Proteins in the nuclear envelope (NE) have been shown to be important regulators of cell function and development, and disruption of these proteins leads to a range of diseases termed laminopathies. The Torsin family of AAA+ ATPases have recently been proposed to regulate NE proteins. Mutation in the human gene that encodes TorsinA, DYT1, leads to the neuromuscular disease early onset dystonia. Mammalian cells with out TorsinA function have NE structural defects, and the outer nuclear membrane KASH domain protein Nesprin-3 can interact with TorsinA. However, how TorsinA functions at the NE, and how this function relates to the disease phenotype is not known. The long term goal of this project is to understand the cell biology of Torsin function at the NE, and how Torsins regulate later development. In this proposal, I will investigate Torsin function at the NE using C. elegans as a model. Our laboratory previously identified ooc-5, a C. elegans homolog of TorsinA. Mutations in ooc-5 lead to smaller oocytes and polarity and spindle positioning defects in early embryos. Interestingly, it has been shown that depletion of nucleoporins, components of nuclear pore complexes, in C. elegans leads to an ooc-5 like phenotype, suggesting a link between Torsins and nuclear pores in the worm. Furthermore, preliminary studies have revealed defects in the localization of multiple nuclear envelope proteins in ooc-5 mutants. Using GFP reporter strains and RNAi-mediated gene depletion, I propose to determine which NE protein defects are primary, and which are secondary downstream defects. I will also determine if nuclear envelope barrier and transport functions are disrupted in ooc-5 mutants by using nuclear-localized GFP markers and dextran dye injections. This will be the first test of the functional consequences of nuclear envelope defects in a Torsin mutant model. Finally, I will test which defects at the NE cause later polarity phenotypes, providing a link between the cell biology of Torsin function and the regulation of development. Results from this study will be applicable to both human Torsin function and mechanisms underlying early onset dystonia, and also to the study of embryonic polarity establishment in C. elegans.
Recent studies have found that the membrane surrounding the nucleus, called the nuclear envelope, is an important regulator of normal development and disease. TorsinA protein is encoded by the gene DYT1, and has unknown functions at the nuclear envelope. Mutations in DYT1 lead to the neuromuscular disease early onset dystonia. Using the nematode C. elegans as a model system, this study will investigate how Torsin functions at the nuclear envelope in development and disease.