A wide range of eukaryotic organisms generate motile cilia and flagella. These slender organelles beat rhythmically to move the surrounding fluid or to propel cells in aqueous environment. Organisms use these powerful yet nimble organelles to forage, evade, adapt and mate. The machinery that drives this tightly controlled movement is the sophisticated microtubule-based axoneme. As it is critical for the survival of individual species, this machinery has largely been preserved to the molecular level throughout evolution. Proteomic studies have shown that most proteins in this biological machine consist of molecular modules commonly used in the cell body. But the usage of these modules is clearly diverged in many cases. This defined machinery with diverged applications provides an opportunity to understand the true capacity of the conserved modules. One example is the radial spoke (RS) that controls the oscillatory beating. This macromolecular complex contains complementary molecular modules that are responsible for localizing cAMP-dependent protein kinase (PKA) in the cell body. However, the RS does not have the features that account for the effector mechanisms of PKA and thus the mechanism discovered for localizing PKA has a broader role that has previously not been recognized. The work described in this dissertation discovered that the core of the RS utilizes two similar sets of PKA anchoring modules for four distinct effector mechanisms that underlie the assembly and function of this regulatory complex. These results elucidate the function of this complex and are applicable to more than 600 diverged proteins that also share the docking module of PKA. Some of them have been shown to play vital roles in myriads of cellular reactions ranging from flagellar beating to trans-Golgi trafficking to chromosome modifications. Founded on this discovery, new reagents and assays were engineered. These tools could be used for the exploration of proteins with similar docking and anchoring modules. Together, these findings will accelerate the advancements in the field of anchoring and docking of proteins in the cell
