Thesis (M.S. in Cancer Biology)--Vanderbilt University, Aug. 2007. / Title from title screen. Includes bibliographical references.

Thesis (Ph. D. in Molecular Physiology and Biophysics)--Vanderbilt University, Aug. 2007. / Title from title screen. Includes bibliographical references.

Thesis (Ph. D.)--University of Missouri-Columbia, 2006. / "December 2006" The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Vita. Includes bibliographical references.

Thesis (Ph. D.)--University of Washington, 2007. / Vita. Includes bibliographical references (leaves 55-62).

Thesis (Ph.D.)--Ohio University, November, 2006. / Title from PDF t.p. Includes bibliographical references.

Thesis (Ph. D.)--University of Oregon, 2006. / Typescript. Includes vita and abstract. Includes bibliographical references (leaves 80-86). Also available for download via the World Wide Web; free to University of Oregon users.

Thesis (Ph. D.)--University of Chicago, Dept. of Molecular Genetics and Cell Biology, March 2001. / Includes bibliographical references. Also available on the Internet.

xiii, 135 p. ; ill. (some col.) A print copy of this title is available through the UO Libraries. Search the library catalog for the location and call number. / This thesis explores the origin and evolution of animal germ layers via evolutionary-developmental analyses of the GATA family of transcription factors. GATA factors identified via a conserved dual zinc-finger domain direct early germ layer specification across a wide variety of animals. However, most of these developmental roles are characterized in invertebrate models, whose rapidly evolved sequences make it difficult to reconstruct evolutionary relationships. This study reconstructs the stepwise evolution of metazoan GATA transcription factors, defining homologous developmental roles based upon clear orthology assignments. We identified two GATA transcription factors ( PdGATA123 and PdGATA456 ) from the marine annelid Platynereis dumerilii to aid comparison of protostome and deuterostome GATA factors. Our phylogenetic analyses defined these as protostome orthologs of GATA1/2/3 and GATA4/5/6 vertebrate subfamilies, while the mRNA localization of the Platynereis GATAs showed ectodermal versus endomesodermal germ layer restrictions, similar to their vertebrate orthologs. To define the phylogenetic relationships of more divergent genes in the invertebrate models, we identified GATA homologs from recently sequenced protostome genomes. Molecular phylogenetic analyses, comparisons of intron/exon structure, and conserved synteny confirm all protostome GATA transcription factor genes are members of either the GATA123 or GATA456 class. These data allowed us to identify multiple protostome-specific duplications of GATA456 homologs and reconstruct the origin and relationships of all arthropod GATA genes. To probe GATA transcription factor evolution in deuterostomes, including vertebrates, we identified GATA factors in basal deuterostomes, including the cephalochordate Branchiostoma floridae and the hemichordate Saccoglossus kowalevskii. Phylogenetic analyses of these data independently confirmed that the ancestral deuterostome and chordate--like the bilaterian ancestor--possessed only two GATA transcription factors. This work was facilitated by a bioinformatics platform we are developing to identify gene families from preassembled genomic sequence. We generated anti- PdGATA antibodies to further explore the role of Platynereis GATAs in germ layer formation. We identified multiple presumptive endomesodermal cells in which nuclear localization of PdGATA456 protein first occurs and utilized PdGATA456 protein localization to follow endomesodermal cell populations throughout development. These analyses represent some of the first cellular and molecular analyses of Platynereis germ layer formation. This dissertation includes both my previously published and unpublished co-authored material. / Adviser: Stephan Q. Schneider

