Consequences of gain-of-function RPA1 mutations on telomere function and hematopoiesis

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Project Summary Inherited mutations in genes governing telomere maintenance cause Telomere biology disorders (TBD), a group of diseases with a progressive course and poor prognosis characterized by pulmonary fibrosis, liver cirrhosis, and bone marrow failure (BMF). Understanding the molecular mechanisms governing telomere preservation and identifying TBD associated genes are established gaps. We recently published novel germline heterozygous missense variants in RPA1 in four unrelated probands presenting with TBD. We also demonstrated that RPA1 mutants have elevated affinity to random and telomere single strand DNA, which causes a unique gain-of- function (GoF) effect, defective hematopoiesis due to short telomeres in RPA1 mutant iPSC cells and extensive somatic rescue mosaicism. The molecular mechanism through which RPA1 GoF mutations cause telomere shortening, and TBD is unknown. I hypothesize that RPA1 mutations cause TBD by outcompeting essential telomere maintenance proteins for binding to ssDNA, thereby limiting stem cell replicative potential. The proposed studies will investigate these questions through two specific aims: 1) determine the molecular mechanism by which GoF RPA1E240K causes telomere dysfunction and 2) determine the effect of RPA1E240K on telomere maintenance and hematopoiesis in vivo. A Mentored Clinical Scientist Research Career Development Award (K08) will provide the candidate with the amount of protected time needed to achieve her career goal of independence as a physician scientist and focus on improving the clinical outcomes of patients with BMF and DNA repair disorders through understanding the mechanisms of their disease pathogenesis. A strong career development plan including experienced and successful faculty advisor and co-advisor committee, clear plans for progress assessments, and attendance at numerous courses, lectures, and workshops to increase proficiency in technical and management skills will accompany these research goals to ensure success as an independent physician scientist. This award will be completed at St. Jude Children’s Research Hospital (SJCRH), one of the world’s leading academic institutions focused on the research and treatment of pediatric catastrophic diseases, making it an exemplary location for an early career physician scientist to develop his career. In addition to the strong institutional support, St. Jude offers unmatched research support with facilities including the Flow Cytometry and Cell Sorting Shared Resource, Center for Advanced Genome Engineering (CAGE), Light Microscopy Division of Cell and Tissue Imaging Center, Animal Resource Center, Cytogenetics Core, and Center for Proteomics.