Primary cilia are microtubule-based organelles projectingfrom the surface of nearly all cells. Primary cilia are complexorganelles and are structurally divided longitudinallyinto sub-compartments that include the basal body, thetransitional zone, the ciliary shaft and the tip. Nephronophthisis(NPHP) is an autosomal recessive cystic kidneydisease with 11 responsible genes (NPHP1-11) thus farbeing identified. The causative gene products, nephrocystins,are divided into at least two groups based on theselocalizations. Group I nephrocystins (Nphp1, 4, 5, 6 and 8)are localized in the transitional zone, whereas group IInephrocystins are localized in the Inv compartment(Nphp2, 3 and 9). Here, we show the localization mechanismof Nphp3. We generated a series of GFP-tagged deletionconstructs of Nphp3 and tried to find the ciliarytargeting sequences of Nphp3. We found that the N-terminalfragments, Nphp3 (8–201), Nphp3 (52–201) andNphp3 (96–201), that contain the CC domains, targetedthe basal body, but could not enter into the ciliary shaft.Further analysis revealed that an N-terminal glycine (G2),which is a conserved myristoylation site among vertebrates,is also essential for trafficking of Nphp3 to the ciliaryshaft. We revealed that Inv/Nphp2 is not required forentry of Nphp3 into the ciliary shaft. Following entry ofNphp3 into the ciliary shaft, Inv/Nphp2 is required for thelocalization of Nphp3 to “the Inv compartment”. Ourresults showed the importance of myristoylation in ciliarytrafficking, and suggest that Nphp3 ciliary localizationoccurs in a three-step process.
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