Thesis (Ph. D.)--University of Hong Kong, 2009. / Includes bibliographical references (leaves 224-250) Also available in print.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1969. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliography.

In U.S. America and much of the Western world, natural is a venerated symbolic placeholder for any number of assumed virtues and ideals. Present conflicts have brought forward questions about what natural (which I argue functions as an ideograph) should mean in contexts that seem to call for a formal, enforceable definition. In this study, I use the vocabulary of Deleuze and Guattari (1987) and the context of bodybuilding to work towards a theory of how ambiguous ideographs become "striated" or “crystallized.” Within this discussion I present instances where natural has been employed as a vehicle to cause harm, and I offer an advisement to rhetorical scholars on how we might approach striated ideographs in the future.

published_or_final_version / Physiology / Doctoral / Doctor of Philosophy

Ciliopathies are a group of disorders that arise from ciliary dysfunction. Meckelin (MKS3 or TMEM67) is a conserved transmembrane protein found at the transition zone of ciliated cells. In humans MKS3 is one of 3 genes linked to the ciliopathy Meckel Syndrome. This disease is characterized by occipital meningioencephalocoele, polycystic kidneys, fibrotic changes to the liver, postnatal polydactyly and situs inversus. Paramecium tetraurelia is a single celled ciliated eukaryote. Its surface is organized of a meshwork of cortical units that run the length of the cell. At the center of the cortical units are either one or two basal bodies. In two basal body units only the posterior basal body is ciliated. From the ciliated basal body, three rootlets project in stereotypical orientations: the post-ciliary rootlet projects posteriorly, the transverse microtubule projects toward the adjacent basal body row and the striated rootlet projects anteriorly. Both the post-ciliary rootlet and transverse microtubule are microtubule-based structures. The striated rootlet is composed of multiple subunits that are predicted to have conserved segmented coiled coil domains known as SF-Assemblin domains. In Picariello at al., 2014, we showed that MKS3 is present in the transition zone of Paramecium tetraurelia and that RNAi for MKS3 leads to global ciliary loss. Additionally, RNAi for MKS3 results in the disorganization of the basal body rows. Within the areas of disorganization, the basal bodies along with their striated rootlets, post-ciliary rootlets and transverse microtubules are rotated away from their expected orientation. Interestingly, the post-ciliary rootlet and transverse microtubule are still attached at the expected angles relative to each other within the areas of disorganization. Initial GST pull-down experiments using the coiled coil domain of MKS3 suggest a potential interaction between MKS3 and the striated rootlet family members KdC1 and KdB2. To test potential interactions between MKS3 and the striated rootlet we identified 27 potential striated rootlet family members in Paramecium. Full-length sequences for 13 of these genes were marked at their N-terminus with a 3x FLAG sequence. Components with a conserved SF-Assemblin domain were distributed uniformly within the striated rootlet. Components lacking the SF-Assemblin domain were found in various cellular locations, but not within the striated rootlet. GST pull-down experiments utilizing the MKS3 C-terminus as bait were performed using cells expressing the FLAG-tagged striated rootlet family members. Unfortunately a clear interaction between MKS3 and the striated rootlet remains elusive. The organized nature of the surface of Paramecium has allowed us to identify a previously unrealized function for MKS3. Our immunofluorescence data suggest that MKS3 functions outside the transition zone to maintain basal body row organization by potentially contributing to a link between the basal body and the striated rootlet. Without the link, the migrating basal bodies are free to rotate and project their rootlets in the wrong directions. Although the nature of the link remains elusive, the identification of disorganized basal body rows upon MKS3 reduction suggests that, in addition to ciliary dysfunction, basal body polarity defects may contribute to the development of MKS.

