The primary cilium,an important microtubule-based organelle,protrudes from nearly all the vertebrate cells.The motility of cilia is necessary for various developmental and physiological processes.Phos-phoinositides(Pls)and its metabolite,Ptdlns(4,5)P2,have been revealed to contribute to cilia assembly and disassembly.As an important kinase of the PI pathway and signaling,phosphatidylinositol 4-kinase β(PI4KB)is the one of the most extensively studied phosphatidylinositol 4-kinase isoform.However,its potential roles in organ development remain to be characterized.To investigate the developmental role of Pi4kb,especially its function on zebrafish ciliogenesis,we generated pi4kb deletion mutants using clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9 tech-nique.The homozygous pi4kb mutants exhibit an absence of primary cilia in the inner ear,neuromasts,and pronephric ducts accompanied by severe edema in the eyes and other organs.Moreover,smaller otic vesicle,malformed semicircular canals,and the insensitivity on sound stimulation were characteristics of pi4kb mutants.At the protein level,both in vivo and in vitro analyses revealed that synthesis of Pi4p was greatly reduced owing to the loss of Pi4kb.In addition,the expression of the Pi4kb-binding partner of neuronal calcium sensor-1,as well as the phosphorylation of phosphatidylinositol-4-phosphate down-stream effecter of Akt,was significantly inhibited in pi4kb mutants.Taken together,our work uncovers a novel role of Pi4kb in zebrafish inner ear development and the functional formation of hearing ability by determining hair cell ciliogenesis.