The microtubule-associated proteins CKAP2 and its paralog CKAP2- Like control ciliogenesis in human cells

Abstract

Ciliogenesis is an evolutionarily conserved process that leads to the assembly of cilia. This process relies on microtubule-associated proteins (MAPs) to regulate axonemal microtubule dynamics. Misregulation of MAPs often leads to changes in ciliary homeostasis, contributing to numerous ciliopathies. Although CKAP2 and CKAP2-Like are best known for regulating microtubule dynamics during cell division, they are also components of ciliary organelles. Notably, CKAP2 overexpression promotes chromosomal instability and cancer, whereas CKAP2-Like deficiency causes Filippi syndrome, a developmental disorder with clinical features overlapping those of ciliopathies. How both MAPs operate at primary cilia remains poorly understood. Here, we identify both MAPs as axonemal components of motile and primary cilia in human cells. We find that CKAP2-positive cilia are linked to cell cycle progression and that the conserved intrinsically disordered microtubule- binding domain of CKAP2, which is essential for microtubule polymerization and stabilization, is required for its ciliary localization. Unexpectedly, CRISPR-mediated loss of either paralog results in elongated cilia without affecting ciliogenesis, accompanied by increased ciliary enrichment of the remaining paralog and impaired Gli2 accumulation at the ciliary tip. In contrast, simultaneous deletion of both MAPs suppresses ciliogenesis and has minimal effect on the ciliary length as compared to wild-type levels. These findings uncover that CKAP2 and CKAP2-Like are axonemal MAPs that contribute to ciliogenesis, partially compensating for each other in human cells.