DMPK and myotonic dystrophy : effects of CTG trinucleotide expansion upon DMPK and their contribution to DM pathogenesis

Myotonic dystrophy (DM), an autosomal dominantly inherited form of muscular dystrophy, results from expansion of a CTG trinucleotide repeat within the 3' untranslated region of the gene myotonic dystrophy protein kinase (DMPK). The mutant DMPK allele is transcribed in cultured DM myoblasts, and mutant transcripts are spliced and polyadenylated normally, despite the presence of long CUG tracts. However, mutant DMPK transcripts do not exit nuclei by standard pathways, and they never reach the cytoplasm in differentiated, non-mitotic muscle cells. Instead, they form stable clusters ("foci") that are tightly linked to the nuclear matrix. Nuclear retention of mutant transcripts prevents their translation; consequently DMPK levels are reduced by at least 50% in DM myoblasts. This DMPK deficiency is the probable cause of many of the muscular symptoms of DM; however, it is unlikely to account for all aspects of the disease. DMPK is postulated to play a role in excitation/contraction coupling in muscle tissue, but its precise role there has not been defined, nor has its role been determined within non-muscle tissues. The predominant alternatively spliced forms of DMPK are differentially localized within cultured cells: DMPK+4 is found throughout the cytoplasm, while DMPK-4 is targeted to the periphery of mitochondria. As a result, these splice forms may have distinct regulators and activities, and each form's absence in DM cells may induce a subset of DM's symptoms. Thesis Supervisor: David Evan Housman Novartis Professor of Biology Acknowledgements: Many people have contributed, either directly or indirectly, to this work, and I am truly grateful for their help and support; without it, this thesis would probably never have been completed. I would particularly like to thank and acknowledge: * lain Bason, for encouraging me and believing in me all these years, and for listening to me talk about things he didn't understand. * past and present members of the Housman lab, who provided invaluable suggestions, encouragement, and moral support, as well as for being wonderful people to work with. I am especially indebted to Julian Borrow, Alain Charest, Alex Kazantsev, John Landers, and Elizabeth Preisinger, who gave freely of their time and knowledge, and to Mila McCurrach, who established the experimental system that formed the basis for this work. * Ben Blencowe, who provided a wealth of technical information and insight into fields that were completely foreign to me, as well as help and encouragement in finally getting the writing done. * Ken Moberg, who was a patient and always accessible teacher when I made antibodies; I couldn't have done it without him. * Tony Albino, who gave me my first opportunities in science and showed me the spirit in which it should be done, and who had faith in me long before I had any in myself. * my parents and sister, for being willing listeners and supporters, especially during hard times. * all the long-term members of my thesis committee David Housman, Phil Sharp, and Tyler Jacks for helping me to grow up as a scientist and for making excellent suggestions along the way. * the newest members of my thesis committee Frank Solomon and Alan Beggs who gave their time to make this final rite of passage possible. * my advisor, David Housman, for consistently supporting my research and for providing me with invaluable opportunities for learning to be independent. The help of these people was central to my graduate school experience, and I feel very fortunate to have encountered and interacted with them all.