Biochemical and Functional Characterizations of Dyskerin Complexes

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Research of RNA-containing enzyme complexes has gained much momentum in the past decade. Despite this upswing, details about the assembly, and the precise cellular functions, of many RNA-containing enzymes have remained poorly defined. Understanding these cellular processes will undoubtedly reveal biological pathways essential to human health. Dyskerin is a housekeeping enzyme that participates in two important cellular processes involving enzymatic RNAs: the repair of chromosome ends, and the regulation of protein synthesis. Heritable mutations in the dyskerin locus lead to multiple human diseases with overlapping phenotypes. At present, the specific mechanisms by which these genetic changes impact dyskerin's multiple functions in RNA-containing enzyme complexes, and to what extent, are not known.******My current and proposed projects include biochemical and cell-biology studies of dyskerin functions, and how its activity impacts other ribonucleoproteins. Our first project will focus on dyskerin's regulatory roles in chromosome-end maintenance by the RNA-enzyme telomerase. We will determine the mechanism by which changes in dyskerin sequence leads to a loss in telomerase-RNA stability, by measuring the rate of enzymatic complex formation. Our second project will probe the structure-function relationship of dyskerin's RNA modification functions, implicated in the regulation of cellular protein synthesis capacity. We will study how the loss of individual site-specific pseudouridine residues impacts both normal, and stress-induced, protein synthesis pathways mediated by ribosomes, the protein-synthesis machinery. We will also measure the physiological consequences (cell viability) of sub-optimal ribosomal RNA modification. Our data will identify the precise molecular characteristics of dyskerin-containing complexes responsible for its multiple cellular activities, and map the mechanisms by which changes can affect these cellular roles. ******Studies of RNA-containing enzyme complexes and their functions have revealed many exceptions to the central dogma of the DNA-protein relationship. Our study will contribute to this by deducing how dyskerin-RNA complexes modulate the functions of biological pathways and impact normal physiology. In addition, this study will provide information to predict genotype-phenotype relationships for inheritance of variant dyskerin isoforms, and will contribute to the modeling of ribosomal function regulation.**