title: ATP Consumption of Eukaryotic Flagella Measured at a Single-Cell Level.
abstract: The motility of cilia and flagella is driven by thousands of dynein motors that hydrolyze adenosine triphosphate (ATP). Despite decades of genetic, biochemical, structural, and biophysical studies, some aspects of ciliary motility remain elusive, such as the regulation of beating patterns and the energetic efficiency of these nanomachines. In this study, we introduce an experimental method to measure ATP consumption of actively beating axonemes on a single-cell level. We encapsulated individual sea urchin sperm with demembranated flagellum inside water-in-oil emulsion droplets and measured the axoneme's ATP consumption by monitoring fluorescence intensity of a fluorophore-coupled reporter system for ATP turnover in the droplet. Concomitant phase contrast imaging allowed us to extract a linear dependence between the ATP consumption rate and the flagellar beating frequency, with ∼2.3 × 10(5) ATP molecules consumed per beat of a demembranated flagellum. Increasing the viscosity of the aqueous medium led to modified beating waveforms of the axonemes and to higher energy consumption per beat cycle. Our single-cell experimental platform provides both new insights, to our knowledge, into the beating mechanism of flagella and a powerful tool for future studies.
authors.affiliation: Martin Fisher School of Physics, Brandeis University, Waltham, Massachusetts.
authors.forename: Daniel T N
authors.initials: DTN
authors.lastname: Chen
authors.affiliation: Martin Fisher School of Physics, Brandeis University, Waltham, Massachusetts; Graduate Program in Biophysics and Structural Biology, Brandeis University, Waltham, Massachusetts.
authors.forename: Michael
authors.initials: M
authors.lastname: Heymann
authors.affiliation: Martin Fisher School of Physics, Brandeis University, Waltham, Massachusetts.
authors.forename: Seth
authors.initials: S
authors.lastname: Fraden
authors.affiliation: Department of Biology, Brandeis University, Waltham, Massachusetts. Electronic address: nicastro@brandeis.edu.
authors.forename: Daniela
authors.initials: D
authors.lastname: Nicastro
authors.affiliation: Martin Fisher School of Physics, Brandeis University, Waltham, Massachusetts. Electronic address: zdogic@brandeis.edu.
authors.forename: Zvonimir
authors.initials: Z
authors.lastname: Dogic
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D054468:Axoneme / Q000378:metabolism; D004398:Dyneins / Q000378:metabolism; D005407:Flagella / Q000378:metabolism*; D007700:Kinetics; D047331:Lytechinus / Q000166:cytology*; D008297:Male; D009068:Movement; D059010:Single-Cell Analysis; D013094:Spermatozoa / Q000166:cytology
chemical_list: D000255:Adenosine Triphosphate; D004398:Dyneins