Thesis advisor: Marc A.T. Muskavitch / The Notch pathway is an evolutionarily conserved mechanism of intercellular signaling that plays a central role in the development of metazoans. Here I summarize two genetic screens that utilize a rough eye phenotype created by Delta overexpression in the Drosophila eye to identify modulators of Notch pathway signaling activity. Among the many "hits" obtained from both screens, I have mapped to the Not1 gene a single complementation group that exhibits strong genetic interactions with Notch pathway mutants. NOT1 is a component of the CCR4-NOT complex, a global regulator of gene expression that exerts its effects through a variety of mechanisms, including mRNA deadenylation and direct transcriptional repression. I have conducted a series of genetic and molecular experiments in an effort to obtain more insight into the relationship between the CCR4-NOT complex and the Notch pathway. Both Not1 EMS mutations and RNAi-mediated knockdown of NOT1 expression produce phenotypes that mimic those of Notch loss-of-function pathway mutants. Knockdown of NOT1 in the developing bristle organ disrupts Notch-mediated inhibition of neuronal specification, resulting in supernumerary neurons and aberrant sheath cell specification. Knockdown of NOT1 within the developing wing margin disrupts expression of the Notch target genes Cut and Wingless, as well as the Notch ligand Delta. Phenotypic rescue experiments imply that Not1 functions downstream of Notch signal activation and acts directly on Notch target gene expression. These results suggest that NOT1 is required for Notch signal transmission in certain developmental contexts and implicate the CCR4-NOT complex as a positive regulator of the Notch pathway. / Thesis (PhD) — Boston College, 2009. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.

Peptídeos sinais determinam o crescimento, desenvolvimento e defesa das plantas. RALF (Rapid Alkalinization Factor) é um peptídeo de sinalização ubíquo no reino vegetal e que está envolvido com a expansão celular. Os peptídeos RALF em arabidopsis estão organizados em uma família multigênica de 37 membros, alguns com expressão tecido-específica, outros expressos em toda a planta. Os mecanismos envolvidos na expansão celular são regulados por vários hormônios, entre os quais as citocininas. A relação existente entre os peptídeos RALF e os demais hormônios é pouco conhecida e um melhor entendimento dessa relação poderá auxiliar na modulação dos processos de crescimento e desenvolvimento vegetal por engenharia genética. O objetivo desse trabalho foi estudar a relação entre o peptídeo AtRALF e as citocininas, principalmente no que diz respeito aos efeitos de ambos no crescimento e desenvolvimento das raízes. Para isso, selecionou-se as isoformas AtRALF1, AtRALF19 e AtRALF34 que apresentam diferentes padrões de expressão. Os resultados sugerem que AtRALF19 e AtRALF34, ambas expressas em toda a planta, contribuem mais com a transdução de sinal da citocinina do que a isoforma AtRALF1, com padrão de expressão específico de raízes. Os peptídeos AtRALF19 e 34 reprimem parcialmente a expressão dos genes reguladores de resposta, ARRs tipo-A, que são reguladores negativos da via de sinalização de citocinina. / Peptides signals influence the growth, development and plant defense. RALF (Rapid Alkalinization Factor) is a ubiquitous signaling peptide in the plant kingdom and is involved in cell expansion. The RALF peptides in arabidopsis are organized in a multigene family of 37 members, some with tissue-specific expression, others expressed throughout the plant. The mechanisms involved in cellular growth are regulated by various hormones, including cytokinins. The relationship between RALF peptides and other hormones is poorly understood and a better understanding of this relationship assists in modulating the processes of plant growth and development. The aim of this work was to study the relationship of AtRALF peptide with cytokinins, especially with regard to the effects of both in the growth and development of roots. For this, we selected the AtRALF1, AtRALF19 and AtRALF34 isoforms that have different expression patterns. The results suggest that AtRALF19 and AtRALF34, both expressed throughout the plant, contribute more to cytokinin signal transduction than isoform AtRALF1, with specific expression pattern in roots. The AtRALF19 and 34 repressed the expression of type-A Arabidopsis Response Regulators (ARRs), whose products act as negative regulators of cytokinin signaling.

