Doublet microtubule inner junction protein FAP20 recruits tubulin to the microtubule lattice

Abstract

Motility of organisms involves beating of cilia and flagella that are composed of doublet microtubules. The doublets are made of A- and B-tubules that fuse together at two junctions. Among these, the outer junction is made of tripartite tubulin connections, while the inner junction contains distinct non-tubulin elements. The latter includes Flagellar-associated protein 20 (FAP20) and Parkin co-regulated gene protein (PACRG) that together link the A- and B- tubules at the inner junction. While the structures of doublet microtubules reveal molecular details, their assembly is poorly understood and examining proteins at the junctions can provide important clues. In this study, we purified recombinant FAP20 and characterized its effects on microtubule dynamics by TIRF microscopy. Using in vitro reconstitution and cryo-electron microscopy, we conclusively show that FAP20 recruits free tubulin to the existing microtubule, where it mediates flexible lateral interactions between the microtubule lattice and tubulin dimers. Our structure of microtubule:FAP20:tubulin complex partially resembles the inner junction architecture, further providing insights into assembly steps involved in closure of B- tubule in a doublet microtubule.