Genetic analysis of neurodegeneration in Caenorhabditis elegans

Expansion of a polyglutamine (polyQ) tract in the huntingtin protein causes neuronal degeneration and death in Huntington's disease, but the molecular mechanisms that cause polyQ-mediated cell death are unclear. Previous data suggest that expanded polyQ tracts alter transcription by sequestering glutamine rich transcriptional regulatory proteins thereby perturbing their function. I tested this hypothesis in a Caenorhabditis (C.) elegans model of polyQ toxicity. Roles of C. elegans homologs of CREB, CBP, and HDAC in polyQ-induced neurodegeneration were examined using RNA interference (RNAi) and loss of function alleles. Deletion of CREB ( crh-1) enhanced polyQ toxicity in C. elegans neurons. Loss of one copy of CBP (cbp-1) enhanced polyQ toxicity. Loss of function alleles and RNAi were used to systematically reduce function of each C. elegans HDAC in a neuronal model of polyQ toxicity. Generally, knockdown of individual C. elegans HDACs enhanced Htn-Q150 toxicity, but knockdown of Cehda-3 suppressed polyQ toxicity. Increased expression of Cehda-1 in Htn-Q150 expressing neurons protected those cells from Htn-Q 150 toxicity. Neuronal expression of Cehda-3 restored Htn-Q150 toxicity and suggested that CeHDA-3 protein acts in the neurons to promote degeneration in response to Htn-Q150. Cehda-3 loss of function suppressed degeneration in crh-1;Htn-Q150 animals suggesting that CeHDA-3 and CRH-1 directly oppose each other in regulating transcription of genes involved in polyQ toxicity. Cehda-3 loss of function failed to suppress increased neurodegeneration in Cehda-1;Htn-Q150 animals. These studies confirmed the role of the CREB/CBP pathway in polyQ expansion diseases. Cehda-3 loss of function failed to suppress increased neurodegeneration in Cehda-1/+;Htn-Q150 animals suggesting that CeHDA-1 and Ce HDA-3 have different targets with opposing effects on polyQ toxicity. In addition, differences in CeHDAC modulation of polyglutamine toxicity suggested that CeHDA-3 represses genes that protect against polyQ toxicity while CeHDA-1 repressed genes that induce degeneration in response to polyQ expression. These differences between CeHDACs open new avenues for research and therapeutic drug design.; A second section of this dissertation focuses on a mutation that was isolated from a screen for enhancers of polyglutamine toxicity. The mutation caused neurodegeneration independent of polyglutamine expression and defined a new allele of the sodium channel, degenerin-1 (deg-1). deg-1 is one of many epithelial sodium channels (ENaC) that can be mutated to cause necrotic neuronal death. The deg-1(rt70) mutation encodes an arginine to histidine substitution in a conserved CX7RXCL motif in the extracellular, cysteine-rich region of the protein. Interestingly, the rt70 mutation causes a transient, recessive, degeneration phenotype that is dependent on cold temperatures. The extracellular location, conservation of the motif, and transient nature of the mutation suggest that this arginine residue may be important for regulation or gating of sodium channels.