Molecular Mechanism of X-linked Dyskeratosis Congenita

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DESCRIPTION (provided by applicant): In dyskeratosis congenita (DC) and many other disease states, patient health and longevity are limited by insufficient hematopoietic renewal. Because bone marrow transplantation has not been an effective DC therapy, new strategies must be devised. This application tests the hypothesis that X-linked DC derives from a specific telomerase RNA deficiency and investigates telomerase function in lymphoid cells, with the long-term goal of understanding disease molecular mechanism(s) and developing effective disease therapies. AIM I: Telomerase function will be restored in X-linked DC patient fibroblasts expressing altered dyskerin, in parallel with telomerase activation in family-matched X-linked DC carrier fibroblasts expressing wild-type dyskerin. DC patient cells rescued for telomerase function will be tested for the acquisition of normal proliferative capacity and telomere length maintenance. These findings will be relevant for understanding disease mechanism and also may provide proof-of-principle for telomerase activation as a clinical therapy. AIM II: Aim I telomerase-activated cell lines expressing DC or normal dyskerin will be used to investigate potential DC dyskerin defects in the biogenesis of RNAs other than telomerase. Unlike currently available DC patient cell cultures, cell lines +TERT have robust, uniform growth and will therefore avoid misleading characterization of RNA biogenesis defects secondary to changes in proliferation rate or the percentage of cells undergoing active proliferation. Dyskerin-associated RNA accumulation and dyskerin enzyme function in ribosomal RNA precursor modification and cleavage will be assayed. AIM III: Mechanisms of telomerase function in normal lymphoid cells will be studied using a cell culture system that resolves telomere length -dependent and -independent roles for telomerase in promoting cell growth and survival. A potential third role for telomerase in protection against cell death will also be evaluated. These studies will reveal new roles for telomerase in the cells that are most significantly affected in DC.