Title

Assembly, structure and regulation of dyneins in cilia

Date of Completion

January 2009

Keywords

Biology, Molecular|Biology, Cell|Chemistry, Biochemistry

Degree

Ph.D.

Abstract

Cilia are conserved cellular organelles with important roles in human physiology. Dyneins are a family of minus-end directed, microtubule motors that are crucial for cilia function. For example, an isoform of cytoplasmic dynein drives the retrograde intraflagellar transport (IFT); a process required for cilia assembly and maintenance. In addition, a diverse class of dynein motors, namely the outer and inner dynein arms, associate with the axoneme making the entire structure motile. In this study we investigated the role of the lissencephaly protein (Lis1) on the regulation of the outer arm dynein complex. We showed that Lis1 has a conserved function in motile cilia and that a Chlamydomonas ortholog (CrLis1) interacts with the a heavy chain (or the associated LC5 subunit) of the outer arm complex. We also provided evidence to suggest that the association of CrLis1 with the outer arm is regulated by other axonemal subsystems, including the central pair apparatus, radial spokes and inner arm system, as part of singling pathway that controls outer arm motor activity. Light chain subunits that belong to the family of LC8/DYNLL, are also important for outer arm motor function. Here, we have characterized a conserved third member of this family of light chains and showed that each of these three subunits has a distinct role in the assembly and regulation of the outer arm motor. In this study we also investigated the architecture of the dynein motor that powers retrograde IFT; a motor complex whose organization, structural composition and regulation is poorly understood. We showed that this motor consists of a heavy chain dimer, and other conserved subunits including a novel intermediate chain (FAP133) that we identified and characterized. FAP133 is an integral component of the IFT dynein and directly binds LC8, forming a complex that is essential for the loading of the dynein complex onto the anterograde transport system or onto other IFT cargoes. Finally, we explored the use of Schmidtea mediterranea as a biological system to study cilia and cilia-based motility, and demontrated that ciliary motility is required for planarian gliding locomotion. ^