STRUCTURE AND MOTION OF CILIA

Cilia are important motile cell organelles of organisms including man.
This study continues a comprehensive attack on the structure and function
of cilia, centering on ultrastructural correlates of ciliary motion,
including analysis of the mechanism of movement and its regulation. This
approach has been influential in the development of the sliding microtubule
model of ciliary motility, now widely accepted. Despite the success of
this model, fundamental questions remain regarding the basic interaction
responsible for sliding and the hierarchy of regulatory processes between
the sliding event and actual ciliary beat. One major aim of this proposal
is to strengthen the hypothesis that a unique cycle of dynein arm activity
is responsible for microtubule sliding in motile cilia, and to specify the
structural bases of this cycle further. The subunit structure of the arm
will be studied using negative stain, freeze etch and rotary shadow
electron microscopy. Structure will be correlated with polypeptide
composition, location of microtubule attachment points and ATPase
activity. A second aims is to use such structural information to probe
possible cooperative and/or asynchronous arm activity in the axoneme.
Asynchronous arm activity is embodied in several axonemal switching
hypotheses relating sliding of specific microtubules to defined beat
positions. These hypotheses will be tested by direct readout of arm
configuration along single doublets and from doublet to doublet in cilia
treated with various ATP analogs or arrested in specific stroke positions,
for example after treatment with Ca2+ or calmodulin (CaM)-directed drugs.
The possibility that CaM or CaM-binding proteins are part of the dynein arm
in some cilia will be explored. This information should lead to further
understanding of normal ciliary activity and of ciliary malfunction in
respiratory disease.