Abstract
Primary cilia are composed of axonemal microtubules that extend from the mother centriole-derived basal body and are sheathed by the ciliary membrane. Distal appendages (DAP) of the mother centriole play crucial roles as a scaffold to initiate ciliogenesis. Although previous studies indicated that the DAP proteins CEP164 and Tau-tubulin kinase 2 (TTBK2) participate in key events of ciliogenesis, including removal of CP110 from the mother centriole and recruitment of the intraflagellar transport (IFT) machinery to the mother centriole, the overall process involving these DAP proteins remains unclear. We here established CEP164-knockout (KO) and TTBK2-KO cells, and expressed various CEP164 and TTBK2 constructs in these cells. Our results showed that the interaction of TTBK2 with CEP164 and TTBK2 kinase activity is required for the recruitment of IFT machinery components (IFT-A, IFT-B, and dynein-2 complexes) to, and removal of CP110 from the mother centriole. However, CP110 removal is not always coupled with IFT protein recruitment. Analysis using chimeric constructs of CEP164 and TTBK2 indicated that CEP164 homodimerization via its central coiled-coil region is necessary for its mother centriole localization and subsequent TTBK2 recruitment, which are required for the recruitment of IFT machinery components to the mother centriole to trigger ciliogenesis.