Abstract Heart responds to pathological hemodynamic stress by increasing cardiac myocyte size, reprogramming gene expression and enhancing contractile protein synthesis. Neurohumoral factors mediate hypertrophic adaptation either directly via specific receptors or indirectly by increasing blood pressure and cardiac load. The aim of this study was to evaluate the role of angiotensin II (Ang II) in the atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP) gene expression during cardiac overload. Furthermore, the mechanisms of action of Ang II in regulating cardiac gene expression were studied. Hemodynamic stress was produced by Ang II or nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME) administration in conscious rats. Despite hypertension and increased left ventricular ANP and BNP mRNA levels, L-NAME administration for 8 weeks did not induce left ventricular hypertrophy. Ang II type 1 receptor (AT1) antagonism decreased significantly L-NAME-induced hypertension and ventricular ANP gene expression. Ang II-induced cardiac overload produced significant increase in ventricular ANP and BNP mRNA levels at 12 and 72 h, respectively. To study whether the factors synthesized by adrenals modulate the response of Ang II, the effects of adrenalectomy were studied. In Ang II-treated rats, adrenalectomy either abolished or blunted the early activation of ANP and BNP gene expression, respectively. Ang II infusion for 2 weeks increased cardiac mass and blood pressure measured by telemetry, and produced changes in diastolic function detected by echocardiography. By using direct plasmid DNA injections into the rat myocardium, BNP promoter activity was observed to increase at 2 h and remain up-regulated up to 2 weeks of Ang II infusion, except at 12 h. BNP mRNA levels increased at 2 h but decreased to basal levels after 72 h. Mutation of GATA elements of the BNP promoter and DNA binding assays revealed that GATA4 mediates the Ang II-responsiveness of the BNP gene. These results indicate that Ang II plays an important role in regulating natriuretic peptide gene expression during cardiac overload. ANP and BNP gene expression in the rat heart is modulated by the adrenal factors during Ang II-stimulated hemodynamic stress and the AT1 receptor antagonism in NO-deficient hypertension. Moreover, ventricular BNP gene expression in Ang II-induced hypertension in vivo is controlled by posttranscriptional mechanisms and GATA elements.

Abstract Cardiac hypertrophy is an adaptive response of the heart to a variety of mechanical, hemodynamic, neurohumoral, and pathologic stimuli. Prolonged pathophysiological load leads to development of left ventricular hypertrophy and ultimately to heart failure. The natriuretic peptides including the B-type natriuretic peptide (BNP) provide the physiological feedback mechanism to suppress the load signal. The aim of the present study was to evaluate the cis elements within the BNP promoter that mediate the cardiac load responses in vivo, and to study the involvement of paracrine factors, such as endothelin-1 (ET-1) and angiotensin II (Ang II) in activating these transcription factors. In this study, cardiac overload was produced by bilateral nephrectomy, and infusions of arginine8-vasopressin (AVP) or Ang II. In isolated perfused rat heart, the direct wall stretch was achieved by inflating the left ventricular balloon. To identify the cis elements within the BNP promoter that mediate hemodynamic overload response, the approach of DNA injection into the myocardium was used. Mutation or deletion of proximal BNP GATA sites abrogated the response to nephrectomy. AVP-induced acute pressure overload increased left ventricular BNP mRNA and peptide levels. In gel mobility shift assays, pressure overload produced rapid activation of transcription factor GATA4 DNA binding, which was completely inhibited by the ET-1 receptor antagonist bosentan. Both ET-1 and Ang II receptor antagonism inhibited the wall stretch-induced increases in left ventricular GATA4 and AP-1 binding activities in isolated perfused heart preparation. BNP promoter activity and BNP mRNA and peptide levels were regulated distinctly in chronic hemodynamic overload produced by Ang II. In conclusion, GATA4 appears to be necessary and sufficient to confer transcriptional activation of BNP gene during hemodynamic stress in vivo. ET-1 is a signaling molecule mediating the cardiac response to acute pressure overload in vivo. In isolated rat heart, Ang II and ET-1 are required for the stimulation of GATA4 and AP-1 binding activity in response to direct left ventricular wall stretch. Finally, posttranscriptional mechanisms play an important role in the regulation of BNP gene expression in pressure overload produced by Ang II in vivo.