Gemin5 是運動神經元綜合體(SMN Complex)的其中一個組件，這綜合體的主要功能是控制小型胞核核糖核蛋白(UsnRNPs)的合成。這些小型胞核核糖核蛋白組合成剪接核糖核酸前體(pre-mRNA)的剪接體(Spliceosome)，使核糖核酸分子可以用來翻譯成蛋白質。失去運動神經元綜合體功能引致脊髓肌肉萎縮症。果蠅是其中一個用作研究人類疾病重要的生物。更重要的是，部分組成運動神經元綜合體的組件也存在於果蠅。是次研究是利用遺傳方式在果蠅內研究Gemin5 的同源基因-- rigor mortis (rig) 的作用。果蠅帶有rig 突變基因表現神經肌肉接頭(neuromuscular junction)上的缺陷和異常的運動行為。這表明，rig 的功能可能與神經退化性疾病有關。為了進一步了解rig 的功能途徑(functional pathway)，已進行了一個利用移除突變體的基因過濾實驗，研究鎖定了 12 個染色體部份可能和rig 有基因上的相互作用，進一步研究與rig 有相互作用的基因有助於了解rig 的功能及研究脊髓肌肉萎縮症的發病機制。 / Gemin5 is a component of the Survival of Motor Neuron (SMN) complex, which is a protein complex regulating biogenesis of various Uridine-enriched small nuclear ribonucleoproteins (UsnRNPs). These UsnRNPs form the molecular machinery spliceosome, which mediates pre-messenger RNA splicing, an important mechanism before an mRNA molecule can be used to translate proteins. Loss-of-function of the SMN complex is now known to cause the neurodegenerative disease, Spinal Muscular Atrophy. Drosophila is one of the well-characterized model organisms for studying human diseases. More importantly, components of the SMN complex are also found in Drosophila. Here, I studied the function of rigor mortis (rig), the Gemin5 orthologue, in Drosophila using a genetic approach. Drosophila carrying mutations in the rig gene show defects in the neuromuscular junction (NMJ) and display abnormal motor behavior. This suggests that the function of rig may link to the neurodegenerative disease. To further characterize the function of rig, a genetic screen was carried out. Twelve chromosomal regions encoding possible rig-interacting genes were identified. Further characterization of these rig-interacting genes may help us better understand the function of rig. / Detailed summary in vernacular field only. / Cheng, Yat Pang. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 120-125). / Abstracts also in Chinese. / ABSTRACT --- p.i / ABSTRACT IN CHINESE --- p.ii / ACKNOWLEDGEMENT --- p.iii / LIST OF ABBREVIATIONS --- p.iv / LIST OF FIGURES --- p.v / LIST OF TABLES --- p.vi / TABLE OF CONTENTS --- p.vii / Chapter CHAPTER 1. --- INTRODUCTION / Chapter 1.1 --- Introduction of rigor mortis / Chapter 1.1.1 --- Orthologue of Gemin5 in Drosophila --- p.1 / Chapter 1.1.2 --- Published Phenotypic Analyses of rig Mutants --- p.1 / Chapter 1.2 --- Introduction of Gemin5 / Chapter 1.2.1 --- Introduction of Gemins --- p.4 / Chapter 1.2.2 --- Structural Properties of Gemin5 --- p.4 / Chapter 1.2.3 --- Gemin5-interacting partners --- p.7 / Chapter 1.2.4 --- Gemin5 as a Component of the Survival of Motor Neuron (SMN) Complex --- p.7 / Chapter 1.2.5 --- Function of the SMN Complex and Spinal Muscular Atrophy --- p.11 / Chapter 1.3 --- Drosophila as a Model Organism / Chapter 1.3.1 --- Advantages of Using Drosophila as a Model Organism --- p.11 / Chapter 1.3.2 --- Neuromuscular Junction of Drosophila --- p.15 / Chapter 1.4 --- Aim of the Present Study --- p.19 / Chapter CHAPTER 2. --- MATERIALS AND METHODS / Chapter 2.1 --- Drosophila Culture / Chapter 2.1.1 --- Culture Medium --- p.20 / Chapter 2.1.2 --- Drosophila Stocks and Crosses Maintenance --- p.20 / Chapter 2.1.3 --- Larvae Collection --- p.21 / Chapter 2.1.3.1 --- Reagents --- p.21 / Chapter 2.1.3.2 --- Procedures --- p.21 / Chapter 2.2 --- Cell culture / Chapter 2.2.1 --- Reagents --- p.23 / Chapter 2.2.2 --- Drosophila S2R⁺ Cell Culture --- p.24 / Chapter 2.2.3 --- Establishment of Stable S2R⁺ Cells --- p.24 / Chapter 2.3 --- Genomic Polymerase Chain Reaction (PCR) / Chapter 2.3.1 --- Reagents --- p.25 / Chapter 2.3.2 --- Genomic DNA Extraction from a Single Larva --- p.26 / Chapter 2.3.3 --- Primer Design --- p.26 / Chapter 2.3.4 --- Polymerase Chain Reaction (PCR) --- p.27 / Chapter 2.4 --- Behavioral Assay / Chapter 2.4.1 --- Stable S2R⁺ Cell Staining --- p.29 / Chapter 2.4.1.1 --- Reagents --- p.29 / Chapter 2.4.1.2 --- Procedures --- p.30 / Chapter 2.4.2 --- Larvae Staining --- p.31 / Chapter 2.4.2.1 --- Reagents --- p.31 / Chapter 2.4.2.2 --- Larvae Dissection --- p.32 / Chapter 2.4.2.3 --- Larval Muscle Staining --- p.33 / Chapter 2.4.2.4 --- Larval Neuromuscular Junction Staining --- p.33 / Chapter 2.5 --- Microscopy / Chapter 2.5.1 --- Light Microscopy --- p.34 / Chapter 2.5.1.1 --- Microscopic Observation of Larval Movement --- p.34 / Chapter 2.5.1.2 --- Quantification of Larval Contraction Rate --- p.34 / Chapter 2.5.1.3 --- Quantification of Larval Travelling Distance --- p.34 / Chapter 2.5.2 --- Fluorescence Microscopy --- p.35 / Chapter 2.5.2.1 --- Microscopic Observation of Larval Muscle --- p.35 / Chapter 2.5.2.2 --- Microscopic Observation of Stable S2R⁺ Cells --- p.35 / Chapter 2.5.3 --- Confocal Microscopy --- p.36 / Chapter 2.5.3.1 --- Microscopic Observation of Larval Neuromuscular Junction --- p.36 / Chapter 2.5.3.2 --- Quantification of Larval Neuromuscular Junction --- p.36 / Chapter 2.6 --- Generation of transgenic fly lines expressing rig transgene / Chapter 2.6.1 --- Polymerase Chain Reaction --- p.36 / Chapter 2.6.2 --- Agarose Gel Electrophoresis --- p.38 / Chapter 2.6.2.1 --- Reagents --- p.38 / Chapter 2.6.2.2 --- Procedures --- p.39 / Chapter 2.6.3 --- Restriction Digestion --- p.39 / Chapter 2.6.4 --- Ligation Reaction --- p.39 / Chapter 2.6.5 --- Bacterial Transformation --- p.40 / Chapter 2.6.5.1 --- Reagents --- p.40 / Chapter 2.6.5.2 --- Procedures --- p.40 / Chapter 2.6.6 --- Bacterial Glycerol Stock for Long-term Storage --- p.41 / Chapter 2.7 --- Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and Immunoblotting / Chapter 2.7.1 --- Reagents --- p.41 / Chapter 2.7.2 --- Lysate Preparation of Stable S2R⁺ Cells, Adult Fly Heads and Larvae --- p.44 / Chapter 2.7.2.1 --- Stable S2R+ Cells --- p.44 / Chapter 2.7.2.2 --- Adult Fly Heads --- p.44 / Chapter 2.7.2.3 --- Larvae --- p.45 / Chapter 2.7.3 --- SDS-Polyacrylamide Gel Electrophoresis --- p.45 / Chapter 2.7.4 --- Immunoblotting --- p.45 / Chapter CHAPTER 3. --- PHENOTYPIC CHARACTERIZATION OF RIG MUTANT / Chapter 3.1 --- Introduction --- p.48 / Chapter 3.2 --- Re-balancing of rig Mutant Fly Lines Over the Cy; Tb Compound Balancer for Genotype Identification --- p.48 / Chapter 3.3 --- Verification of Model Genotype --- p.49 / Chapter 3.4 --- rig Mutant Larvae Displayed Abnormal Motor Behavior / Chapter 3.4.1 --- Contraction Rate of rig Mutant Larvae --- p.54 / Chapter 3.4.2 --- Traveling Distance of rig Mutant Larvae --- p.56 / Chapter 3.5 --- rig Mutant Larvae Showed Normal Body Wall Musculature --- p.58 / Chapter 3.6 --- rig Mutant Larvae Displayed Defects in the Neuromuscular Junction / Chapter 3.6.1 --- rig Mutant Larvae Showed Branching Defects --- p.60 / Chapter 3.6.2 --- rig Mutant Larvae Showed Fewer Boutons Number --- p.62 / Chapter 3.7 --- rig Mutant Larvae Showed Normal Active Zone Pattern --- p.64 / Chapter 3.8 --- Discussion --- p.66 / Chapter CHAPTER 4. --- A GENETIC SCREEN TO IDENTIFY GENES THAT INTERACT GENETICALLY WITH RIG / Chapter 4.1 --- Introduction --- p.71 / Chapter 4.2 --- Candidates and Design of the Screen --- p.72 / Chapter 4.3 --- Re-balancing of Deletion Lines Over the Cy; Tb Compound Balancer --- p.75 / Chapter 4.4 --- Identification of Chromosomal Regions That Genetically Interact With rig --- p.75 / Chapter 4.5 --- Identification of NMJ Genes That Genetically Interact With rig --- p.80 / Chapter 4.6 --- Discussion --- p.83 / Chapter CHAPTER 5. --- ATTEMPTS TO INVESTIGATE RIG FUNCTION IN PRE-AND POST-SYNAPTIC REGIONS OF THE NMJ / Chapter 5.1 --- Introduction --- p.89 / Chapter 5.2 --- Transgenic Rescue Experiment by Transgenic Expression of rig in rig Mutant / Chapter 5.2.1 --- Design of the Rescue Experiment --- p.90 / Chapter 5.2.2 --- Construct of pUAST-rig-FLAG --- p.93 / Chapter 5.2.3 --- Construct of the pUAST-myc-rig --- p.98 / Chapter 5.3 --- Tissue Specific Knockdown of rig expression --- p.102 / Chapter 5.4 --- Discussion --- p.105 / Chapter CHAPTER 6. --- ESTABLISHMENT OF AN INDUCIBLE S2R⁺ CELL MODEL FOR RIG OVEREXPRESSION / Chapter 6.1 --- Introduction --- p.108 / Chapter 6.2 --- Detection of Rig Protein in S2R⁺ Cells by Immunoblotting --- p.111 / Chapter 6.3 --- Detection of Rig Protein in S2R⁺ Cells by Immunostaining --- p.111 / Chapter 6.4 --- Detection of RNA in Immunopurified Rig Protein --- p.113 / Chapter 6.5 --- Discussion --- p.115 / Chapter CHAPTER 7. --- GENERAL DISCUSSION --- p.117 / References --- p.120 / Appendices --- p.126

