| The radial spoke (RS) is a signature feature in the 9+2 axonemes in most motile cilia and flagella. It is critical for synchronized oscillatory beating and the regulation of dynein motors. Yet, only a few subunits are apparently conserved, including a homodimeric RSP3, one of the A-kinase anchoring proteins (AKAPs) that bind mammalian RII in vitro. The binding in theory entails RIIa, the dimerization and docking (D/D) domain in RII subunit of cAMP-dependent protein kinase (PKA) and an amphipathic helix in AKAPs. Intriguingly, RS contains four non-PKA subunits with a D/D domain: two with RIIa while the others with a homologous DPY-30. Like many of the other 400 members in the RIIa clan superfamily, these four RS subunits contain additional diverged sequences that are inconspicuous or related to nucleotide metabolism and calcium/calmodulin signaling. Mutants defective in these RIIa clan molecules display complete or reversible paralysis or abnormal calcium-dependent steering. Using both in vivo and in vitro approaches, we discovered that in fact RIIa and DPY-30 domains bind to two distinct regions in RSP3. Interestingly, only DPY-30 domain showed cross recognition in vitro, suggesting that the targeting principles of these two D/D domains are similar but additional cues differentiate the specificity in vivo. Based on the biochemical deficiencies, we postulate that RSP3 forms a dimeric structural scaffold anchoring to particular locations the two types of D/D domains that in turn tether four different effector modules, each conferring to RS unique regulatory mechanisms. These findings reveal the extraordinarily versatile usage of D/D in eukaryotes. |