Abstract
Axonemal dynein is a microtubule-based molecular motor that drives ciliary/flagellar beating in eukaryotes. In axonemal dynein, the outer-arm dynein (OAD) complex, which comprises three heavy chains (α, β, and γ), produces the main driving force for ciliary/flagellar motility. It has recently been shown that axonemal dynein light chain-1 (LC1) binds to the microtubule-binding domain (MTBD) of OADγ, leading to a decrease in its microtubule-binding affinity. However, it remains unclear how LC1 interacts with the MTBD and controls the microtubule-binding affinity of OADγ. Here, we have used X-ray crystallography and pulldown assays to examine the interaction between LC1 and the MTBD, identifying two important sites of interaction in the MTBD. Solving the LC1-MTBD complex from Chlamydomonas reinhardtii at 1.7 Å resolution, we observed that one site is located in the H5 helix and that the other is located in the flap region that is unique to some axonemal dynein MTBDs. Mutational analysis of key residues in these sites indicated that the H5 helix is the main LC1-binding site. We modeled the ternary structure of the LC1-MTBD complex bound to microtubules based on the known dynein-microtubule complex. This enabled us to propose a structural basis for both formations of the ternary LC1-MTBD-microtubule complex and LC1-mediated tuning of MTBD binding to the microtubule, suggesting a molecular model for how axonemal dynein senses the curvature of the axoneme and tunes ciliary/flagellar beating.
Highlights
Axonemal dynein is a microtubule-based molecular motor that drives ciliary/flagellar beating in eukaryotes
Solving the light chain-1 (LC1)-microtubulebinding domain (MTBD) complex from Chlamydomonas reinhardtii at 1.7 Å resolution, we observed that one site is located in the H5 helix and that the other is located in the flap region that is unique to some axonemal dynein MTBDs
This enabled us to propose a structural basis for both formations of the ternary LC1-MTBD-microtubule complex and LC1-mediated tuning of MTBD binding to the microtubule, suggesting a molecular model for how axonemal dynein senses the curvature of the axoneme and tunes ciliary/flagellar beating
Summary
The complex of outer-arm dynein light chain-1 and the microtubule-binding domain of the ␥ heavy chain shows how axonemal dynein tunes ciliary beating. It has recently been shown that axonemal dynein light chain-1 (LC1) binds to the microtubulebinding domain (MTBD) of OAD␥, leading to a decrease in its microtubule-binding affinity. Because the ATPase activity of the heavy chain is increased in the presence of microtubules [10], the above-mentioned physiological and biochemical analyses imply that LC1 might change the ATPase activity of the AAAϩ ring of OAD␥ from the remote MTBD region and thereby fine-tune ciliary/ flagellar beating. Our findings enable us to provide the first structural model of the LC1 accessory function, showing how LC1 is able to fine-tune the ATPase activity of the heavy chain without any direct interaction with the AAAϩ ring or microtubule, because of its tight binding to the MTBD. Our structural model addresses the possible functional role of the flap region that is directly bound to LC1, leading to a unique stepping model of OAD␥ on the microtubule based on tethering and release of the flap region by LC1
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