The interplexiform cells(IP cells) are the most recently discovered neurons in the retina and their function is to provide centrifugal feedback in retina. The anatomical structure of the IP cells has been well studied, but the function of these neurons is largely unknown. I systematically studied the excitatory and inhibitory inputs from IP cells in salamander retina. I found that L-EPSCs in IP cells are mediated by AMPA and NMDA receptors; in addition, L-IPSCs are mediated by glycine receptors and GABAC receptors. In response to light, IP cells reaction potentials transiently at the onset and onset of light stimulation. The major neural transmitter of IP cells in salamander retina is glycine. We also studied the distribution and function of glycine transporters. Our result indicates that GlyT1- and GlyT2-like transporters were present in Muller cells and neurons. The glycine feedback at outer plexiform layer (OPL) has effects on both the bipolar cell dendrites and rod photoreceptor terminals. At bipolar cell dendrites, glycine selectively depolarizes rod-dominant On-bipolar cells, and hyperpolarizes Off- bipolar cells. At rod photoreceptor terminals, 10 M glycine activates voltage-gated Ca2+ channels. These effects facilitated glutamate vesicle release in photoreceptors. It increases the sEPSC in OFF bipolar cells. The combined effect of glycine at rod terminals and bipolar cell dendrites leads to enhanced dim light signal transduction in the rod photoreceptor to ganglion cell pathway. This study provides a model that displays the function of centrifugal feedback through IP cells in the retina. / by Zheng Jiang. / Thesis (Ph.D.)--Florida Atlantic University, 2009. / Includes bibliography. / Electronic reproduction. Boca Raton, Fla., 2009. Mode of access: World Wide Web.

We have studies oxidative damage of RNA, a major type of cellular macromolecules. RNA is a primary target of reactive oxygen species (ROS). Under oxidative stress, most nucleic acid damages in Escherichia coli (E.coli) are present in RNA as shown by high levels of 8-oxo-G, an oxidized form of guanine. Increased RNA oxidation is closely correlated to cell death under oxidative stress. Surprisingly, neither RNA structure nor association with proteins protects RNA from oxidation... Our results demonstrate a major role for RNA degradation in controlling oxidized RNA. We have identified activities that may work in specific pathways for selectively degrading damaged RNA. These activities may play pivotal rold in controlling oxidized RNA and protecting cells under oxidative stress. / by Min Liu. / Thesis (Ph.D.)--Florida Atlantic University, 2012. / Includes bibliography. / Mode of access: World Wide Web. / System requirements: Adobe Reader.

We employed three genotypes of GAD 65, wildtype (GAD 65 +/+), heterozygous (GAD 65 +/-) and knockout (GAD 65 -/-) to investigate the role of GAD 65 in survival of pancreatic islets. We analyzed the mRNA expression of pro-survival proteins including Bcl2 and Bax in pancreas of wildtype, heterozygous and knockout using Reverse Transcriptase Polymerase Chain Reaction (RTPCR). The level of expression of Bcl2 mRNA was down regulated in knockout mice pancreas and Bax to Bcl2 ratio was found higher in knockout mice pancreas suggesting higher cell death rate. However, further studies are required to recognize and understand the specific connections between apoptotic pathways and GAD 65 in pancreatic islets. / by Neeta Kumari. / Thesis (M.S.)--Florida Atlantic University, 2012. / Includes bibliography. / Electronic reproduction. Boca Raton, Fla., 2012. Mode of access: World Wide Web.

