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Dr. Gillespie’s laboratory is broadly focused on the mechanism of mechanotransduction by sensory hair cells of the inner ear. Although the lab uses a multidisciplinary approach, including molecular biology, imaging, genetics, electrophysiology, and other approaches, they specialize in the application of biochemical methods to characterizing hair-cell transduction. Dr. Gillespie has a broad background in biochemistry, with a particular emphasis on the sensitive techniques required for inner-ear biochemistry, given the small amount of protein present in each ear. As a graduate student, he was trained in the laboratory of Joe Beavo, an enzymologist interested in phosphodiesterases. As a postdoc, he worked with Jim Hudspeth and developed the hair-bundle purification that forms the basis for mass spectrometry experiments they use widely in the lab. Dr. Gillespie’s lab has exploited this technique for over twenty years to carry out hair-bundle biochemistry experiments.
Seminar Abstract:
The mechanotransduction apparatus of vertebrate hair cells, the sensory cells of the inner ear, remains incompletely characterized. We are taking a biochemical approach to the description of molecules key for hair-cell transduction. Using the "twist-off" hair-bundle preparation pioneered twenty years ago, we isolate bundles that are >95% pure. Employing sensitive mass-spectrometry approaches with differential quantitation in bundles and whole epithelium, we identified >400 proteins in the bundle preparation with high confidence. Of these, ~75 are highly enriched in hair bundles, and ~175 are at roughly similar concentrations in bundles and epithelium. Of the highly enriched bundle proteins, many are encoded by previously identified "deafness genes," genes that lead to deafness when mutated. Because estimates suggest that only a fraction of deafness genes have been identified, many more must be present in the list of enriched hair-bundle proteins. We will use this strategy of complete description of the bundle proteome, as well as targeted immunoprecipitation approaches, to describe molecules essential for mechanotransduction.
The hair bundle's protein constellation [electronic resource] / Peter Gillespie.
Series:
NIH neuroscience seminar series
Author:
Gillespie, Peter. National Institutes of Health (U.S.)
Publisher:
[Bethesda, Md. : National Institutes of Health, 2010]
Other Title(s):
NIH neuroscience seminar series
Abstract:
(CIT): Dr. Gillespie's laboratory is broadly focused on the mechanism of mechanotransduction by sensory hair cells of the inner ear. Although the lab uses a multidisciplinary approach, including molecular biology, imaging, genetics, electrophysiology, and other approaches, they specialize in the application of biochemical methods to characterizing hair-cell transduction. Dr. Gillespie has a broad background in biochemistry, with a particular emphasis on the sensitive techniques required for inner-ear biochemistry, given the small amount of protein present in each ear. As a graduate student, he was trained in the laboratory of Joe Beavo, an enzymologist interested in phosphodiesterases. As a postdoc, he worked with Jim Hudspeth and developed the hair-bundle purification that forms the basis for mass spectrometry experiments they use widely in the lab. Dr. Gillespie"s lab has exploited this technique for over twenty years to carry out hair-bundle biochemistry experiments. Seminar Abstract: The mechanotransduction apparatus of vertebrate hair cells, the sensory cells of the inner ear, remains incompletely characterized. We are taking a biochemical approach to the description of molecules key for hair-cell transduction. Using the "twist-off" hair-bundle preparation pioneered twenty years ago, we isolate bundles that are >95% pure. Employing sensitive mass-spectrometry approaches with differential quantitation in bundles and whole epithelium, we identified >400 proteins in the bundle preparation with high confidence. Of these, ~75 are highly enriched in hair bundles, and ~175 are at roughly similar concentrations in bundles and epithelium. Of the highly enriched bundle proteins, many are encoded by previously identified "deafness genes," genes that lead to deafness when mutated. Because estimates suggest that only a fraction of deafness genes have been identified, many more must be present in the list of enriched hair-bundle proteins. We will use this strategy of complete description of the bundle proteome, as well as targeted immunoprecipitation approaches, to describe molecules essential for mechanotransduction.
