JHY enables the transition from switchable to fixed ciliary waveforms in metazoan evolution.

Motile cilia are evolutionarily conserved protrusions critical for motility and homeostasis. Their rhythmic movements require the central pair microtubules (CP-MTs). While the initial CP-MT assembly in mammals is mediated by WDR47 and microtubule minus-end-binding CAMSAPs, the mechanism by which CP-MTs are stabilized remains unclear. Here, we demonstrate that WDR47 coordinates JHY and SPEF1 to maintain the stability of mammalian CP-MTs. By generating a proximity interactome of WDR47, we identify a group of CP-MT-associated proteins, including SPEF1 and JHY. WDR47 enriches JHY and SPEF1 to the central lumen and tip of nascent cilia, whereas SPEF1 recruits WDR47 and JHY to CP-MTs through direct interactions. Jhy deficiency in mice preferentially disrupts distal CP-MTs, resulting in rotatory ciliary beats. Phylogenetic analyses suggest conserved functions of WDR47 and SPEF1 in protozoa and metazoans, as well as a role for JHY in animals with radial or bilateral body symmetry. We propose that JHY emerges to further reinforce CP-MTs, enabling the transition from switchable to fixed ciliary waveforms in metazoan evolution.