by Wai-on Fu. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1993. / Includes bibliographical references (leaves 88-95). / Chapter Chapter I --- Introduction --- p.1 / Chapter I .1 --- Structure and functions of the epididymis --- p.1 / Chapter I.2 --- Cellular mechanisms for electrolyte secretion --- p.3 / Chapter I.3 --- Second-messenger modulation of chloride secretion in epididymis --- p.5 / Chapter I.3.1 --- Cyclic AMP pathway --- p.5 / Chapter I.3.2 --- Ca2+ as second messenger --- p.6 / Chapter I.4 --- Pathophysiology of electrolyte transport in the epididymis --- p.6 / Chapter I.5 --- Objectives of the study --- p.9 / Chapter Chapter II. --- Materials and Methods --- p.10 / Chapter II. 1 --- Materials --- p.10 / Chapter II. 1.1 --- Culture media and enzyme --- p.10 / Chapter II. 1.2. --- Drugs --- p.10 / Chapter II. 1.3 --- Chemicals --- p.11 / Chapter II. 1.4. --- Animals and human tissue --- p.11 / Chapter II.2 --- Preparation of solutions --- p.12 / Chapter II. 3. --- Preparation of cultured cells --- p.12 / Chapter II.3 .1 --- Culture of rat epididymal epithelial cells --- p.12 / Chapter II.3.2 --- Cultured of human epididymal epithelial cells --- p.16 / Chapter II.4. --- Patch-Clamp technique --- p.21 / Chapter II.4.1 --- Electrode --- p.23 / Chapter II.4.2 --- Pulling of electrode --- p.23 / Chapter II.4.3 --- Coating of electrode --- p.23 / Chapter II.4.4 --- Polishing of the electrode --- p.24 / Chapter II.4.5 --- Filling of the electrodes --- p.26 / Chapter II.4.6 --- Mounting of Electrode to the Headstage Pipette Holder --- p.26 / Chapter II.4.7 --- Electrical Isolation --- p.28 / Chapter II.4.8 --- Vibration Isolation --- p.28 / Chapter II.4.9 --- "Formation of ""Giga-seal"" and Whole-cell configuration" --- p.28 / Chapter II.4.10 --- Correction for liquid junction potential --- p.30 / Chapter II.4.11 --- "Data Acquisition and Analyses," --- p.32 / Chapter II.4.12 --- Statistics --- p.34 / Chapter Chapter III. --- Results --- p.35 / Anion Secretion in human epididymal cells --- p.35 / Chapter III. 1 --- Whole-cell current in human epididymal cells --- p.35 / Chapter III. 2 --- Effect of adrenergic receptor blockers on whole-cell current --- p.38 / Chapter III.3 --- Effect of inhibitors of the cyclic AMP pathway --- p.42 / Chapter III.4 --- Effect of altering intracellular calcium concentration --- p.42 / Anion secretion in rat epididymal cells --- p.46 / Chapter III. 5 --- Effect of cAMP on whole cell C1- current in rat epididymis --- p.46 / Chapter III.6 --- Effect of ionomycin on whole cell C1- current --- p.53 / Chapter III.7 --- Differences between cAMP- and ionophore- dependent C1- current --- p.57 / Chapter III. 8 --- The swelling-induced whole-cell currents --- p.63 / Chapter III.9 --- The swelling-induced current was mainly C1- selective --- p.65 / Chapter III. 10 --- Anion selectivity of the swelling-induced C1- current --- p.68 / Chapter III. 11 --- Inhibition of the swelling-induced C1- conductance by anion channel blockers --- p.68 / Chapter III. 12 --- The role of Ca2+ and cAMP in swelling-induced C1- conductance --- p.74 / Chapter Chapter IV. --- Discussion --- p.79 / Signal transduction mechanism of adrenaline stimulated C1- current in human epididymal cell --- p.79 / Anion secretion in rat epididymal cell The role of Ca2+ and cAMP --- p.83 / Chapter Chapter V. --- Reference --- p.88

Ling, Yick Hin. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 72-77). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts also in Chinese.

Cheung, Hei Nga. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 63-72). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts also in Chinese.

by Wen-Liang Zhou. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1996. / Includes bibliographical references (p. 125-144). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web.

Notch signaling plays an important role in developmental and pathological angiogenesis. Notch ligands, Dll4 and Jag1, have been implicated in tumor angiogenesis. Inhibition of Dll4-mediated Notch signaling results in hypersprouting of non-functional vasculature in tumors. We have constructed and analyzed pan-Notch ligand inhibitors, Notch1 decoys 1-24 and 1-36, which are based on the extracellular EGF-like repeats of Notch1. Both Notch1 decoys block angiogenesis in in vitro endothelial cell-based assays and in the mouse retina. We also show that they similarly inhibit Dll4- and Jag1-induced Notch signaling in vitro and result in a significant decrease in tumor growth and tumor vasculature in mouse and human tumor xenograft models. Interestingly, truncated Notch1 decoy variants, Notch1 decoys 1-13 and 10-24, act as ligand-specific Notch inhibitors. Notch1 decoy 1-13 is Dll4-specific whereas Notch1 decoy 10-24 is Jag1-specific. Ligand-specific Notch1 decoys effectively reduce tumor growth in tumor xenograft models in the mouse, including Mm5MT-FGF4, KP1-VEGF, LLC, and B16-F10. Notch1 decoy 1-13 has been demonstrated to increase tumor vasculature by increasing endothelial sprouting and number of tip cells. However, similar to the previously reported effects of Dll4 blockade, the tumor vessels are poorly perfused and hardly functional. On the other hand, Jag1-specific Notch1 decoy 10-24 significantly reduces tumor vessel density and disrupting endothelial-pericyte interactions, causing the impaired vascular structure and attenuated vascular perfusion. In addition, Notch1 decoys 1-13, 10-24, and 1-24 show an anti-metastatic potential in causing a delay of lung metastasis in the B16-F10 tumor model. Unlike gamma-secretase inhibitors and Dll4-blocking agents, Notch1 decoys do not cause GI-associated toxicity or vascular neoplasms. Therefore, our Notch1 decoys may represent a novel alternative and may hold future promise for Notch-targeted cancer therapy.