Date of Award

Fall 2011

Document Type


Degree Name

Doctor of Philosophy (PhD)


Biological Sciences

First Advisor

Yang, Pinfen

Second Advisor

Karrer, Kathleen

Third Advisor

Downs, Stephen


The goal of this thesis is to elucidate the assembly mechanism of macromolecular complexes in the microtubule-based axoneme in flagella. The axoneme is comprised of more than 400 distinct polypeptides. While all axonemal proteins are synthesized in the cell body, they need to be imported to the tip of the growing flagella for the final assembly. They are then assembled into distinctive structural complexes and integrated into precise positions relative to each other. The assembled macromolecular complexes operate in concert enabling the axoneme to beat rhythmically as a biological machine. This arduous process involves multiple reactions and meticulous regulation. However, the assembly process in the cell body and at the tip of flagella remains largely unknown.

This thesis took advantage of a flagella model organism Chlamydomonas and a rather simple and a well-characterized axonemal complex, the radial spoke, to identify the key assembly mechanisms. Genetic studies have revealed two spoke components, RSP3 and LC8, that are particularly important for the assembly of the entire complex. Mutants defective in either gene generate paralyzed flagella devoid of the RS. Using a variety of approaches that are possible for this model organism, this study revealed that a stack of LC8 dimers directly binds to RSP3 at the tip of the flagella to promote a series of events, including phosphorylation of RSP3, formation of the base of the RS and docking of the spoke to the axoneme.

These findings have shed light on the series of events that occur between transport and assembly. These findings also have broad implications, as LC8 is a promiscuous molecule that interacts with many flagellar and non-flagellar proteins in the cell body. Many of them play vital roles in the normal functioning of cells or in the pathogenesis of microorganisms. Some LC8 target proteins exist in the molecular complexes, as the radial spoke. However, the LC8-involved reactions remain poorly defined. The findings from this thesis show that LC8 bindings can trigger pleiotropic effects on a single target protein. Together, these conclusions shed critical insight on the biology of flagella and LC8.

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