The broad objective of this application is to study the molecular basis of cardiac contractility using zebrafish, a model vertebrate system. The proposed research will be conducted within the sound scientific environment at the University of California, San Francisco (UCSF). Dr. Sehnert, a fellow in Pediatric Cardiology at UCSF, has dedicated the past two years to full-time research in the sponsor's laboratory as a fellow of the Pediatric Scientist Development Program (PSDP). She is currently funded by the PSDP for a third year in the laboratory, and her immediate goals are to bring her preliminary work to publication. The candidate's long-term career goals are to establish an investigative research career studying congenital and genetically acquired heart disease. The career development plan incorporated in this application includes mentorship by a scientific sponsor and advisory committee; participation in relevant course work, seminars, and scientific retreats both locally and nationally; and an academic appointment providing protected research time to ensure her success in achieving her career goals. The proposed research plan specifically addresses the affect of cardiac troponin T (cTnT) mutations on cardiac contractility. cTnT is a crucial thin filament protein of the contractile apparatus, or sarcomere. It is clinically relevant to the human genetic disorder, familial hypertrophic cardiomyopathy (FHC). 15 percent of FHC cases result from cTnT mutations, which for unknown reasons predispose patients to sudden death despite minimally detectable hypertrophy. We hypothesize that a genetic mutation affecting cTnT expression results in the non-contractile heart defect observed in zebrafish silent heart mutant embryos. Furthermore, we believe that the visual transparency and genetic manipulability of zebrafish embryos offer distinct advantages for studying the affect of induced cTnT mutations on heart function. Therefore, we propose the following specific aims: 1) To isolate and characterize the silent heart gene in zebrafish using genetic techniques, 2) To rescue the silent heart phenotype by replacing the wild-type gene in mutant embryos, and 3) To generate transgenic zebrafish for combined in vivo and in vitro evaluation of mutant cTnT expression on heart function. Accomplishing these will advance our understanding of the regulation and in vivo role of cTnT in sarcomere assembly and cardiac contractility and will offer insight into the molecular mechanisms leading from cTnT mutation to disease.
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| Sehnert, Amy J (2006) Use of denaturing high-performance liquid chromatography to detect mutations in pediatric cardiomyopathies. Methods Mol Med 126:257-70 |
| Sehnert, Amy J; Stainier, Didier Y R (2002) A window to the heart: can zebrafish mutants help us understand heart disease in humans? Trends Genet 18:491-4 |
