Polyglutamine (polyQ) diseases describe a group of late-onset progressive neurodegenerative disorders which are caused by the CAG triplet repeat expansion in the coding region of disease genes. Such expansions result in expanded polyQ tracts in the disease proteins which confer neurotoxicity. To date, nine such diseases are reported including Huntington's disease and several types of spinocerebellar ataxias. Misfolding of polyQ proteins and formation of intracellular SDS-insoluble protein aggregates are closely associated with the toxicity of these diseases. In particular, impairment of the ubiquitin-proteasome system (UPS) which is responsible for protein degradation has been observed in polyQ diseases. Recently, ubiquitin ligases, which govern substrate specificity of the UPS, have gained huge attention in polyQ disease pathogenesis studies. In humans, cullin (Cul) proteins, including Cul1, 2, 3, 4 & 5, are integral components of a group of ubiquitin ligases called cullin- RING ubiquitin ligases (CRLs). Each CRL displays distinct substrate specificity through specific substrate receptors. Cullin proteins are evolutionarily conserved and Cul orthologues are found in the Drosophila genome. In the present study, it was found that individual Culs displayed distinct effects on polyQ pathogenesis in Drosophila polyQ models. Particularly, it was found that Cul1 modulated polyQ-induced toxic phenotype. This modification was accompanied with an alteration in the ubiquitination level and SDS-solubility properties of expanded polyQ protein. Through genetic interaction studies and biochemical analyses, it is suggested that Cul1-based CRL specifically targets SDS-insoluble species of expanded polyQ protein for ubiquitination via selective recognition by CG2010 substrate receptor. On the other hand, it was found that expanded polyQ protein induced accumulation of CRL substrates in cells. Current data support a hypothesis that polyQ protein would impair the ubiquitin ligase activity of CRLs upon expansion of the polyQ domain, through interfering with neddylation of cullin and other uncharacterized mechanisms. Taken together, the present study identifies Cul1-CRL as a novel E3 ligase that modifies polyQ toxicity through modulating ubiquitination of expanded polyQ protein, and demonstrates a pathological mechanism by expanded polyQ protein through impairing CRL activity. These findings would lead to a better understanding of polyQ pathogenesis and give insights on developing treatments against polyQ diseases. / Wong, Kam Yan. / Adviser: Ho-Yin Edwin Chan. / Source: Dissertation Abstracts International, Volume: 73-01, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 260-273). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

A confocal scanning laser ophthalmoscope (CSLO) was used to image the axonal and dendritic aborizations of RGCs in the Thy-1 YFP mice. With quantitative analysis of cell body area, axon diameter, dendritic field, number of terminal branches, total dendritic branch length, branching complexity, symmetry and distance from the optic disc, the morphologies of RGCs and the patterns of axonal and dendritic degeneration were analyzed. After optic nerve crush, RGC damage was observed prospectively to begin with progressive dendritic shrinkage, followed by loss of the axon and the cell body. Similar pattern of RGC degeneration was observed after 90 minutes of retinal ischemia although no morphological changes were detected when the duration of ischemia was shortened to 30 minutes. The rate of dendritic shrinkage was variable and estimated on average 2.0% per day and 11.7% per day with linear mixed modeling, after optic nerve crush and retinal ischemic injury, respectively. RGCs with a larger dendritic field had a slower rate of dendritic shrinkage. / In summary, we demonstrated that dendritic shrinkage could be evident even before axonal degeneration after optic nerve crush and retinal ischemic injury. We have established a methodology for in vivo and direct visualization of RGCs and retinal microglia, which could provide reliable and early markers for neuronal damage. Measuring the rate of dendritic shrinkage and tracking the longitudinal activation of microglia would provide new paradigms to study the mechanism of neurodegenerative diseases and offer new insights in testing novel therapies for neuroprotection. / Progressive neuronal cell death and microglial activation are the key pathological features in most neurodegenerative diseases. While investigating the longitudinal profiles of neuronal degeneration and microglial activation is pertinent to understanding disease mechanism and developing treatment, analyzing progressive changes has been obfuscated by the lack of a non-invasive approach that allows long term, serial monitoring of individual neuronal and microglial cells. Because of the clear optical media in the eye, direct visualization of the retinal ganglion cells (RGCs) and microglia is possible with high resolution in vivo imaging technique. In this study, we developed experimental models to visualize and characterize the cellular morphology of RGCs and retinal microglia in vivo in the Thy-1 YFP and the CX3CR1 +/GFP transgenic mice, described the patterns of axonal and dendritic shrinkage of RGCs, discerned the dynamic profile of microglial activation and investigated the relationship between RGC survival and microglial activation after optic nerve crush and retinal ischemic injury induced by acute elevation of intraocular pressure. / The longitudinal profile of microglial activation was investigated by imaging the CX3CR1GFP/+ transgenic mice with the CSLO. Activation of retinal microglia was characterized with an increase in cell number reaching a peak at a week after optic nerve crush and retinal ischemic injury, which was followed by a gradual decline falling near to the baseline at the 4 th week. The activation of retinal microglia was proportional to the severity of injury. The number of RGCs survival at 4 weeks post-injury was significantly associated with the number of activated retinal microglia. / Li, Zhiwei. / Adviser: Leung Kai Shun. / Source: Dissertation Abstracts International, Volume: 73-02, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 50-66). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.