Analysis of Huntingtin-interacting protein 1 in Drosophila melanogaster

Moores, Justin Nathaniel (2007) Analysis of Huntingtin-interacting protein 1 in Drosophila melanogaster. Doctoral (PhD) thesis, Memorial University of Newfoundland.

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Abstract

Huntington's disease (HD) is associated with a wide range of effects including selective neuronal death and altered levels of neurogenesis that are ultimately dependent upon altered activities of Huntingtin (Htt) interacting proteins. These effects are similar in nature to those observed with mutations in the Notch signal transduction pathway. Huntingtin interacting protein 1 (Hip1) shows decreased binding to expanded Htt. Hip1 plays a key role in endocytosis and intracellular transport and activation of the Notch signal requires both. Based on this observation links between Hip1 and Notch-dependent neurogenesis were investigated. In Drosophila two hip1 mRNAs may be naturally produced through the use of alternative splicing of the first exon: full length hip1 with lipid binding ANTH domain and hip1ΔANTH lacking this domain. Directed expression of hip1 decreases while expression of hip1ΔANTH increases microchaetae density in the dorsal notum, a field of sensory bristles on the fruit fly back, suggesting a functional role for Hip1 in neurogenesis. The following studies demonstrate genetic interaction between hip1 and deltex, a key mediator of Notch signaling, with hip1 enhancing and hip1ΔANTH suppressing deltex phenotypes. Reduction of bristle microchaetae density associated with NotchMCD alleles is sensitive to hip1 and hip1ΔANTH. This pathway is shown to be independent of classical Notch control through E(sp1) and tightly controlled by both GSK3β and achaete levels. Building on this the possibility of an analogous pathway in human neuronal development was investigated using the Ntera-2/D1 neuronal precursor cell line along with siRNA and antibody technologies. These studies demonstrate that the Hip1 functions in a similar deltex-dependent, HES1-independent process of neuronal differentiation. Hip1's novel role in neurogenesis provides a functional link between Notch signaling and proteins related to HD advancing the understanding of HD neurogenic phenotypes. -- During the course of these investigations it was noted by others in our laboratory that high levels of Ga/4 in the developing eye result in elevated apoptosis in the eye imaginal disc. Suppression of apoptosis by expression of the caspase inhibitor p35 prevented this phenotype. These studies were extended to analyze Gal4 phenotypes associated with microchaetae density using the pannier-Ga/4 transgenic line. The reduction in microchaetae density associated with pannier-Ga/4 is suppressed by co-expression of p35 but unaffected by expression of GFP. These results show that Gal4 has effects outside the Drosophila eye and that similar mechanisms of disruption are at work at least in the eye and dorsal notum.

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