The BAR domain-containing protein ciBAR1 requires Cby1 for efficient binding to phosphoinositide-enriched membranes...

Bin/Amphiphysin/Rvs (BAR) domain-containing proteins participate in membrane-remodeling processes, including vesicle trafficking, cell division, and cell migration. We previously demonstrated that ciBAR1 interacts with Cby1 and colocalizes at the ciliary base to promote ciliogenesis. However, the function of the Cby1/ciBAR1 complex remains poorly understood. Here, using recombinant proteins and reconstituted liposomes, we show that ciBAR1 alone exhibits little or no lipid-binding activity, whereas Cby1 promotes ciBAR1 binding to liposomes. In nanobar assays, Cby1 also promotes ciBAR1 membrane association and enrichment at regions of high membrane curvature. The Cby1/ciBAR1 complex preferentially associates with liposomes containing PI(4,5)P₂ and PI(3,4,5)P₃, two phosphoinositides (PIs) enriched near the ciliary base. Consistent with these findings, ciBAR1 mutants defective in either membrane or Cby1 binding fail to localize to the ciliary base and are unable to rescue ciliogenesis defects in ciBAR1-KO NIH3T3 cells. Furthermore, ultrastructural analysis reveals that ciBAR1-KO cells exhibit defects in preciliary vesicle recruitment and an accumulation at the ciliary vesicle stage, suggesting impaired progression to ciliary membrane extension and ciliary bud formation. Collectively, our findings support a model in which Cby1 potentiates ciBAR1 binding to PI-enriched membranes at the basal body, where the Cby1/ciBAR1 complex mediates membrane-associated processes during early ciliogenesis.