I. Control of Microtubule Sliding in Cilia We will directly test the current model of how the force-generating machinery in cilia is controlled to produce bending motion. By using a uniquely advantageous system - macrocilia from the ctenophore Beroe - we will apply the ATP-induced sliding disintegration method of Summers and Gibbons (1971), together with cinemicrography and electron microscopy, to see if bending is caused by alternate activation and inactivation of dynein arms on opposite sides of the axoneme. Because ciliary and flagellar motion is responsible for fluid and particle transport in human respiratory and reproductive systems, as well as for sperm motility, the project is strongly related to certain health problems. II. Electrical Control of Ciliary Motor Responses in Metazoans The activity of cilia and flagella can be modified by the organism to make adaptive responses to environmental stimuli. We will use newly-discovered ciliary motor responses of ctenophore comb plates - a system allowing clear visualization of ciliary beating and cellular electrophysiology - to investigate the ionic and nervous control of ciliary motion in metazoans. Extracellular stimulation of ciliary responses, intracellular recordings from comb plate cells, and current injection will be combined with high-speed video microscopy to give simulataneous recording of electromechanical coupling. This project complements Part I by providing information on the bioelectric control of ciliary motor states in metazoans. III. Mechanism of an Eukaryotic Rotary Motor We will investigate the mechanism of a unique type of motility in an eukaryotic cell: continual, unidirection rotation of one part relative to the other, driven by a rotary axostyle. We will determine whether the rotary motor operates by a conventional actomyosin mechanism, but with a circular geometry of interacting elements. Recently devised ATP-reactivated models of the rotary axostyle will be modified to permit access of specific macromolecular probes. These inhibitors will also be microinjected into living cells and ATP-models. Identity and polarity of presumed actin filaments will be tested by electron microscopic cytochemistry. The protein composition of the axostyle will be analyzed by biochemical methods. This project will enlarge our understanding of the types of motility and filament geometry possible based on actomyosin mechanochemistry.
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