deficiency of Lgl1 is associated with arrested alveolarization and contributes to the pathophysiology of BPD, an Lgl1 knockout mouse model was created. Lgl1 deficiency revealed a complex phenotype including: goblet cell hyperplasia, mucus production, increase expression of interleukins 4 and 13, disruption of lung development, elastin fragmentation and disruption of lung function suggesting that Lgl1 deficiency may contribute to BPD in the neonatal period. / La dysplasie bronchopulmonaire (DBP), une maladie caractérisée par l'arrêt du développement alvéolaire, demeure une des causes principales de morbidité chez les nouveau-nés de faible poids [1, 2]. Plus d'informations sur les mécanismes et les gènes impliqués dans l'alvéolarisation sont nécessaires afin de mieux comprendre la physiopathologie de cette maladie et aider au développement de nouvelles interventions thérapeutiques. Lgl1 est une glycoprotéine sécrétée dans le poumon qui est induite par les glucocorticoïdes et finement régulée lors du développement [3]. Dans les modèles animaux de toxicité d'O2 qui mimique la DBP, les niveaux de Lgl1 sont réduits puis redeviennent normaux lors de la récupération dans l'air [4]. Afin de déterminer si une déficience de Lgl1 est associée à une arrestation de l'alvéolarisation et contribue à la physiopathologie de la DBP, une souris mutante pour le gène Lgl1 a été générée. Un déficit de Lgl1 a révélé un phénotype complexe, incluant notamment : l'hyperplasie des cellules caliciformes, la production de mucus, l'augmentation de l'expression des interleukines 4 et 13, la perturbation du développement pulmonaire, la fragmentation de l'élastine et la perturbation de la fonction mécanique du poumon suggérant que la carence de Lgl1 pourrait contribuer à la DBP dans la période néonatale.

Low plasma high-density lipoprotein cholesterol (HDL-C) is a well-established risk factor for coronary artery disease (CAD). The proprotein convertase subtilisin/kexin 5 (PC5/PCSK5) is known to inactivate endothelial lipase, enzyme critical in modulating HDL-C levels. In this study, we investigated the role of human PCSK5 genetic variants on HDL-C. We examined haplotypes at the PCSK5 locus in 9 multigenerational families with HDL-C<10th percentile and discovered segregation with low HDL-C in one family. We genotyped novel single nucleotide polymorphisms (SNPs) found through sequencing and tagSNPs from the HapMap Project (n=182 total) in 457 individuals with CAD and identified 9 SNPs associated with HDL-C (P<0.05), the strongest being rs11144782 (minor allele frequency 0.164, p=0.002). This SNP decreased HDL-C by 0.076 mmol/L in a gene dosage-effect and was also associated with very low-density lipoprotein (P=0.039), triglycerides (P=0.049) and apolipoprotein B (P=0.022) levels. We conclude that variability at PCSK5 influences HDL-C levels and consequently, CAD risk. / Le rôle protecteur des lipoprotéines de haute densité (HDL) envers les maladies cardiovasculaires est documenté par un grand nombre d'études épidémiologiques. Il est bien établi qu'un niveau bas de HDL-cholestérol (HDL-C) représente un facteur de risque des maladies coronariennes. Des études récentes ont démontré que la proprotéine convertase subtilisin/kexin 5 (PC5/PCSK5) est impliquée dans l'inactivation de la lipase endothéliale, enzyme clé dans la modulation des niveaux plasmatiques de HDL-C. Dans cette étude, nous avons examiné la relation entre les variantes génétiques de PCSK5 et les niveaux du HDL-C chez l'humain. L'analyse des haplotypes PCSK5 dans 9 familles multigénérationnelles caractérisées par un HDL-C<10e centile a montré une ségrégation avec des niveaux faibles de HDL-C chez une famille. D'autre part, l'analyse génotypique des polymorphismes de nucléotide simple (SNP) trouvés par séquençage et des tagSNPs du projet HapMap (n=182 en total) dans 457 personnes coronariennes, a permis l'identification de 9 variantes dans PCSK5 associées à une déficience en HDL-C (P<0.05), la plus forte représentant le rs11144782 (fréquence d'allèle mineure 0.164, P=0.002) et causant une diminution de HDL-C par 0.076 mmol/L dans une relation gène dose-effet. Celui-ci à également été associé à des niveaux faibles de lipoprotéines de très basse densité (P=0.039), des triglycérides (P=0.049) et de l'apolipoproteine B (P=0.022). En conclusion, nos travaux indiquent que la variabilité du gène PCSK5 influence les niveaux du HDL-C et conséquemment, le risque des maladies coronariennes.