Abstract not available

Thesis (Ph. D.)--York University, 1999. Graduate Programme in Biology. / Typescript. Includes bibliographical references. Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://wwwlib.umi.com/cr/yorku/fullcit?pNQ56221.

Smyd1 is the founder of a gene family whose members contain split SET and MYND Domains. Smyd1 has several SET dependent lysine methyl-transferase substrates, including multiple histone lysines and at least one non-histone protein, skNAC. The MYND domain of Smyd1 is required for protein interactions, such as that with skNAC. Conventional Smyd1 knockouts die at E10 due to cardiac defects, including an enrichment of cardiac jelly, a decrease in trabeculation, and the loss of ventricular septation. dHand, a transcription factor specific for right ventricular development, and Irx4, a ventricle specific gene, are down regulated. I have shown that an approximately one kb stretch of DNA sequence upstream of the muscle specific first exon of Smyd1 is sufficient to drive expression of a reporter in transgenic mice. Cardiac specific expression is mediated by a proximal Mef2 binding site whereas skeletal muscle expression is dependent on E-boxes. I have fully analyzed this stretch of sequence via computational methods and made predictions on other potential regulatory factors. Through the use of Cre mediated conditional knockouts, I have shown that the phenotype of the conventional knockout was not due to the introduction of the Neomycin cassette at the gene locus or due to cell non-autonomous effects on the heart. Smyd1 is not only essential for cardiac septation, but throughout embryonic cardiac development, during embryonic skeletal muscle development, and in adult cardiac tissue. Conditionally deficient Smyd1 embryonic hearts are less affected than conventional Smyd1 knockouts, but are embryonically lethal and show poor trabeculation, cardiac hemorrhaging, and a pericardial edema. I detail that the Nkx2.5-Cre mediated Smyd1 deletion phenocopies the skNAC conventional knockout and that both knockouts have similar changes in the expression levels of several genes. Furthermore, when Smyd1 is conditionally removed from adult cardiac tissue, survival rates are diminished. Surprisingly a skeletal muscle specific CKO of Smyd1 mediated by Myogenin-Cre has resulted in perinatal lethality, with a visible phenotype as early as E15. Evident in the phenotype is a large edema between the epithelium and skeletal muscle, fewer myoblasts, decreased muscle mass, increased degenerating cells, and a potentially defective differentiation process. / text

published_or_final_version / abstract / toc / Physiology / Master / Master of Philosophy